The Oakland School District needs to learn how to teach proper English, not how to coddle inappropriate speech in the classroom
Sounds like a quoted source or personal opinion?
- Last 7 days
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go.gale.com go.gale.com
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The real objective for ebonics, like Afrocentric education, is to immobilize and often to discredit the public school systems efforts to teach the standard English.
Afrocentric education agenda?
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The failure to recognize and re-teach Ebonics users may cuase a lifetime of struggle due to language barriers.
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making the child unfit for later career development in a post modern information society
Inability to essentially "re-learn" the standard English language may hinder the development of the individual. Esepcially deeper into college and career life should re-education fail to take place.
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It matters little if the inappropriate speech pattern derives from poverty, rap music, hip hop culture, a dysfunctional ghetto family, or historical speech variations from Africa or the plantation
A wide variety of factors contribute to the inception of Ebonics.
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are appropriately thought to have language disorders and often with learning disabilities
Is there a correlation between the percentage of children that live in poverty and the percentage of children that are considered to have learning disabilities?
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Source reporting on the controversy of Ebonics and the role it plays on the youth.
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show disrespect and to mislead the child and an entire race of African-American people
Continued conformity to slang in the classroom may present the child as incapable of improvement or proper communication
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some attempts to legitimize such poor language habits as being a culturally legitimate African or English language do more harm than good to students
The false varification of this slang fails to align with either traditional African or English language values, thus rendering it useless.
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It involves information economy where standard English determines one's career success and failure. Let us face it, ebonics is a fancy political cover for abnormal, defective, or dysfunctional speech
Guiding claim/overshadowing argument.
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laughing-bassoon-7p6pm2g.pages.github.io laughing-bassoon-7p6pm2g.pages.github.io
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Consent screen
How this screen is display? Client should identify the user came from MCP context and redirect it after the login process is ended successfully
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make the issued token something the platform already trusts and that we can revoke in a single call — instead of a signed token that would need new signing infrastructure.
Need to make sure Kong identified it as apikey for manage the rate limit of the account.
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Revoke = delete the credentialOne Admin-SDK call kills a connection — no blocklist, no waiting for a signature to expire.
Where this option is located?
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What stays identical: the proxy still forwards ?apikey=; Kong still validates a key with the same plugin; api.tomorrow.io is untouched; existing x-api-key callers (server-to-server, n8n) keep working unchanged.
Does it return the existing API key to Claude? You should provide another secret key. It's not clear to me who converts this key to the API key.
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stunlaw.blogspot.com stunlaw.blogspot.com
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cusp of a transformation
I think here is the goal! The governments want to end the struggle with the universities by controlling them with AI. They want to narrow the gap when they assumed the technology and AI are the same path to use in education. It is truth the technology has helped us for many years, but AI takes our place as educators and make the fake thoughts as the facts but more shiny.
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The Dangers of an Algorithmic Idea of the University
It's interesting fact that when we students we find AI as a kind angel work for us as we want him to do our assignments with grade A . However, When we be teachers we do not want our student or our kids use it because they will lose their ideas and thoughts plus their imagination to creative thoughts!.
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research.ebsco.com research.ebsco.com
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The reliance on ChatGPT is more concerning when students use it for writing assignments, as dependence could impair the development of critical thinking skills and originality (Azaria et al., [ 2 ]; Rane et al., [37]). Azaria et al. stressed the tool's limitations in logical reasoning and the risk of generating inaccurate responses (i.e., hallucinating), which may lead to overconfidence in nonexpert users.
Reliance of AI is what brings the concerns and because of the imperfection of AI it causes Hallucinations. Overconfidence in non-educated users who think AI is perfect has a lot to do with Hallucinations from AI.
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Understanding student perceptions toward ChatGPT is crucial for navigating its implications for education, especially because students have generally embraced ChatGPT for its ability to facilitate various aspects of learning (Adeshola & Adepoju, [ 1 ]).
Understanding how AI is seen is necessary to know how to implement AI in education.
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The integration of ChatGPT into education has sparked a dynamic dialog among researchers, educators, and students. ChatGPT's application within educational settings heralds a significant shift toward personalized learning, offering a range of opportunities to enhance both teaching and student engagement (Kasneci et al., [21]). Adeshola and Adepoju ([ 1 ]) highlighted the technology's capacity for content generation and natural language processing, which can serve as powerful tools in the development of learning materials and the facilitation of a more interactive learning experience.
Impact of AI on researchers, educators, and students.
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To address this issue, the present study seeks to determine whether active discussions—a form of participatory learning—about potential ChatGPT concerns among students can help reduce uncritical positivity toward ChatGPT, such as usefulness and favorability, that will eventually reduce the overreliance on it among college students.
Developing ways to counteract the issues forming because of AI (GPT). Over time the issues will be reduced (reliance on AI).
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Furthermore, the overreliance on ChatGPT for academic writing raises questions about the impact on students' development of critical thinking and writing skills—essential competencies in higher education and beyond (Adeshola & Adepoju, [ 1 ]; Lo, [32]). This problem is becoming more serious as the use of AI writing tools among college students is drastically increasing, with nearly half of students reportedly using AI for writing, according to survey data from Fall 2023 (Shaw et al., [41])
Issues arising due to the influence of AI towards students. This is impacting student's critical thinking and writing skills.
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students use AI applications to generate content ideas, get help in language editing, and even conduct literature reviews (Rezaei et al., [38]). Using AI for academic tasks has opened new avenues for learning and exploration, suggesting a potential paradigm shift in educational methodologies.
What AI has been used for and how it has opened up a new view of education for students and teachers.
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The integration of ChatGPT into educational settings has sparked a significant shift in how learning is approached, offering both promising opportunities and unique challenges that warrant a closer examination.
What AI (GPT) is changing in the educational landscape, and how it benefits us.
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Although artificial intelligence (AI) tools (e.g., ChatGPT) have potential to act as a transformative educational tool, the widespread adoption of such technology also brings forth concerns related to academic integrity and learning gaps.
Things meant to be used for good can also have its risks and concerns. Some concerns like "academic integrity and leaning gaps."
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www.jstor.org www.jstor.org
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are many culturally significant ways of using languages, such as speaking, reading, writing,and listening. Therefore, if a language does become dormant following a period of endan-germent or a significantly impactful event, languages can still be culturally meaningfulin their ‘afterlife’, even if no longer used in everyday spoken communication (i.e., Latin,Classical Greek, and Hebrew). While not the desired outcome of linguists and Indigenouscommunities, without intense intervention this may be the future of Indigenous languageswithin North America and worldwide.
I like this section because it describes how even though these languages might not be spoken everyday or in day to day life it still can be incredibly helpful to know these languages to understand the history of these cultures.
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Knowledge about geography is transmitted from generation togeneration and is extremely important to the way of life of many Indigenous commu-nities. For example, since many Indigenous communities moved seasonally to differenthunting and fishing grounds, they relied heavily on their lived experiences and storiespassed down through generations to navigate the landscape. In addition, the words usedto describe the landscape and specific geographical markers demonstrate a connection,further illustrating the correlation language has to cultural practices and geography.5
This shows the connection between geography and how the Inuit traveled with the language itself as the language could describe how to get to certain hunting ground and settlements through the year. Without their language they'll lose their ability to travel as their ancestors did.
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On the other hand, although the English language can describe these snow types,there are only a handful of words for snow or similar weather phenomena; however, withinInuktitut, there are dozens. This directly highlights that, given their environment, the Inuitdeemed that the varieties of snow and ice were things they needed to differentiate andtalk about, whereas this distinction is not so relevant to English culture. Ice and snowwithin their community influence how people “choose sledge routes, find water or snowfor drinking, select places where they can make a house, consider which surfaces are safeto walk or sledge across,”47 and more.
This is a great example of how Language directly reflects the environment and the culture that was being built by Inuit. The environment was snow filled so they saw and spoke the world through snow.
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Specifically, they construct the “Whiteman” using jokes in English.Although “it is impossible to determine exactly when Apache’s living at Cibecue firstemployed English to perform joking imitations of Anglo-Americans,”50 Basso explainsthat jokes in English were a new phenomenon when writing his book, first exclusively usedby young children and then by adults as Anglo-Americans became recurrent characters intheir lives.51 Basso also asserts that individuals within the Western Apache communityonly make jokes in English around people that they are close with because jokes can bemisunderstood or offensive. This adverse reaction can destroy relationships unless theyare strong or long-standing. This example helps to reinforce the importance of languagerevitalization in relation to cultural identity. The Western Apache use joking in Englishto construct the white settlers with whom they interact and this joking became ingrainedwithin their cultural practices, resulting in bonding within the community and conse-quences if employed in the wrong setting or with the wrong people.52
This i believe reinforces the thought that language can be miss interpreted if not properly understood by the people who are speaking said language.
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How communities feel about their language can greatly impact whether theywish to continue speaking it. However, language suppression was one of the ways colo-nial powers attempted to assimilate Indigenous communities into Western society. Thishas led to an overwhelming perceived superiority of dominant languages within society,which along with many other factors, have inspired the decline of Indigenous languageswithin North America and worldwide.40 These linguistic ideologies continued to evolvewithin Indigenous communities because of often unspoken and unconscious trauma andintergenerational trauma. These beliefs are ‘naturalized’ or ‘normalized’ within societies,leading to the eventual endangerment or dormancy of languages.
I like this section of text because I can most likely use this as a connection between my own up brining and the suppression of my own families native language in favor for standard English due to embarrassment of our native ancestry.
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the Sapir-Whorf hypothesis of linguistic relativity or linguistic determinism. In its weaker form(linguistic relativity), this hypothesis “asserts a correlation between language and thinkingeither with regard to the specifics, the structure, the categories, or some other generalitiesof each.”42 Then, in its stronger form (linguistic determinism), this hypothesis postulatesthat “in some way, the language of any given culture is the causal determinant of thepatterns of thinking in that culture.”43 The hypothesis argues that the language we speakinfluences how we, as speakers, perceive and conceptualize the world.44In addition to the case studies cited herein, this hypothesis can also be applied to theInuit who live in what is known as Northern Canada, the United States, and Greenland.45For example, in Inuktitut, a language spoken by the Inuit, there are many words forsnow/ice. This can influence how speakers conceptualize reality or are predisposed to
I just like this section because it explains why language changes our world perspective and how languages in term reflect the society that was built on said language.
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Additionally, as indicated in the three case studies, all these communities are focusingtheir efforts on children. It can be argued that this is important; as, according to thecritical period hypothesis, there is a limited period of time in childhood development
This shows why the focus is on children to learn their native tongue over teaching adults how to speak the native language.
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Although on the brink of dormancy, the Lakota Nation is actively working to revital-ize their language with a strong emphasis within their community on teaching childrenthe language. They have implemented immersion language nests, including daycare fortoddlers33 and immersion elementary schools.34 In both these environments studentslearn all the same concepts as in traditional schools, but classes are taught in Lakota
How the Lakota people are trying to revitalize their languages.
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Oftenreferred to as the Great Sioux Nation, the Lakota people can be divided into three distinctgroups based on their language and geography: Dakota (Santee, Eastern Dakota), Lakota(Teton, Western Dakota) and Nakota (Yankton, Central Dakota).”
Specific Lakota groups in North and South Dakota.
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Additionally, many of the film’sspeakers highlight how influential language is to their community, arguing that languageloss has contributed to issues of drugs and alcohol within their reservation
Additional information on how the loss of language affects the native culture and society for the worse.
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y eradicating Indigenous languagesin children in an attempt to Americanize Indigenous populations, the government waseffectively cutting off the cultural pathway between these children, their ancestors, andfuture generations.
This is a great quote to use to explain why the eradication of native languages does harm as it severs their cultural pathways.
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One program is a Cherokeeschool where children are educated in all the same areas they would be in a traditionalschool, but classes are taught in Cherokee
This is my favorite of all the programs so far. A school that teaches everything that they usually need to learn but in Cherokee. It intertwines modern learning with the deep history of their language, how cool.
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nother initiative is the language documentation program where L1 and L2 speakersare recorded using their language as a means of building resources for language documen-tation and acquisition.21 Language documentation programs like this one are valuable asthey create resources that “serve to link language users to past practices, affirm identityand support language revitalization and reclamation.
This is another program that helps young generations learn their native language.
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For example,they created the Tahltan Language Revitalization Program, a working group inspired bythe desire to restore interest in the language, whose mandate is to create programs thatencourage language acquisition and maintenance. One of the programs developed by thisgroup is called language nests. Language nests are daycare programs in which babiesand toddlers are immersed in the language to encourage L1 and L2 language acquisi-tion.19
I feel this is important as this is somewhat the formula to bring back native language speakers, by bringing programs that help younger kids learn their native tongue.
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this paper focuses onthe Tahltan Nation of what is now known as northwestern British Columbia, the CherokeeNation of what is now known as Oklahoma, and the Lakota Nation of what is now knownas North and South Dakota.
Thse are the people that this Journal will be discussing in terms of their language revitalization.
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One of the catalysts sparked by colonization and globalizationthat impacted the vitality of Indigenous languages in what is now known as Canada andthe United States is the residential ‘school’ system. Through these programs, Indigenouschildren were taken away from their homelands and punished for using their language(s),among many other things, resulting in intergenerational trauma, causing a direct declinein language use.6
This represents how originally natives were thrown into school systems that taught them to forget their native language in favor of a universal proper English. This is how the government was able to control native language and force them into a new one.
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The United Nationspresents that “out of the 6,700 languages spoken worldwide, forty percent are in danger ofdisappearing [and of that] most of the languages that are under threat are Indigenous lan-guages.”5
Wow I'm surprised not only by the seer number of languages but how many actually are indangered of being lost forever. That's generations of history that we be lost with the languages.
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For many people, theirancestral language is integral to cultural expression and continuity.”2 Therefore, whenlanguages become endangered or dormant, this directly impacts culture and culturalpractices. This is specifically true for Indigenous cultural practices since traditions andcustoms are passed down orally from one generation to the next.3 Consequently, if theselanguage(s) were to become endangered or dormant, not only would Indigenous peopleslose their language, but they also risk losing the cultural knowledge that is deep-seated inthe language.
This highlights the true reason behind the want to revitalize indigenous languages as a loss of language for those people directly correlates to the loss of culture and history since it's very much intertwined.
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Historically (and currently) Indigenous languages have been suppressed and marginalized within so-ciety, inspiring declining levels of language usage and L1 and L2 speakers. It can be argued that thesedeclining levels of speakership have impacted the tangible and intangible elements of Indigenous culturalpractices. Although Indigenous peoples have faced punishment for using their languages, the reclama-tion or revitalization of Indigenous languages can lead to the recovery of cultural knowledge and, in theprocess, help heal the trauma caused by colonization. This article seeks to address the impact languagerevitalization efforts can have on maintaining the cultural practices of Indigenous communities byexamining three case studies of ongoing revitalization efforts: the Tahltan Nation, the Cherokee Nation,and the Lakota Nation. Moreover, a theoretical analysis will be conducted following the Sapir-WhorfHypothesis of linguistic relativity and linguistic determinism. A review of these practical and theoreticalexamples demonstrates that the language we speak can shape our thinking patterns as well as how weare predisposed to view the world.
This is important because this is mostly the thesis or the reason for why this journal was written.
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underactuated.mit.edu underactuated.mit.edu
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trace
middle expression should be right multiplied by x rather than x^T:
x^T\left(Q + K^TRK\right)x
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www.jstor.org www.jstor.org
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Using Standard English isimportant because, if for nothing else, it makesmore sense [than other forms}. It is something thatwe all can understand."
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Students who have significant experiencestudying a foreign language or who grew up speak46 March 2011John W. Whiteing a language other than English tend to graspthe concept even more fully;
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ing a language other than English tend to graspthe concept even more fully;
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Students who are native English speakersand who tend to hold a static view of English cansee—and more importantly experience—how English is an ever-changing language and they can feel,albeit in a nonthreatening way, linguistic alienationwithin the English classroom.
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istance," "Sociolinguistic").To emphasize the ever-changing nature ofEnglish, my students engage in an activity in whichthey read, interpret, and then answer questionsabout two canonical pieces of English literature:
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A language withby far the largest number of words of any language,English is constantly changing and adapting to theneeds of our society.
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RecitingBeowulf aloud and with accuracy (to Old Englishnorms) is almost impossible.
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we could value these uniquediscursive forms, use them for code-switchingpurposes, and thus better induce students to addStandard English "to their existing [language] repertoire"
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oal had been to get the studentsto see that though we should teach the conventionsof Standard English, we should also acknowledgeand even celebrate the unique and highly effectiveforms of discourse that students bring with theminto the classroom.
Learn to embrace outside culture while respecting curriculum.
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My lesson on making use of thedynamic nature of English and the many forms ittakes across and within cultures certainly wasn'tgoing as planned
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In my classes, I'mgoing to require that students speakand use proper English. Period."
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of what counts as appropriate language practices in the classroom. T
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However, few students tend to see theidea of "lost in translation" as it occurs within alanguage. They generally fail to see the dynamic nature of dialects and discourses within modernAmerican English and how such dialects areuniquely positioned to express ideas and feelings
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the original, non-Standard English passage inevitably holds far more emotional and rhetorical power regardless of audience.
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working on this passage, either ingroups or as a whole class, spend inordinateamounts of time trying to decode it to come upwith a meaning that makes sense to them. A
Working far more than should be.
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Students working on this passage, either ingroups or as a whole class, spend inordinateamounts of time trying to decode it to come upwith a meaning that makes sense to them.
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www.youtube.com www.youtube.comYouTube3
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Ongoing Delivery & Demonstrating What's Next: Holochain, Unyt, HoloTap to unmute2xOngoing Delivery & Demonstrating What's Next: Holochain, Unyt, HoloHolochain 3,217 views 2 months agoCopy linkInfoShoppingIf playback doesn't begin shortly, try restarting your device.6:35Pull up for precise seekingMute11:30•You're signed outVideos you watch may be added to the TV's watch history and influence TV recommendations. To avoid this, cancel and sign in to YouTube on your computer.CancelConfirmWhat we've done is we've split the Holochain conductor intoUp nextLiveUpcomingCancelPlay NowShareInclude playlistAn error occurred while retrieving sharing information. Please try again later.6:396:50 / 27:35Live•Watch full video••15:30How the universe generates time and space from a single rewriting rule | Stephen WolframThe Well and 2 more63K views • 12 days agoLivePlaylist ()Mix (50+)56:43This Physicist (Just) Found A New Origin For GravityCurt Jaimungal120K views • 13 days agoLivePlaylist ()Mix (50+)11:19What everyone gets wrong about the double slit experimentSabine Hossenfelder424K views • 6 days agoLivePlaylist ()Mix (50+)12:14The AI Future No One Wants to Talk AboutSabine Hossenfelder364K views • 1 month agoLivePlaylist ()Mix (50+)9:24Neil deGrasse Tyson And Jaron Lanier on the AI IllusionStarTalk Plus604K views • 9 days agoLivePlaylist ()Mix (50+)20:49Why the Speed of Light Is NOT a Speed - Leonard SusskindSusskind Speaks244K views • 4 months agoLivePlaylist ()Mix (50+)16:12Why do people see elves when they take DMT? | Rupert SheldrakeThe Institute of Art and Ideas227K views • 3 months agoLivePlaylist ()Mix (50+)18:05Why Does Time Stop at the SPEED OF LIGHT? Feynman's Mind-Blowing TruthImagine the Physics313K views • 4 months agoLivePlaylist ()Mix (50+)17:04This Is Boring But It Will Make You Scary IntelligentSandeep Swadia542K views • 3 weeks agoLivePlaylist ()Mix (50+)53:57AI and the Battle for the Soul with Iain McGilchrist - Lecture 1: Information is Not UnderstandingRalston College45K views • 5 days agoLivePlaylist ()Mix (50+)18:00You’ll stop using ChatGPT after listening to this | Jonathan Pageau [ARC 2026]Alliance for Responsible Citizenship and Jonathan Pageau915K views • 1 month agoLivePlaylist ()Mix (50+)17:43Scientists Reveal Shocking Genetic Origin of HungariansHuman Discovery 60K views • 2 months agoLivePlaylist ()Mix (50+) NaN / NaN Comments 26 Top Show featured comments Newest Show recent comments, including potential spam In this video Transcript Create clip Gyuri Lajos Public Add a title (required) 0/140 – 30.0 seconds Cancel Share clip Continue clipping after ad finishes Can’t create clip while ad is playing Description Ongoing Delivery & Demonstrating What's Next: Holochain, Unyt, Holo Holochain Holochain 91Likes3,217ViewsMay 62026 Matthew Schutte sits down with Arthur Brock and Eric Harris-Braun to recap what Holochain, Holo, and Unyt shipped in Q1 2026. The key deliverables included: the live HOT to HoloFuel Technical Migration Test, a prototype of Holochain in the browser, and a wave of new Unyt capabilities for peer-to-peer payments and multi-currency accounting. More about Holochain: ✅ Twitter/X: https://x.com/Holochain ✅ Website: https://holochain.org/ ✅ Developer Discord: / discord ✅ GitHub: https://github.com/holochain More about Holo: ✅ Twitter/X: https://x.com/H_O_L_O_ ✅ Website: https://holo.host/ ✅ GitHub: https://github.com/holo-host More about Unyt: ✅ Twitter/X: https://x.com/unytco ✅ Website: https://unyt.co/ ✅ GitHub: https://github.com/unytco …...more ...more Show less How this was madeAuto-dubbedAudio tracks for some languages were automatically generated. Learn more Transcript Follow along using the transcript. Show transcript Holochain 11.6K subscribers VideosAboutTwitterRedditFacebook Transcript Ongoing Delivery & Demonstrating What's Next: Holochain, Unyt, Holo
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A Holo Web conductir
Inbrowser holo chain
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What is Holochain?Tap to unmute2xWhat is Holochain?Holochain 55,454 views 5 years agoInfoShoppingCopy linkIf playback doesn't begin shortly, try restarting your device.Pause•You're signed outVideos you watch may be added to the TV's watch history and influence TV recommendations. To avoid this, cancel and sign in to YouTube on your computer.CancelConfirmand there's nobody to fix itthese are serious usability problemsUp nextLiveUpcomingCancelPlay NowShareInclude playlistAn error occurred while retrieving sharing information. Please try again later.0:401:21 / 12:17Live•Watch full video••17:58Web3, Blockchain, cryptocurrency: a threat or an opportunity? | Shermin Voshmgir | TEDxCERNTEDx Talks615K views • 7 years agoLivePlaylist ()Mix (50+)9:52What Is Holo, Holochain? (HOT) (Whiteboard Animated)Crypto Animations2.9K views • 4 years agoLivePlaylist ()Mix (50+)48:43Dr. Gabor Maté: The #1 Reason You Never Feel Like You’re Enough (And How to Fix it)Jay Shetty Podcast598K views • 3 months agoLivePlaylist ()Mix (50+)27:36Ongoing Delivery & Demonstrating What's Next: Holochain, Unyt, HoloHolochain3.2K views • 2 months agoLivePlaylist ()Mix (50+)55:15A 28-year-old Steve Jobs gives a talk at the 1983 International Design Conference in AspenSteve Jobs Archive524K views • 1 year agoLivePlaylist ()Mix (50+)22:55Blockchain: Massively Simplified | Richie Etwaru | TEDxMorristownTEDx Talks1.8M views • 9 years agoLivePlaylist ()Mix (50+)14:05Blockchain vs Hashgraph vs DAG vs Holochain | Types of DLT ExplainedLearn with Whiteboard19K views • 5 years agoLivePlaylist ()Mix (50+)20:50RWA with Verifiable Data will Change EconomicsHolochain1.4K views • 1 year agoLivePlaylist ()Mix (50+)15:59OpenAI’s ads are failing and it can’t survive without them | Ed ZitronThe Tech Report68K views • 4 hours agoLivePlaylist ()Mix (50+)1:06:24Jövőkutató: Az AI az agyadat támadja, ezért rettegsz tőle igazából | Rab Árpád | Karizma Podcast#141Bolya Imre | Karizma Podcast73K views • 1 month agoLivePlaylist ()Mix (50+)27:13How to Restore a Broken RelationshipPastor Rick102K views • 3 days agoLivePlaylist ()Mix (50+)7:13Holo and HolochainHolo Hosting41K views • 8 years agoLivePlaylist ()Mix (50+) What is Holochain?
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bafkreihuihdcbkvnzrwslnlurofnqts3gvxlyaipzqaktwa5kmbo2mtcbm.ipfs.localhost:8080 bafkreihuihdcbkvnzrwslnlurofnqts3gvxlyaipzqaktwa5kmbo2mtcbm.ipfs.localhost:8080
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from: https://www.youtube.com/watch?v=EUfyHNGvnDo
inbrowser-holo-web-conductor
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academic-oup-com.proxy.binghamton.edu academic-oup-com.proxy.binghamton.edu
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MRNY has achieved a lot, but it has not gone beyond being a mobilizing model. Like most community ‘organizing’ groups, with the exception of the isolationist-inclined Industrial Areas Foundation (IAF)
Why?
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MRNY has adopted a detailed and transparent decision-making process. Most decisions are made by consensus, and rotating leadership is standard practice at meetings. MRNY’s “Decision-Making Authority” document (available to members in both Spanish and English) specifies in detail how people are chosen for every role and every sub-body in the organization, and specifies the authority embodied in each role and sub-body, much like a union constitution.31
Similar to our YDSA chapter 👍🏾
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The idea was to ratchet up the amount of back-pay claims a resistant employer might face, then offer a no-cost alternative: The workers would drop the claims in exchange for the employer’s agreement to not fight the unionization effort.
LEVERAGE!!
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127.0.0.1:8080 127.0.0.1:8080
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what is holochain
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volla.online volla.online
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The first cloud in the world without a data center.
virtual cloud
personal-first virtual cloud
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Holochain. Time for a paradigm shift
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higheredstrategy.com higheredstrategy.com
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revenue from international students
International student!! Also, we have the researches in medicine and technology with other sciences filed get refunds from it. However, the education system can get more money then spend if we use it correctly.
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ageing population
Ageing population!! Is that the real problem, or does the government spend our tax money on other countries for any reason? Because old people still need more spend especially for health, and to make their lives easy and nice, just like they made our lives good when they were young and worked hard with a little technology; they used their brains and bodies to get jobs does not depend on AI!!
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support for public postsecondary education!
The government of all provinces needs to remember the time in 2020 when we all suffered from COVID-19. At that time, all we needed was education: people from doctors, nurses, researchers, and teachers to make our lives safe and continue. Did these education people sacrifice their lives to save the government's life?!!
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Ontario…is Ontario.
WHY Ontario... is Ontario?! Do we need to move t o another province to get a good spot for education, or is it the total subject of life in Ontario? When we came to Canada in the first, we did not know we need to read such an article or report to find what is the best province which province spends more money on the education system. However, Canada is one country, but there is a difference between their provinces.
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More Eating the Future
Interesting title! From the past to the future, education has been an important center point for society and the community, not just in Canada. Each country needs to focus on education first if they want to shine in the future; they need to spend more money on the education system, from teachers and schools to students themselves.
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en.wikipedia.org en.wikipedia.org
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💻/thinkpad/🧊/me/📓/2026/7/0/3/=/PageRank@Wikipedia/
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cielitolindoseniorliving.com cielitolindoseniorliving.com
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www.biorxiv.org www.biorxiv.org
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An external discussion of this preprint is available on PubPeer: https://pubpeer.com/publications/8DA4B0AA40D8625B196C1014E32ED3
The discussion focuses on the study’s cross-validation design, including whether random rather than contiguous splits could allow temporal dependencies to influence the reported encoding results. It also considers how the language features were defined, how the findings compare with concept-cell coding, and how strongly the results support interpretations involving pattern separation and completion. Readers may wish to consult the original discussion for the full context.
paperstack.pub · The trust layer for preprints
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arxiv.org arxiv.org
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novel multi-resolution
This is the new Try
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hypothes.is hypothes.is
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Check out some of the
Who is Public...?
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www.qnap.com www.qnap.com
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Push service uses the myQNAPcloud service as the server.
Does Qmanager also require myQNAPcloud?
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We recommend choosing push service for notifications that are not important or urgent.
Then what is the actual use case?
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www.youtube.com www.youtube.com
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1946 Electromatic Model 01 - IBM's First Typewriter <br /> by [[Tech Tangents]]
Not the best cleaning and repair advice here for a typewriter, but some interesting tidbits of IBM history.
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Tip: of the 3 things WD-40 is actually useful for on a typewriter, one is cleaning and giving a nice sheen to crinkle paint safely. Apply lightly on a clean rag. The other two are cleaning white-out off plastic card guide without fogging the plastic, and removing the calcified masking tape often found on SCM portables.
via Ted Munk as a comment on a YouTube video
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laughing-bassoon-7p6pm2g.pages.github.io laughing-bassoon-7p6pm2g.pages.github.io
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POST /v4/data/exports/models create → 202 + { jobId }
do we also use client account? as in feed configuration?
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www.biorxiv.org www.biorxiv.org
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Note: This response was posted by the corresponding author to Review Commons. The content has not been altered except for formatting.
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Reply to the reviewers
Reviewer #1:
Major comments:
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Lines 103-116 (first paragraph of the results section) describe mainly published data that is more suitable for the introduction section. It is annoying to refer to different published articles in the Results section to strengthen the results instead of showing them. The same goes for paragraphs two and three. Why mention those data in the Results section if they are already published and known?
We have reorganized this material by moving some background information to the Introduction. Our intention was not to incorporate published data to strengthen our results, but rather to provide essential context for interpreting our findings. We have therefore left some of this foundational information in the results section to create a clear narrative flow, enabling readers to understand the basis for our experimental design and interpretations without needing to recall details from earlier paragraphs in the Introduction. For example, we considered it crucial to restate the earlier report of the BiP:sfGFP:HDEL phenotype in Atlastin mutants, since our results supporting luminal ER protein displacement contradict the previous fragmentation model.
The following concept was in line 103 in the Results section, and is now in the introduction in lines 82-92: "Conventional light microscopy, commonly used in studies of neuronal ER structure, lacks the resolution necessary to visualize individual ER tubules in small structures, such as presynaptic terminals. The ER is highly sensitive to fixation, and live imaging experiments in neurons in vivo have been conducted on upright microscopes using water dipping objectives with a typical axial resolution limit of >300 nm, which cannot distinguish the densely packed ER tubules at presynaptic terminals (3,8,21,28,39-42). Electron microscopy offers higher resolution, but cannot be used in live samples and has typically been limited to thin 2D sampling (in which it is difficult to distinguish ER cross-sections from synaptic vesicles) (8,20,22)."
Figure Legends-(in all Figures): The number of experimental repeats must be mentioned in the figure legends.
This information is provided in Supplementary Table 1, which contains detailed information about the genotype, statistical analysis, and number of larvae and NMJs analyzed. If the journal requires this information in figure legends, we can move it.
The way the figures are labeled is worrisome; supplementary figures are not ordered numerically.
We will be happy to rename supplementary figures according to journal guidelines.
The tubule extension in Figure 2D is not convincing. Is there a movie showing those changes? Better images are needed. It is essential to show which supplemental movie corresponds to which panel.
We have now included a corresponding video of the same neuron used as an example of tubule extension. We also added another frame to the figure to provide further information on the tubule event we captured. (Figure 2D, Movie S10)
This is unnecessary in the results section: "To investigate the relationship between ER structure and function at synapses, we examined mutants of Atlastin, a GTPase that regulates ER tubule fusion. Drosophila has a single homolog while mammals have three Atlastin homologs, with Atlastin-1 enriched in the brain (Rismanchi et al., 2008)."
This information was moved to the introduction.
"This reduction in ER membrane marker intensity has also been observed in other HSP mutants, suggesting this is a common feature of ER shaping mutants and could indicate changes in ER membrane composition, integrity, or tubule thickness (Perez-Moreno et al., 2023)." This comparison is important and should be shown in the same settings as for the Atlastin mutant rather than referring to published data.
We agree with the reviewer that it is important to determine whether other ER-shaping proteins, besides Atlastin, also show a decrease in tdTomato:Sec61b to support our claim that this could be a common feature among ER-shaping mutants. To do this, we examined mutants of another ER-shaping protein, Reticulon 1, which regulates membrane bending and stabilization in ER tubules. These loss-of-function mutants were a gift from Dr. Cahir O'Kane at the University of Cambridge and were used in his lab's Pérez-Moreno et al., 2023 publication. We found that in our hands tdTomato:Sec61b levels were reduced in Reticulon 1 mutants, consistent with the results reported by Pérez-Moreno et al. (2023). These results are in Figure 3E-F. We also examined the synaptic distribution of the luminal ER marker, BiP:sfGFP:HDEL, in Reticulon 1 mutants to see if it is displaced to the cytosol. Notably, it remained ER-associated, unlike in Atlastin mutants. These results are in Figure 6F-G, results lines 267-270, and discussion lines 542-545.
Does the distribution of the luminal ER marker in Figure 6F diffuse due to mislocalization or reflux after being localized to the ER and then refluxed to the cytosol as was previously shown for the ER to Cytosol signaling (ERCYS) mechanism? Could you assess other ER-luminal protein localization biochemically? It is highly recommended to look at another soluble ER-protein localization in the Atlastin mutant without overexpression, which can be an artifact.
ER stressors can induce ERCYS, in which some luminal proteins, including PDIA3, DNAJB11, ERp29, and an eroGFP reporter, reflux by 30-70% to the cytoplasm without subsequent degradation (unlike ERAD (ER-associated degradation). This phenomenon has only previously been observed in yeast and glioblastoma tumor cells from mice and human . We believe that our work provide the first suggestion that this may occur in neurons, and particularly in a neurological disease model.
We do not believe that the reflux phenotype for BiP:sfGFP:HDEL is due to its overexpression for two reasons: (1) we observe reflux in our neuronal Atlastin knockdown experiments, even when the levels of BiP:sfGFP:HDEL are significantly reduced artificially because of titration of the GAL4 between the RNAi and the reporter (Figure 7A), and (2) BiP:sfGFP:HDEL overexpression somewhat suppresses endogenous BiP upregulation ((Figure 10 and see Reviewer 1.10), arguing that the transgene does not induce ER stress). We included a new "limitations of the study" section to be transparent about the caveats of the BiP:sfGFP:HDEL reporter (lines 639-664).
Identifying potential endogenous neuronal ERCYS substrates in our in vivo preparation poses several challenges. First, biochemical approaches, such as fractionation, are not possible in our complex in vivo sample because neuronal ER proteins would mix with ER from other tissues upon homogenization. Second, detecting endogenous proteins with antibodies requires fixation and permeabilization, which notoriously disrupts ER structure and even causes our reporter BiP:sfGFP:HDEL to collapse from a smooth distribution, as visualized by live imaging and FRAP, to a punctate distribution. Third, using antibodies rather than neuronally restricted transgenes makes it challenging to determine whether the signal originates from the neuron or from dense ER structures in the surrounding muscle. Fourth, some ER luminal proteins can displace as little as 30% in the ERCYS examples cited above, and the sensitivity of our imaging assays may limit our ability to detect these small changes. Finally, the limited availability of tagged transgenes and antibodies specific to Drosophila luminal ER proteins (see next paragraph) poses additional challenges. These limitations highlight the need for future studies to develop novel tools and techniques to more definitively test whether we are indeed observing ERCYS. We have included a paragraph on these future challenges in our discussion in lines 639-664. Identifying endogenous targets of ERCYS in fly neurons is a worthwhile goal, but beyond the scope of the current study. These next steps will particularly benefit from identifying the machinery involved in the reflux of our BiP:sfGFP:HDEL reporter.
Tools we tested: We investigated several options: (1) a tagged PDI transgene (a gift from Karen Hibbard), which was not detectable at presynaptic terminals, (2) a tagged BiP (FlyORF; F000956) that did not localize to the ER, and (3) full-length endogenous BiP detected by antibody staining. We did not detect obvious reflux of endogenous BiP to the cytoplasm (Figure 9), with the caveat that in fixed samples, the BiP signal was not tightly co-localized with the ER marker even under control conditions. However, we did use this antibody to detect an increase in BiP in Atlastin mutant presynaptic terminals, indicating ER stress (see Reviewer 1.10).
Though we have not identified endogenous targets, we believe that our studies with the exogenous reporter will be of great interest to the field, as they clarify the previously reported Atlastin phenotype and provide the first report of a new defect in a human disease animal model.
In comparison to Summerville et al. (2016) in Figure 7, the experiment was not done in the same way. It is important to keep the same settings for comparison
In Figure 7D-E, we compare the distribution of BiP:sfGFP:HDEL in cell bodies, axons, and muscles between controls and Atlastin mutants. To clarify the experimental approach relative to Summerville et al. (2016): while both our studies examined the same cellular compartments (cell bodies, axons and nerve terminals) using the BiP:sfGFP:HDEL reporter, we employed super-resolution Airyscan microscopy. This enhanced resolution was critical for definitively demonstrating that this is a functional rather than a structural phenotype and that ER displacement is progressive, and repeating this experiment at lower resolution as previously reported does not provide any new information. We identified two distinct distribution phenotypes in Atlastin mutants expressing BiP:sfGFP:HDEL, which were not described in the Summerville et al., 2016 paper. From our manuscript (lines 249-251): "We identified two distinct ER network phenotypes in Atlastin mutants expressing BiP:sfGFP:HDEL: "Partial loss" NMJs retained both diffuse signal and identifiable ER network structures, while "Complete loss" NMJs showed no visible ER network structures. Note that the "Complete loss" phenotype in Atlastin mutants reflects the absence of detectable luminal marker signal in organized ER structures, but not the complete absence of ER membranes, as demonstrated by our ER membrane marker tdTomato:Sec61β results."
Does the Atlastin mutant induce the unfolded protein response and stress within the ER? It is necessary to look for UPR markers in those settings. It was shown previously that ER stress leads to protein reflux from the ER to the cytosol. Is there a difference in the ER stress markers in the presynaptic terminal?
The reviewer suggested that Atlastin mutant synapses may exhibit ER stress. To address this, we examined levels of the ER chaperone BiP, a well-established ER stress marker whose expression increases during UPR activation. We first validated that our BiP antibody can detect changes in ER stress by feeding control larvae with 50mM DTT for 24 hours. These results are in the new Figure 10A. Note that we were unable to test sensitivity to ER stress in this way in Atlastin mutant larvae because they did not consume the DTT-treated food, as assessed by blue food coloring in the larvae's guts.
Using this antibody, we measured baseline BiP levels at NMJs of Atlastin mutants on normal food, and found they were slightly increased compared to controls. We conclude from these experiments that Atlastin mutant synapses have mild ER stress. Notably however, Atlastin mutants co-expressing UAS-BiP:sfGFP:HDEL or UAS-tdTomato:Sec61b did not show significantly increased endogenous BiP levels, suggesting that transgene expression at least partly suppresses the mild ER stress response, even though there is extensive cytosolic displacement. These results argue (1) that the mild ER stress in Atl mutants does not strictly correlate with the reflux phenotype, and (2) that the reflux phenotype is not an artifact of overexpression-induced stress. These results are described on lines 430-436 in the results section and shown in Figure 10B-E, and their implications discussed on lines 585-598.
We also explored another strategy to detect ER stress by assessing eIF2α phosphorylation, a key event in the Unfolded Protein Response (UPR) pathway. We obtained a phospho-eIF2α antibody (Cell Signaling; #3597) that was reported to work in Drosophila. However, when we tested this antibody by Western blot, we were unable to detect a band at the expected molecular weight for phosphorylated eIF2α, even in positive-control samples treated with DTT to induce ER stress. We therefore concluded that this antibody is not suitable for reliably detecting ER stress in our experimental system. The failure of this antibody highlights the challenges of finding robust tools to measure ER stress in Drosophila.
It is important to add biochemical experiments to show that no fragmentation of the ER membrane occurred. It can be simply demonstrated by looking at the redox state of the ER, which would change if it were mixed with the reducing cytosol. Moreover, this can be shown by using an ER-targeted redox-sensitive fluorescent protein that is tethered to the ER membrane to follow changes in the redox state of the ER.
The reviewer asked us to test whether the redox state of the ER is disrupted, which could indicate exchange between the cytosol and ER due to membrane rupture. As noted above, biochemical approaches such as fractionation are not possible in this in vivo sample. We attempted to address this concern by creating a UAS-Sec61β:roGFP construct, using the roGFP sequence from Igbaria et al. (2019) to monitor the ER lumen redox environment in Atlastin mutants. Since Sec61β is membrane-tethered, it should remain in the ER and not undergo reflux, making it an ideal sensor for detecting any mixing between the reducing cytosolic environment and the oxidizing ER lumen that would occur if membrane fragmentation and/or ruptures were present. We tested this approach in wild-type Drosophila S2 cells and used the Gal4-UAS binary expression system to co-express Actin-Gal4 (to drive expression of UAS constructs), UAS-Sec61β:roGFP (redox sensor), and UAS-BiP:Halo:HDEL (as a control reporter insensitive to DTT treatment).
Our experiments showed no detectable changes in the fluorescent properties of UAS-Sec61β:roGFP following 30 min 10mM DTT treatment compared to DMSO vehicle control, including no increase in 405-nm excitation fluorescence or changes in 488nm/405nm excitation ratios. These results suggest that either the roGFP sensor requires further optimization for sensitivity in this cellular system or that additional controls and calibration steps are needed to establish the dynamic range of the assay. We believe this experiment falls beyond the scope of the current study, given the extensive optimization required. However, it represents an important future direction for testing membrane fragmentation as a mechanism underlying the phenotypes observed in Atlastin mutants. The possibility of ER integrity defects is mentioned in the discussion on lines 547-559.
Minor comments:
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It is important to call figures by order. Figure 2C is called before 2A-B. Figure 2B is called before Figure 2A.
The revised manuscript has all figures in order of appearance in the text.
Figure legends (Figure 2): "The same control dataset used in E-G was used in Figure 5 and Figure 5_Supplement." Why is this relevant?
We wanted to be transparent about reusing the same control dataset across multiple figures to avoid any appearance of data duplication. This notation clarifies that, although the data appear in different contexts (Figures 2 and 5. This version does not contain a Figure 5_Supplement), it represents the same biological samples analyzed for different parameters, ensuring readers understand that these are not independent datasets.
Figure 4F is called before Figure-4D-E which are not called.
We revised our manuscript and reorganized Figure 4 to ensure that all figure panels are referenced in sequential order and that panels 4D-E, which were previously not cited in the text, are now properly referenced when discussing their corresponding results.
Figure 5B is called before the previous ones. Same for Figure 5A supplement.
We referenced Figure 5A in lines 211-212, which precedes our discussion of Figure 5B. To clarify the figure order, we removed the early references to Figures 2D-G and Movies 7-14, which were mentioned only to indicate that we were analyzing the same dataset in different ways.
The revised manuscript has all figures in order of appearance in the text.
Referees cross-commenting
I agree with the comments raised by reviewer2 and 3. Basically it is highly important to validate those data by genetic rescue. Moreover, it is essential to know the source of the displaced luminal marker to the cytosol. Is it mislocalization or it is a reflux of pre-existing protein to the cytosol after insertion to the ER. It is also recommended by me and the reviewers and me to test the endogenous protein rather than overexpression.
We have addressed these points in our responses to the following reviewer questions:
- Genetic rescue: Please see our responses to Reviewer 1/Question #10 and Reviewer 2/Question #1.
- Source of displaced luminal marker: We provide some evidence addressing this in our response to Reviewer 3/Question #1.
- Endogenous protein localization: We have examined this and detailed our findings in our responses to Reviewer 1/Question #7 and Reviewer 2/Question #6.
Reviewer #1 (Significance (Required)):
General assessment: This interesting paper shows that proteins can escape the ER under special conditions. However, the authors need more evidence to show that and rely less on the overexpression system, especially of BIP-GFP, which can cause proteostasis stress within the ER. Advance: The results have been oversimplified in their explanations, and some points and complexities of the study need to be addressed further to make the most of them. These are often some of the more interesting concepts in the paper. I think many points can be addressed in the text by the authors being clear and concise with their reporting. At the same time, other experiments would turn this paper from an observational one into a very interesting mechanistic one. This paper is based on previously published articles from the group and other groups, and it is a nice progression. However, as mentioned, this paper depends primarily on published data, and the novelty is somehow lost between all the comparisons to other published data instead of emphasizing that. Without a substantial mechanistic improvement, the paper would remain observatory.
Audience: The microscopy tools can be great addition to researchers in the field to monitor protein trafficking especially Cell biologists (basic research)
My expertise: ER homeostasis, protein trafficking, cell biology
Reviewer #2 (Evidence, reproducibility and clarity (Required)):
Summary The endoplasmic reticulum (ER) is a continuous organelle that extends throughout neurons to regulate fundamental processes. The analysis of ER dynamics at synaptic terminals is limited by the challenge of imaging these structures at high resolution. In this manuscript, the authors use super-resolution (~170 nm) live imaging and a combination of membrane and luminal ER markers at the Drosophila larval NMJ, an important model synapse, to investigate dynamic ER architecture in vivo. They report a detailed characterization of the presynaptic ER organization and dynamics at wild-type and GTPase Atlastin mutant NMJs. Their analysis using the ER membrane marker tdTomato:Sec61b reveals the presence of an intact ER network in Atlastin mutants. This contrasts with the apparent ER fragmentation phenotype previously reported and replicated here when using a luminal marker. Their findings instead point to the progressive displacement of luminal proteins to the cytosol in Atlastin mutants specifically at synapses. The authors propose that the disruption of ER protein dynamics at synapses is a compartment-specific ER stress response. The manuscript is well written, results are clearly presented, and experiments are technically rigorous.
Major comments
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The baseline ER phenotypes in Atlastin mutants are mild with complete loss of ER network only observed in terminal boutons. This interesting and unexpected result should be further confirmed by genetic rescue. The authors can use a UAS rescue line previously reported in PMID: 19341724.
We tested the UAS-Atl-myc rescue line and unfortunately found that even in wild-type neurons, overexpression of Atlastin produced strong ER organization defects that precluded the rescue experiment. Instead, to confirm the cell autonomy of the phenotype and to test it wth an independent tool, we performed a presynaptic knockdown of Atlastin by RNAi and found that BiP:sfGFP:HDEL is displaced, as observed in the Atlastin null mutant. These results are in now shown in Figure 7A-C.
Lines 204-7: It's not clear how a greater coefficient of variation indicates that the marker is more concentrated in subsynaptic structures or what is meant by 'subsynaptic structures.'
We added the following text to explain, in lines 181-183: "A higher CoV indicates an uneven distribution of tdTomato:Sec61β within the presynaptic terminal, with some areas showing higher concentrations than others (in contrast to the uniform, diffuse signal expected from fragmentation)." To avoid confusion with postsynaptic structures called the subsynaptic reticulum, we have removed the term "subsynaptic". The intended meaning is distinct structures found within the presynaptic terminal.
There's a mistake in Figure 6C and the associated text. The summed percentage of the three phenotypic categories adds up to 110% for Atlastin mutants.
The reviewer noted that the summed percentage of the three phenotypic categories in Figure 6C adds up to 110% for Atlastin mutants, which appears to be a mathematical error. However, this is not an error, but rather a reflection of our quantification methodology, in which a single bouton can exhibit more than one type of ER dynamics per movie recorded. Our quantification counts each phenotype independently, so boutons displaying multiple phenotypes contribute to more than one category. This approach provides a more comprehensive view of the range of ER dynamics present in Atlastin mutants, as restricting the analysis to mutually exclusive categories would underrepresent the complexity of the phenotypes observed. To make this point clear, we made the following change to the text in lines 257-259: "We note that the sum of these percentages exceeds 100% because one NMJ exhibited multiple phenotypes: one branch had a complete loss, while the other branch had no phenotype. These phenotypes were counted separately."
Figure 8: the ER looks fragmented in 1st instar controls and mutants. The authors should address this difference from more mature NMJs.
We would like to clarify that the bulk of experiments in this manuscript (including all ER dynamics, luminal marker redistribution, and membrane marker analyses discussed throughout the Results) were performed in 3rd instar larvae, which are more mature larval NMJ preparations standard in the field. Figure 8 was included specifically to test whether the Atlastin mutant phenotype we describe throughout the paper is also detectable at an earlier developmental stage, not to replace or reinterpret our primary findings.
Regarding the specific observation that the ER appears more fragmented in Figure 7F-H relative to the more mature NMJs shown elsewhere: this fragmentation, observed similarly in both control and Atlastin mutant 1st instar larvae, likely reflects technical challenges associated with dissecting these smaller, more delicate early-stage specimens rather than a genotype-specific effect. Because fragmentation occurred similarly in both genotypes, we could still reliably assess the redistribution of BiP:sfGFP:HDEL as our primary phenotypic readout in this experiment. We have added the following text (lines 306-309) to clarify this point: "Note that in 1st instar larvae, both normal networks in controls and residual networks in Atlastin mutants appeared more fragmented than in 3rd instar preparations, likely due to the technical challenges of dissecting these smaller, more delicate specimens. Since ER fragmentation occurred similarly in both genotypes, we could still reliably assess the redistribution of BiP:sfGFP:HDEL as our primary phenotypic readout.
The images in figure 9B do not seem representative of the quantification in Figure 9D. Specifically, the partial loss Atlastin NMJ appears to have recovered as fully as the complete loss Atlastin NMJ.
The images showed FRAP recovery across the entire bouton, but we photobleached only a small region within each bouton and quantified only this region. We have now added outlines to clearly delineate the specific FRAP regions that were analyzed in each image, which clarify that the partial loss Atlastin showed less recovery than the overall bouton. We have also reordered the figures to more clearly convey our message (Figure 9 is now Figure 8).
We also made a few changes to the paragraph on lines 347-350 to clarify our experimental reasoning: "We photobleached en passant boutons using a defined region of 6.8 x 7.8 microns (dashed box in Figure 8D) to ensure that BiP:sfGFP:HDEL could recover from the ER networks surrounding the FRAP region (Movies S20-S23)."
We also added this sentence to the figure legends of Figure 8: "The dashed boxes in (D) indicate areas that were photobleached and analyzed for recovery quantification in (E-F)."
Optional: An overexpressed luminal marker is displaced to the cytoplasm in Atlastin mutants. It would be interesting to know and increase the significance of the findings if the same is true of endogenous luminal proteins under biological stress conditions.
As noted in our response to Reviewer #1 suggested that Atlastin mutant synapses may exhibit ER stress. To address this, we examined levels of the ER chaperone BiP, a well-established ER stress marker whose expression increases during UPR activation. We first validated that our BiP antibody can detect changes in ER stress by feeding control larvae with 50mM DTT for 24 hours. We were unable to perform this experiment in Atlastin mutant larvae because they did not consume the DTT-treated food, as assessed by blue food coloring in the larvae's guts. These results are in Figure 10A. In the future, it will be of interest to establish a protocol to examine Atlastin mutants by feeding or treating larval fillets with DTT.
We measured BiP levels at NMJs of Atlastin mutants and found they were slightly increased compared to controls. Atlastin mutants co-expressing UAS-BiP:sfGFP:HDEL or UAS-tdTomato:Sec61b did not show significantly increased endogenous BiP levels, suggesting that transgene expression suppresses the mild ER stress response. We conclude from these experiments that Atlastin mutant synapses have mild ER stress. These results are in Figure 10B-E).
Optional: Applying this approach in stimulated conditions (high potassium, increased temperature) might reveal a greater activity-dependent role for Atlastin at synaptic terminals.
This is a very interesting idea, as we have only examined synapses at rest. However, this is beyond the scope of this paper.
Minor Comments
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Line 16: Atlastin should be italicized.
Thank you for catching this typo. We have fixed it.
Figure 5A: Based on the relative intensities, it appears that control and mutant images are not contrast matched but this isn't stated.
Thank you for catching this omission. We added to the figure legend: "Control and Atlastin mutant images are not contrast matched."
Line 822: The number of static Atlastin mutant boutons used for analysis is missing.
Thank you for catching this omission. We have fixed this supplementary table.
Figure 9: The blue arrows are not annotated in the figure legend.
Thank you for catching this omission. We have fixed this figure legend.
Reviewer #2 (Significance (Required)):
Atlastin is linked to Hereditary Spastic Paraplegia (HSP) and this study changes our understanding of the compartment-specific impacts of its loss. This study reveals the importance of using both membrane and luminal ER markers to accurately interpret phenotypes as well as the importance of considering compartment-specific effects on ER. These findings represent significant mechanistic and conceptual advances. The lack of genetic rescue is a limitation and adding an investigation of an endogenous luminal protein under basal and stress conditions would add significantly to our understanding of Atlastin dysfunction in HSP. Notably, the in vivo imaging approach introduced here can be adapted broadly for live imaging of Drosophila larvae. Thus, this work will be of interest to both neuronal cell biologists and the wider Drosophila community. This review is based on our expertise in neuronal cell biology.
Reviewer #3 (Evidence, reproducibility and clarity (Required)):
In this manuscript, the authors investigate the structural dynamics of the endoplasmic reticulum (ER) in Drosophila neurons and examine the role of the ER-shaping protein Atlastin in ER morphology. Their discovery on the neuromuscular junction (NMJ)-specific contribution of Atlastin to ER integrity is intriguing and may provide valuable insights into the pathological mechanisms underlying Atlastin mutations associated with hereditary spastic paraplegia (HSP) and hereditary sensory neuropathy. The key observation on ER protein showing an aberrant cytoplasmic localisation in mutant cells appears convincing. Though this phenomenon's characterisation stays at the point of primary observation with its mechanics unclarified, establishing this new and unexpected functional rather than structural Atl effect is important and useful for the field. The observation that ER is structurally preserved in this mutant with absolute lack of Atl are also extremely useful.
It is unclear if the cytoplasmic localisation affects an exogenous overexpressed ER marker or endogenous protein would also appear in cytoplams, the authors should consider adding an immunostaining data to test that.
Authors offer speculations on potential reasons for the cyto localisation of the ER marker suggesting that relocation at the cell periphery specifically combined with slow clearance there is the most likely explanation (still unclear what stops the marker from spreading through the entire cell). They suggest that decrease in cotranslational translocation is unlikely as this would result in somatic accumulation of the marker. However, if the clearance in the periphery is less efficient than in soma, the accumulation there might reflect a compromised translocation. Any clarifying experiments, if practical, to directly demonstrate how ER proteins in relocates to the cytoplasm in atl mutant would help understanding better the phenomenon. For example, would proteasomal inhibition make the marker accumulate more across the cell? Authors also suggest links to ER stress. Would stress induction phenocopy the mutant?
Reviewer #3 asked whether defective proteasomal clearance underlies the cytosolic accumulation of BiP:sfGFP:HDEL in Atlastin mutants. We addressed this directly. First, proteasome function appears intact in the mutants: baseline ubiquitinated protein levels (FK1 antibody) were comparable between control and Atlastin mutants, and MG132 treatment produced a similar increase in ubiquitination in both genotypes, confirming both antibody specificity and normal proteasome activity. We then examined BiP:sfGFP:HDEL directly. In controls, MG132 caused the marker to accumulate at axons and presynaptic terminals, showing that it is normally cleared from these compartments by the proteasome. Critically, this accumulated marker remained associated with intact ER networks: MG132 did not induce diffuse cytosolic BiP:sfGFP:HDEL in any compartment (cell bodies, axons, or presynaptic terminals), even where levels rose substantially. Thus, blocking proteasomal clearance raises ER-localized marker but does not generate the cytosolic pool seen in Atlastin mutants, indicating that impaired clearance is not sufficient to cause the displacement phenotype. We separately noted that BiP:sfGFP:HDEL was already elevated in Atlastin mutant axons without MG132, paralleling the axonal tdTomato:Sec61β accumulation in Figure 4, consistent with reduced baseline clearance specifically in mutant axons, but this does not lead to cytosolic displacement. This experiment is now shown in Figure 11, described in Results (lines 445-475), and discussed in lines 576-581.
Minor comments:
Line 146:
"fast dynamics (Thank you for catching this mistake. We have corrected it.
Fig. 2D: The data representation of "Tubule displacement" image is unclear. The ER tubule indicated by the red arrow does not seem to show any changes over time (like static). time 0 in stamp appears behind the image.
Thank you for catching the typo. We have fixed it. Additionally, we added black arrows to highlight a tubule that is not moving, allowing the reader to compare it with the moving tubule. We also included a video of all types of ER tubule dynamics to ensure the reader can also look at the raw data (Movies S9-11).
Line 157-158 (and relevant method sections):
The definition of static and dynamic boutons is ambiguous. The author should describe in more detail this point including how long they observed the structure to define the changes in ER tubule dynamics.
We provide in the methods (lines 779-791) a detailed explanation of how we categorized boutons as dynamic or static. In addition, we added the following to explain in the results section how we defined static vs dynamic:
Old sentence: We qualitatively categorized boutons as "static" if we observed no change in ER network structure or "dynamic" if we observed at least one change.
New sentence in lines 143-147: "We imaged boutons for 40 sec at 0.92 sec intervals to capture ER dynamics over this observation period. Boutons were qualitatively categorized as "static" if we observed no detectable changes in ER network structure throughout the entire 40 sec imaging session, or "dynamic" if we observed at least one of the three defined dynamic events during this time window."
Fig. 2E: What n=75 and n=29 represent is unclear, are these the number of boutons in en passant and terminal subjected for qualitative analysis?
We removed these n values from the figure and added this information to the Supplementary Table 1, which contains detailed information about the genotype, statistical analysis, and number of larvae and NMJs analyzed.
Fig. 2: What the qualitative analysis represents is unclear, are the points pulled from different experiments?
The data in Fig. 2 E-F comes from movies acquired in the same experiment. The number of independent animals and NMJs imaged is described in Table 1.
* *Line 231: Regarding "...we found a small but significant reduction in dynamic boutons in Atlastin mutants (76%), ...", how do the authors assess significance. If proportion of static/dynamic ER in boutons was obtained from multiple experiments, it should be presented e.g. as in average {plus minus} standard deviation, or clarify that the proportion is representative of x independent experiments.
The videos used for this figure were acquired from a single experiment. We use a chi-square test to determine significance relative to the "expected" distribution of dynamics types from controls, as these are categorical rather than continuous data (see PMID 31145670). Information regarding genotype, statistical analysis and number of larvae and NMJs can also be found in Supplementary Table 1.
Line 267-269 and Fig. 6B: The author's conclusion that "Complete loss of ER network structure in NMJ of BiP:sfGFP:HDEL overexpressing Atl mutant" seem to be based on the lack of signal from luminal marker, which may be undetectable due to changes to tubular volume or marker loss to the cytoplasm, as suggested by the authors, while the membranous ER structure is intact. It would be useful to discuss this point and potentially add ER membrane-stained control.
We agree with the reviewer that Atlastin mutants categorized as 'complete loss mutants' do not actually lack ER at synapses. We think this is an important point so we added the following to the results in lines 251-254: "Note that the "Complete loss" phenotype in Atlastin mutants reflects the absence of detectable luminal marker signal in organized ER structures, not the complete absence of ER membranes, as demonstrated by our ER membrane marker tdTomato:Sec61β results."
We attempted to co-label the ER membrane and ER lumen, but these crosses yielded very few live larvae (in either controls or Atlastin mutants, and those that survived had severely deformed NMJs. We added Figure 6-Supplement showing the results of this experiment, and described them on lines 270-273.
Fig. 6C: In Atl mutant, why does the total of the proportion exceed 100% (10 + 45 + 55)?
The reviewer noted that the summed percentage of the three phenotypic categories in Figure 6C adds up to 110% for Atlastin mutants. This is not an error, but rather a reflection of our quantification methodology because a single bouton can exhibit more than one type of ER dynamics per movie recorded. Our quantification counts each phenotype independently, so boutons displaying multiple phenotypes contribute to more than one category. This approach provides a more comprehensive view of the range of ER dynamics present in Atlastin mutants, as restricting the analysis to mutually exclusive categories would underrepresent the complexity of the phenotypes observed. To make this point clear, we made the following change to the text in lines 257-259: "We note that the sum of these percentages exceeds 100% because one NMJ exhibited multiple phenotypes: one branch had a complete loss, while the other branch had no phenotype. These phenotypes were counted separately."
Fig. 9C, line 342-344: In FRAP experiment using CD8, it seems that the Partial loss Atl mutant shows slower recovery that control. There seems to be a mismatch in triangle symbols of Partial loss Atl mutant between legend and plot (one is filled and the other is empty). This should be clarified.
Thank you for catching this mistake. We have fixed the figure.
fig. 10 is a clever way to verify the cytoplasmic localization of the ER marker; however, its description and annotation can be improved, and it would be stronger if 4 curves in F for mutant and controls with the trap and normal were shown.
The reviewer suggested merging our graphs but we believe that keeping them separate is clearer.
Line 495: Drosophila have ReepA and ReepB, but not Reep1-4. If the authors discuss their speculation based on their observation (using Drosophila), the gene names should be unified in the same species, and explain the corresponding genes to mammalian cells.
We made the following changes to address the reviewer's concern about gene nomenclature consistency (lines 502-506): "These ER-derived vesicles are likely to involve ReepA and ReepB, the Drosophila orthologs of mammalian REEP1-4, which regulate ER vesicle formation in mammalian cells (67). Notably, while overexpression of Atlastin can regulate REEP vesicle fusion in mammalian systems (67), it is not essential for vesicle formation, suggesting similar regulatory relationships may exist between Atlastin and Reep genes in Drosophila."
Line 548; should UPR be Unfolded Protein Response?
Thank you for catching the typo. We have fixed it.
Reviewer #3 (Significance (Required)):
This study advances the understanding of how ER morphogens affect neuronal cells specifically, the lack of which limits researchers ability to comprehend the neuronal pathologies associated with ER structure-function. The observation on ER content aberrant localisation caused by the lack of key structural protein should be of a great interest for cell and neuronal biologists and researchers of the associated diseases and shows the field a new direction. Though, mechanistic details remain to be unraveled, it constitutes a fundamental, conceptual advance.
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Referee #3
Evidence, reproducibility and clarity
In this manuscript, the authors investigate the structural dynamics of the endoplasmic reticulum (ER) in Drosophila neurons and examine the role of the ER-shaping protein Atlastin in ER morphology. Their discovery on the neuromuscular junction (NMJ)-specific contribution of Atlastin to ER integrity is intriguing and may provide valuable insights into the pathological mechanisms underlying Atlastin mutations associated with hereditary spastic paraplegia (HSP) and hereditary sensory neuropathy. The key observation on ER protein showing an aberrant cytoplasmic localisation in mutant cells appears convincing. Though this phenomenon's characterisation stays at the point of primary observation with its mechanics unclarified, establishing this new and unexpected functional rather than structural Atl effect is important and useful for the field. The observation that ER is structurally preserved in this mutant with absolute lack of Atl are also extremely useful.
It is unclear if the cytoplasmic localisation affects an exogenous overexpressed ER marker or endogenous protein would also appear in cytoplams, the authors should consider adding an immunostaining data to test that.
Authors offer speculations on potential reasons for the cyto localisation of the ER marker suggesting that relocation at the cell periphery specifically combined with slow clearance there is the most likely explanation (still unclear what stops the marker from spreading through the entire cell). They suggest that decrease in cotranslational translocation is unlikely as this would result in somatic accumulation of the marker. However, if the clearance in the periphery is less efficient than in soma, the accumulation there might reflect a compromised translocation. Any clarifying experiments, if practical, to directly demonstrate how ER proteins in relocates to the cytoplasm in atl mutant would help understanding better the phenomenon. For example, would proteasomal inhibition make the marker accumulate more across the cell? Authors also suggest links to ER stress. Would stress induction phenocopy the mutant?
Minor comments:
Line 146: "fast dynamics (<1 sec)" a velocity should be presented as distance/time
Fig. 2D: The data representation of "Tubule displacement" image is unclear. The ER tubule indicated by the red arrow does not seem to show any changes over time (like static). time 0 in stamp appears behind the image.
Line 157-158 (and relevant method sections): The definition of static and dynamic boutons is ambiguous. The author should describe in more detail this point including how long they observed the structure to define the changes in ER tubule dynamics.
Fig. 2E: What n=75 and n=29 represent is unclear, are these the number of boutons in en passant and terminal subjected for qualitative analysis?
Fig. 2: What the qualitative analysis represents is unclear, are the points pulled from different experiments?
Line 231: Regarding "...we found a small but significant reduction in dynamic boutons in Atlastin mutants (76%), ...", how do the authors assess significance. If proportion of static/dynamic ER in boutons was obtained from multiple experiments, it should be presented e.g. as in average {plus minus} standard deviation, or clarify that the proportion is representative of x independent experiments.
Line 267-269 and Fig. 6B: The author's conclusion that "Complete loss of ER network structure in NMJ of BiP:sfGFP:HDEL overexpressing Atl mutant" seem to be based on the lack of signal from luminal marker, which may be undetectable due to changes to tubular volume or marker loss to the cytoplasm, as suggested by the authors, while the membranous ER structure is intact. It would be useful to discuss this point and potentially add ER membrane-stained control.
Fig. 6C: In Atl mutant, why does the total of the proportion exceed 100% (10 + 45 + 55)?
Fig. 9C, line 342-344: In FRAP experiment using CD8, it seems that the Partial loss Atl mutant shows slower recovery that control. There seems to be a mismatch in triangle symbols of Partial loss Atl mutant between legend and plot (one is filled and the other is empty). This should be clarified
fig. 10 is a clever way to verify the cytoplasmic localisatoin of the ER marker, however its description and annotation can be improved, and it would be stronger if 4 curves in F for mutant and controls with the trap and normal were shown.
Line 495: Drosophila have ReepA and ReepB, but not Reep1-4. If the authors discuss their speculation based on their observation (using Drosophila), the gene names should be unified in the same species, and explain the corresponding genes to mammalian cells.
Line 548; should UPR be Unfolded Protein Response?
Significance
This study advances the understanding of how ER morphogens affect neuronal cells specifically, the lack of which limits researchers ability to comprehend the neuronal pathologies associated with ER structure-function. The observation on ER content aberrant localisation caused by the lack of key structural protein should be of a great interest for cell and neuronal biologists and researchers of the associated diseases and shows the field a new direction. Though, mechanistic details remain to be unraveled, it constitutes a fundamental, conceptual advance.
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Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.
Learn more at Review Commons
Referee #2
Evidence, reproducibility and clarity
Summary
The endoplasmic reticulum (ER) is a continuous organelle that extends throughout neurons to regulate fundamental processes. The analysis of ER dynamics at synaptic terminals is limited by the challenge of imaging these structures at high resolution. In this manuscript, the authors use super-resolution (~170 nm) live imaging and a combination of membrane and luminal ER markers at the Drosophila larval NMJ, an important model synapse, to investigate dynamic ER architecture in vivo. They report a detailed characterization of the presynaptic ER organization and dynamics at wild-type and GTPase Atlastin mutant NMJs. Their analysis using the ER membrane marker tdTomato:Sec61b reveals the presence of an intact ER network in Atlastin mutants. This contrasts with the apparent ER fragmentation phenotype previously reported and replicated here when using a luminal marker. Their findings instead point to the progressive displacement of luminal proteins to the cytosol in Atlastin mutants specifically at synapses. The authors propose that the disruption of ER protein dynamics at synapses is a compartment-specific ER stress response. The manuscript is well written, results are clearly presented, and experiments are technically rigorous.
Major comments
- The baseline ER phenotypes in Atlastin mutants are mild with complete loss of ER network only observed in terminal boutons. This interesting and unexpected result should be further confirmed by genetic rescue. The authors can use a UAS rescue line previously reported in PMID: 19341724.
- Lines 204-7: It's not clear how a greater coefficient of variation indicates that the marker is more concentrated in subsynaptic structures or what is meant by 'subsynaptic structures.'
- There's a mistake in Figure 6C and the associated text. The summed percentage of the three phenotypic categories adds up to 110% for Atlastin mutants.
- Figure 8: the ER looks fragmented in 1st instar controls and mutants. The authors should address this difference from more mature NMJs.
- The images in figure 9B do not seem representative of the quantification in Figure 9D. Specifically, the partial loss Atlastin NMJ appears to have recovered as fully as the complete loss Atlastin NMJ.
- Optional: An overexpressed luminal marker is displaced to the cytoplasm in Atlastin mutants. It would be interesting to know and increase the significance of the findings if the same is true of endogenous luminal proteins under biological stress conditions.
- Optional: Applying this approach in stimulated conditions (high potassium, increased temperature) might reveal a greater activity-dependent role for Atlastin at synaptic terminals.
Minor Comments
- Line 16: Atlastin should be italicized.
- Figure 5A: Based on the relative intensities, it appears that control and mutant images are not contrast matched but this isn't stated.
- Line 822: The number of static Atlastin mutant boutons used for analysis is missing.
- Figure 9: The blue arrows are not annotated in the figure legend.
Significance
Atlastin is linked to Hereditary Spastic Paraplegia (HSP) and this study changes our understanding of the compartment-specific impacts of its loss. This study reveals the importance of using both membrane and luminal ER markers to accurately interpret phenotypes as well as the importance of considering compartment-specific effects on ER. These findings represent significant mechanistic and conceptual advances. The lack of genetic rescue is a limitation and adding an investigation of an endogenous luminal protein under basal and stress conditions would add significantly to our understanding of Atlastin dysfunction in HSP. Notably, the in vivo imaging approach introduced here can be adapted broadly for live imaging of Drosophila larvae. Thus, this work will be of interest to both neuronal cell biologists and the wider Drosophila community. This review is based on our expertise in neuronal cell biology.
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Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.
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Referee #1
Evidence, reproducibility and clarity
In the present manuscript, the authors address an important question related to the ultrastructure and the dynamics of the ER in HSP. In contrast to previous studies, the authors here show (by using a membrane and luminal protein markers) that in the presynaptic terminals, the overexpressed BIP "mislocalizes" to the cytosol without affecting (or with minimal effect) the integrity of the ER membrane. Although they used an artificial system by overexpressing (overexpression) of BIP-sfGFP-HDEL (fused protein), the findings lack validation of the endogenous protein by biochemical and fluorescent tools.
Concerns:
I am worried about how the article is presented, mainly in the results section, as most of it refers to published data. The Results section is reserved for presenting new findings without external interpretation or comparison. The paper is written mainly as a comparison paper with other studies or relies on previous studies to strengthen their findings rather than coming up with novel findings. Up to figure 4, I missed the relevance of the new findings. The manuscript needs rewriting to emphasize its novelty and significance without comparing it to previous data. Moreover, the manuscript emphasizes the technology and the findings of the localization of the ER luminal proteins to the cytosol (which is not novel and was previously reported in other settings). Those two aspects were not given enough focus. Here are the main major and minor comments:
Major comments:
- Lines 103-116 (first paragraph of the results section) describe mainly published data that is more suitable for the introduction section. It is annoying to refer to different published articles in the Results section to strengthen the results instead of showing them. The same goes for paragraphs two and three. Why mention those data in the Results section if they are already published and known?
- Figure Legends-(in all Figures): The number of experimental repeats must be mentioned in the figure legends.
- The way the figures are labeled is worrisome; supplementary figures are not ordered numerically.
- The tubule extension in Figure 2D is not convincing. Is there a movie showing those changes? Better images are needed. It is essential to show which supplemental movie corresponds to which panel.
- This is unnecessary in the results section: "To investigate the relationship between ER structure and function at synapses, we examined mutants of Atlastin, a GTPase that regulates ER tubule fusion. Drosophila has a single homolog while mammals have three Atlastin homologs, with Atlastin-1 enriched in the brain (Rismanchi et al., 2008)."
- "This reduction in ER membrane marker intensity has also been observed in other HSP mutants, suggesting this is a common feature of ER shaping mutants and could indicate changes in ER membrane composition, integrity, or tubule thickness (P.rez-Moreno et al., 2023)." This comparison is important and should be shown in the same settings as for the Atlastin mutant rather than referring to published data.
- Does the distribution of the luminal ER marker in Figure 6F diffuse due to mislocalization or reflux after being localized to the ER and then refluxed to the cytosol as was previously shown for the ER to Cytosol signaling (ERCYS) mechanism? Could you assess other ER-luminal protein localization biochemically? It is highly recommended to look at another soluble ER-protein localization in the Atlastin mutant without overexpression, which can be an artifact
- "(data not shown)" in line 288. This affects the process of judging those data.
- In comparison to Summerville et al. (2016) in Figure 7, the experiment was not done in the same way. It is important to keep the same settings for comparison
- Does the Atlastin mutant induce the unfolded protein response and stress within the ER? It is necessary to look for UPR markers in those settings. It was shown previously that ER stress leads to protein reflux from the ER to the cytosol. Is there a difference in the ER stress markers in the presynaptic terminal?
- It is important to add biochemical experiments to show that no fragmentation of the ER membrane occurred. It can be simply demonstrated by looking at the redox state of the ER, which would change if it were mixed with the reducing cytosol. Moreover, this can be shown by using an ER-targeted redox-sensitive fluorescent protein that is tethered to the ER membrane to follow changes in the redox state of the ER.
Minor comments:
- It is important to call figures by order. Figure 2C is called before 2A-B. Figure 2B is called before Figure 2A.
- Figure legends (Figure 2): "The same control dataset used in E-G was used in Figure 5 and Figure 5_Supplement." Why is this relevant?
- Figure 4F is called before Figure-4D-E which are not called.
- Figure 5B is called before the previous ones. Same for Figure 5A supplement.
Referees cross-commenting
I agree with the comments raised by reviewer2 and 3. Basically it is highly important to validate those data by genetic rescue. Moreover, it is essential to know the source of the displaced luminal marker to the cytosol. Is it mislocalization or it is a reflux of pre-existing protein to the cytosol after insertion to the ER. It is also recommended by me and the reviewers to test the endogenous protein rather than overexpression.
Significance
General assessment: This interesting paper shows that proteins can escape the ER under special conditions. However, the authors need more evidence to show that and rely less on the overexpression system, especially of BIP-GFP, which can cause proteostasis stress within the ER.
Advance: The results have been oversimplified in their explanations, and some points and complexities of the study need to be addressed further to make the most of them. These are often some of the more interesting concepts in the paper. I think many points can be addressed in the text by the authors being clear and concise with their reporting. At the same time, other experiments would turn this paper from an observational one into a very interesting mechanistic one. This paper is based on previously published articles from the group and other groups, and it is a nice progression. However, as mentioned, this paper depends primarily on published data, and the novelty is somehow lost between all the comparisons to other published data instead of emphasizing that. Without a substantial mechanistic improvement, the paper would remain observatory.
Audience: The microscopy tools can be great addition to researchers in the field to monitor protein trafficking especially Cell biologists (basic research)
My expertise: ER homeostasis, protein trafficking, cell biology
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go.gale.com go.gale.com
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in some of the courses of the program, there was a consistency of microaggressions that I have observed. For example, during discussions in several classes, I observed white students routinely interrupt Black and Brown students and, at times, not hesitate to interrupt white women instructors while they lecture.
Bias in academia favored towards white students
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For example, instructors wanted grad students to engage in discussions about the content of the class. Most times, however, I sat quietly because I was not entirely sure that I was in a safe space. I would remain quiet for several weeks until I was sure that I was in a classroom where professors and students saw me as a person.
Fear of being treated as less than in academic settings based on color
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Fortunately, during my undergraduate years, a Black male professor took me under his wing and mentored me throughout my undergraduate and graduate years of study, giving me sound advice on life and my academic career. The most profound advice he gave me was, "Never apologize for wanting better for yourself."
academic support from those with shared experiences as POC
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I have witnessed Black and Brown students give their perspectives and share personal experiences in the classroom, only to hear the responses of white students who believed they could speak to those same experiences
academic settings being dismissive and not favoring towards POC
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the contagious assumption that we are living in a postracial world is simply an assumption. Universities and colleges pride themselves on inclusion and diversity but do faculty and staff truly understand and embrace diverse student-body members?
key point to reference in Sarah Alvarez reading from week 7
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This essay will focus on inclusion in social spaces, specifically in educational settings where there is the presumption of inclusion, yet my experiences indicate otherwise.
main purpose
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From my teenage years, I have managed to train myself to speak and act in a manner that is nonthreatening to whites, making them feel comfortable, all the while not invoking any stereotypical behavior or speech that would embarrass myself or Black people as a whole.
pressure from society to speak in a standardized way so as to not sound different
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esmodules.com esmodules.com
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The export statement makes functions, objects, or values available to other modules.
function, objects values
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www.jstor.org www.jstor.org
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Code-switching helps us understand how andwhen we (and literary characters) change languagevarieties according to situation, audience, and purpose.
key theme
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hroughout, Joe speaks in the vernacular: "Lemmespeak to mah wife a minute and Ah'm goin' seede man" (Hurston 37). In this way, Joe signals ingroup status by using his speech mirroring the language of those he lead
example of how important code switching can be
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In the United States, standardized English is the variety used by businesses, education,and the government. However, at times, other varieties convey power within a community.
point of interest as to why code switching is so important in society
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If contrastive analysis helps us identify what aspectsof language contrast in dialectally diverse texts,code-switching reveals how characters' languagereflects context.
point to connect with Youngs reading from week 6
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Language comes in varieties (sometimescalled dialects).• All language varieties are structured.• Language varieties are linguistically equal(Adger, Wolfram, and Christian; Wolframand Schilling-Estes)
key points
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When teaching texts that usedifferent varieties of English, we must assume students are unfamiliar with reading that variety, evenif you hear them speak it (and even write it) on aregular basis.
talking point to explore in the paper
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e'll explore how studentscan use language to interrogate issues of identity,power, prestige, and prejudice in society.
Main theme of the reading
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tesda-course.pages.dev tesda-course.pages.dev
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A common trap is to read the slip once and then work from memory. Memory drops a line — most often the middle one. If your reading is rusty, that mistake is normal, not a failure. Keep the slip in front of you and check each line off as you finish it.
same with my previous comment
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tesda-course.pages.dev tesda-course.pages.dev
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StepWhat to do1Put on gloves.2Wipe the bed frame with disinfectant (a cleaning liquid that kills germs).3Let the frame air-dry.4Put on fresh sheets.
Review correctness of the procedure.
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tesda-course.pages.dev tesda-course.pages.dev
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A diagram is a picture with labels that point to its parts.
Can we make use of definition from dictionary? I think this explanation is oversimplified and might not really capture what this is
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A chart, such as a bar chart, shows numbers as bars. A taller bar means a bigger number.
Include other commonly used charts as well, not only bar chart. It would be also helpful to explain when these different charts are used.
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Careful — here is the common trap. The Bread (1 piece) row sits right below the egg row. Read across the wrong row and you answer 5, the price of bread. If that happens, it is rusty reading, not failure. Match the row label to your question first, then slide across
The materials may benefit from a more varied and intentional instructional flow. At present, the lessons appear to follow a very similar structure, which may feel repetitive or overly templated. While a consistent format can support familiarity, the design should also allow for deeper learning, progression, and increasing complexity across activities and examples.
Even the phrases are the same through out "rusty reading" etc.
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Bread (1 piece)5
Double check sample. Buying 1 piece of bread worth 5 pesos is possible in a bakery, but i dont think that is the case for sari sari store. Usually they sell it in pack or bag
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link.springer.com link.springer.com
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Molecular diagnosis of putative Stargardt disease probands by exome sequencing
PMID: 22863181
Gene: ABCA4
HGNC ID: 34
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pubmed.ncbi.nlm.nih.gov pubmed.ncbi.nlm.nih.gov
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Functional Relevance and Structural Correlates of Near Infrared and Short Wavelength Fundus Autofluorescence Imaging in ABCA4-Related Retinopathy
PMID: 31879568
Gene: ABCA4
Disease: ABCA4-Related Retinopathy
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pubmed.ncbi.nlm.nih.gov pubmed.ncbi.nlm.nih.gov
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Monoallelic ABCA4 Mutations Appear Insufficient to Cause Retinopathy: A Quantitative Autofluorescence Study
PMID: 26720470
Gene: ABCA4
Disease: retinopathy
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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Screening of reported pathogenic variants in ABCA4 for Stargardt (STGD) The disease prevalence of STGD is estimated as 1 in 10000 individuals4. It has been estimated that about 70% of STGD patients carry variants in ABCA45. Therefore, this represents the scenario of a recessive disease with a relatively homogeneous genetic cause. We screened 945 reported pathogenic variants in ABCA4 genes collected in HGMD. Among them, 11 variants are likely benign, as their population AF in is higher than 0.7% (1/20000‾‾‾‾‾‾‾‾√)<math xmlns:mml="http://www.w3.org/1998/Math/MathML" display="inline" id="M11"><mrow><mrow><mo>(</mo><mrow><msqrt><mrow><mn>1</mn><mo>/</mo><mn>20000</mn></mrow></msqrt></mrow><mo>)</mo></mrow></mrow></math>, the cutoff based on STGD disease prevalence, therefore were excluded from further analysis. The remaining 934 variants were subjected to our test model. As a result, 26 variants with the AF in the range of 0.46% to 0.03% were identified as likely benign (Binomial test1, Bonferroni correction p-value ≤ 0.05/934 and test2 Bonferroni correction p-value > 0.05/934) (Figure 3A).
This variant is reported in Table S6, but only location, predictions, frequencies, etc are reported for it, not cases.
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Screening of reported pathogenic variants in ABCA4 for Stargardt (STGD) The disease prevalence of STGD is estimated as 1 in 10000 individuals4. It has been estimated that about 70% of STGD patients carry variants in ABCA45. Therefore, this represents the scenario of a recessive disease with a relatively homogeneous genetic cause. We screened 945 reported pathogenic variants in ABCA4 genes collected in HGMD. Among them, 11 variants are likely benign, as their population AF in is higher than 0.7% (1/20000‾‾‾‾‾‾‾‾√)<math xmlns:mml="http://www.w3.org/1998/Math/MathML" display="inline" id="M11"><mrow><mrow><mo>(</mo><mrow><msqrt><mrow><mn>1</mn><mo>/</mo><mn>20000</mn></mrow></msqrt></mrow><mo>)</mo></mrow></mrow></math>, the cutoff based on STGD disease prevalence, therefore were excluded from further analysis. The remaining 934 variants were subjected to our test model. As a result, 26 variants with the AF in the range of 0.46% to 0.03% were identified as likely benign (Binomial test1, Bonferroni correction p-value ≤ 0.05/934 and test2 Bonferroni correction p-value > 0.05/934) (Figure 3A).
This variant is reported in Table S6, but only location, predictions, frequencies, etc are reported for it, not cases.
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www.dovepress.com www.dovepress.com
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Stargardt Disease Due to an Intronic Mutation in the ABCA4: A Case Report
PMID: 36471740
Gene: ABCA4
HGNC ID: 34
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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ABCA4
Unable to find this variant in the text or supplementary even though Mastermind says it's in supplemental MOESM1
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pmc.ncbi.nlm.nih.gov pmc.ncbi.nlm.nih.gov
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F17-003
Case#: Patient 225, Female, age of onset 7 y.o, Poland
DiseaseAssertion: STGD-1
FamilyInfo: no given family information.
CasePresentingHPOs: HP:0007722, HP:0000608, HP:0025158
CaseHPOFreeText: RPE atrophy, macular degeneration, central hyper-autofluorescence in fundus autofluorescence
CaseNotHPOs: n/a
CaseNotHPOFreeText: n/a, non-proband identified HPO's mentioned, but not assignable to individual proband.
Genotyping Method: DNA isolated from peripheral blood from patients and relatives via MagNA Pure 24, samples screened with MIPs targeting 108 genes involved in pathogensis of IRD's. PCR completed on library, analysed with NGS fragment analysis kit.
PreviouslyPublished: yes
Variant: c.[1622T>C;3113C>T]
ClinVar: 99067, 7894
CAID: n/a
gnomeAD 0.0001266 allele frequency
SupplementalData: Fig1: List of families displaying pseudo-dominant inheritance. Fig2: Number of alleles for most common variants.
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www.tandfonline.com www.tandfonline.com
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WDR19-associated retinopathy presenting with adult-onset Stargardt-likephenotype
PMID:39967245
Gene: ABCA4
HGNC ID: 34
Case#:39 man
DiseaseAssertion:NA
FamilyInfo:NA
CasePresentingHPOs:Snellen in both eye, visual impairment with night blindnessisual acuity was 20/20Snellen in both eyes, with a minor correction for astig-matism. The anterior segment and intraocular pressurewere within normal limits. On fundus examination, dif-fuse fleck-like lesions were scattered both inside and out-side the arcades, while sharply demarcated areas ofmacular atrophy with foveal sparing, more pronouncedin the left eye, were visible.
CaseHPOFreeText:NA
CaseNotHPOs:NA
CaseNotHPOFreeText:
Genotyping Method:Next-Generation Sequencing (NGS), using theTruSight One Clinical Exome sequencing panel on anIllumina NexSeq500 platform, enriching for 4800 genesincluding ABCA4, CNGB3, ELOVL4, PROM1, and PRPH2
PreviouslyPublished:Under refernces?
Variant:WDR19 variants:the novel deletion at c.1777 + 1 within the donor splicingsite (class 4) and the rare c.1430 G>T variant causing theamino-acid substitution p.(Arg477Leu) (class 3) in the putative protein. Additionally, a heterozygous c.1793A>G(class 3) variant in the CDH23 gene was found, though it was deemed as not contributive to the patient’s clinical phenotype. All reported variants were confirmed throughSanger sequencing
ClinVar:NA
CAID:NA
SupplementalData:NA
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pmc.ncbi.nlm.nih.gov pmc.ncbi.nlm.nih.gov
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Focal choroidal excavation in Stargardt’s dystrophy
PMID: 32843395
Gene: ABCA4
Disease: Stargardt
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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The STGD patient from Family 12 is a compound heterozygous with p.Val931Met and a novel nonsense mutation at exon 33 (p.Glu1574X; Figure 1B). Disease onset for this patient was at age 43. Ophthalmic examination revealed moderate central retinal changes, decreased mfERG responses exclusively in the central 15 degrees, and decreased visual acuity.
Case#: Family 12 Proband, male, 43yo at onset, Portuguese
DiseaseAssertion: Stargardt
FamilyInfo: no affected family members in pedigree (Fig. 1)
CasePresentingHPOs: HP:0007663
CaseHPOFreeText: "The criteria for STGD phenotype included bilateral central vision loss and pigmentary macular lesions, normal caliber of retinal vessels, absence of pigmented bone spicules, and compatibility with recessive mode of inheritance." Moderate central retinal changes, decreased mfERG responses exclusively in the central 15 degrees
CaseNotHPOs:
CaseNotHPOFreeText:
PreviouslyPublished: n/a
Variant: p.Glu1574X; p.Val931Met. Several other polymorphisms also reported. ABCR400 gene chip microarray, DHPLC
ClinVar: 1460063
CAID: CA341283936
SupplementalData: n/a
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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Personalized genetic counseling for Stargardt disease: Offspring risk estimates based on variant severity
PMID: 35120629
Gene: ABCA4
Disease: Stargardt disease
PAPER USED TO SCORE PS4
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pubmed.ncbi.nlm.nih.gov pubmed.ncbi.nlm.nih.gov
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ABCA4 gene sequence variations in patients with autosomal recessive cone-rod dystrophy
PMID: 12796258
Gene: ABCA4
Disease: autosomal recessive cone-rod dystrophy
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pmc.ncbi.nlm.nih.gov pmc.ncbi.nlm.nih.gov
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Phenotype/genotype correlation in a case series of Stargardt's patients identifies novel mutations in the ABCA4 gene
PMID: 23949494 HGNC: 34 Gene: ABCA4 DiseaseAssertion: Stargardt Disease
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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Mutation scanning and direct DNA sequencing of all 50 exons of ABCR were completed for 150 families segregating recessive Stargardt disease (STGD1). ABCR variations were identified in 173 (57%) disease chromosomes, the majority of which represent missense amino acid substitutions. These ABCR variants were not found in 220 unaffected control individuals (440 chromosomes) but do cosegregate with the disease in these families with STGD1, and many occur in conserved functional domains. Missense amino acid substitutions located in the amino terminal one-third of the protein appear to be associated with earlier onset of the disease and may represent misfolding alleles. The two most common mutant alleles, G1961E and A1038V, each identified in 16 of 173 disease chromosomes, composed 18.5% of mutations identified. G1961E has been associated previously, at a statistically significant level in the heterozygous state, with age-related macular degeneration (AMD). Clinical evaluation of these 150 families with STGD1 revealed a high frequency of AMD in first- and second-degree relatives. These findings support the hypothesis that compound heterozygous ABCR mutations are responsible for STGD1 and that some heterozygous ABCR mutations may enhance susceptibility to AMD.
Annotating here since the full text is a PDF.
Case#: Family AR321 proband, US, 6yo at onset
DiseaseAssertion: Stargardt
FamilyInfo: proband and two other siblings are affected
CasePresentingHPOs: The essential and defining features of STGD were (1) pedigrees with at least one living affected individual compatible with autosomal recessive inheritance; (2) an ophthalmoscopically characteristic retinal disorder in families with both parents living; (3) bilateral central visual loss with both “beaten metal” elliptical foveal dystrophy and temporal pallor of the optic discs, documented by retinal color photography, with or without yellow-pigment epithelial flecks in the macular and/or retinal “near periphery”; and (4) the characteristic fluorescein angiographic feature of a dark choroid (Blacharski 1988).
CaseHPOFreeText:
CaseNotHPOs:
CaseNotHPOFreeText: (1) evidence of autosomal dominant inheritance; (2) any history of night blindness, loss of peripheral vision, or "retinitis pigmentosa"; (3) cataracta complicata or cells in the vitreous; (4) substantially abnormal electroretinographic or electrooculographic responses; (5) no fluorescein angiography performed or no dark choroid documented; (6) neurological disease (including loss of cognition or seizures); 7) drug exposures (especially to antimalarial and agents known to cause crystalline retinopathies); or (8) any "atypical" maculopathies in which a unique diagnosis of STGD could not be established.
PreviouslyPublished: PMID: 8533764
Variant: c.3113C>T p.A1038V; c.1715G>C p.R572P . Heteroduplex and SSCP analyses were used to screen the 50 exons of ABCA4. Linkage analysis and haplotype analysis were previously performed
ClinVar: 99073
CAID: CA226919
SupplementalData: n/a
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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Functional Analysis and Classification of Homozygous and Hypomorphic ABCA4 Variants Associated with Stargardt Macular Degeneration
PMID: 32845050
Gene: ABCA4
Disease: Stargardt Macular Degeneration
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pmc.ncbi.nlm.nih.gov pmc.ncbi.nlm.nih.gov
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JB260 Stargardt ABCA4 c.6119G>A p.Arg2040Gln rs148460146 Zernant et al (2014)50 c.2879del p.Ala960Aspfs*17 N/A
Case#: Bryant Subject JB260, US
DiseaseAssertion: Stargardt
FamilyInfo:
CasePresentingHPOs: "Stargardt disease is a childhood-onset macular degeneration and is most commonly caused by mutations in ABCA4. Characteristic yellow flecks are typically seen under the macula during a fundus exam."
CaseHPOFreeText:
CaseNotHPOs:
CaseNotHPOFreeText:
GenotypingMethod: WES; previously screened using arrayed primer extension (APEX) multigene panels for the relevant disease and no disease-causing variants had been identified; PCR and Sanger for verification
PreviouslyPublished: n/a
Variant: c.6119G>A p.Arg2040Gln; c.2879del p.Ala960Aspfs*17
CAID: CA232815
SupplementalData:
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pubmed.ncbi.nlm.nih.gov pubmed.ncbi.nlm.nih.gov
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Molecular testing for hereditary retinal disease as part of clinical care
PMID: 17296903
Gene: ABCA4
Disease: hereditary retinal disease
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www.e-mjm.org www.e-mjm.org
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An uncommon case of retinitis pigmentosa patients basedon clinical and genetic studyAyudha Bahana Bahana Ilham Perdamaian, MSc2, Dewi Kartikawati Paramita, PhD3, Riris Istighfari Jenie,PhD4, Supanji Supanji, PhD11Universitas Gadjah Mada Fakultas Kedokteran Kesehatan Masyarakat dan Keperawatan, 2Doctorate Program of Health andMedicine Science, Faculty of Medicine, Public Health, and Nurse, Universitas Gadjah Mada, Yogyakarta, Indonesia. Departmentof Ophthalmology, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, 3Department of Histology andMolecular Biology, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta, Indonesia,Integrated Research Laboratory, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakar,4Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Gadjah Mada University, Yogyakarta, IndonesiaCASE REPORTThis article was accepted: 25 August 2024Corresponding Author: Supanji SupanjiEmail: supanji@ugm.ac.id19-An uncommon00304.qxp_3-PRIMARY.qxd 29/08/2024 3:47 PM Page 98
PMID:39215425
Gene: ABCA4
HGNC ID: 34
case27-year-old male, the brother of case 1
DiseaseAssertion: Table 1 The summary of the clinical assessment of IRD patients’ family in this research fro there down they did a whole pannel on the family
Pedigree one can be fore form the beggginnings of case presention section?
CasePresentingHPOs: Case 2, a 27-year-old male, the brother of case 1 had blurry vision which was not corrected with an eyeglass and inconveniences under bright light starting from 14 years ago. Case 2 also underwent a fundus examination after finding that case 1 was RP. In further examination of those patients and their family members found that case 1 was confirmed as RP and case 2
CaseHPOFreeText:NA
CaseNotHPOs:NA
CaseNotHPOFreeText:NA
Genotyping Method:NA
PreviouslyPublished:NA
Variant:NA
ClinVar:
CAID:NA
SupplementalData:NA
Inheritance pattern Autosomal Recessive
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www.ncbi.nlm.nih.gov www.ncbi.nlm.nih.gov
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See Supplementary Table S2 for a complete genotypic glossary of the cohort.
Case#: Patients were identified from the inherited retinal disease (IRD) database at UC San Diego (UCSD).
DiseaseAssertion: RP with macular edema
FamilyInfo:
CasePresentingHPOs:
CaseHPOFreeText: Dx of RP based on "a history of progressive peripheral vision loss or nyctalopia, and ocular examination findings of RP including bone spicule pigmentation, disc pallor and attenuated vessels and genetic confirmation."
CaseNotHPOs:
CaseNotHPOFreeText:
GenotypingMethod: Next-generation sequencing (NGS), exome sequencing, and/or targeted Sanger sequencing were the primary genetic testing approaches.
PreviouslyPublished: PMID:10206579 is referenced but it seems a reference to the variant and not the proband
Variant: c.6383A>G (p.His2128Arg); c.3G>T (p.Met1?). phase unknown
ClinVar: 99455
CAID: CA227399
SupplementalData: Variant is found in table S2
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socialsci.libretexts.org socialsci.libretexts.org
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laughing-bassoon-7p6pm2g.pages.github.io laughing-bassoon-7p6pm2g.pages.github.io
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ems (vs. rubber-stamp
trying this annotation
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hypothes.is hypothes.is
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Create a note by selecting some text and clicking the button
Let's try if this will appeare later on
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socialsci.libretexts.org socialsci.libretexts.org
- Jul 2026
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newsletter.squishy.computer newsletter.squishy.computer
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cloud.google.com cloud.google.com
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LLM Wiki gist. "LLMs don't get bored, don't forget to update a cross-reference, and can touch 15 files in one pass,
LLM Wiki gist
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OKF formalizes the small set of conventions needed to make these patterns interoperable.
make these patterns interoperable
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Just YAML frontmatter
YAML formatter
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www.youtube.com www.youtube.com
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Andrew Krapivin
Andrew Krapivin
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www.yudkowsky.net www.yudkowsky.net
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The Gatekeeper must actually talk to the AI for at least the minimum time set up beforehand. Turning away from the terminal and listening to classical music for two hours is not allowed.
'just say no' would be this (the kind of 'buy in' that is required, and super hard to enforce surely)
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I wish I’d thought to specify this protocol in the earlier tests.
😭
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github.com github.com
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see the User Manual.
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Local-first knowledge app that turns documents, images, and web pages into a connected, searchable, Obsidian-compatible Markdown wiki.
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github.com github.com
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My Second Brain by YFW — User Manual
from: https://hyp.is/dU5m-IjZEfGxfufpohGIAA/github.com/EricWcr7/My-Second-Brain-by-YFW
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gist.github.com gist.github.com
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readable by humans, editable by agents
flip that
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retrieval is still useful, but mainly as a navigation layer
retrieval as navigation layer
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local-first application that turns documents, images, PDFs, and web pages into a connected, searchable, Obsidian-compatible Markdown wiki.
connected searcheable
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JSON-LD
why bother with JSON-LD when you can have IPLD?
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every article keeps its unstructured body and a structured JSON slot side by side
structured JSON slot side by side
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What actually compounds is structure - typed entities
typed structure compound
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the thing that doesn't scale is unqualified links + free text.
Yep
unqualified links and free text
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might want a completely different set of output formats
can do that with IndyWiki
Do It yOurselves
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optional and modular
make it omni optional, holonic, evergreen, co-evolvable co-laborative
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the connections between documents as valuable as the documents themselves.
and if you make these connections intentionally named, interpretable, high resolution, bi-directional, outside as well not just inside the file,
dsicoverable
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Vannevar Bush's Memex (1945) — a personal, curated knowledge store with associative trails between documents.
memex associative trails
except that Bush anvisaged it as interpersonal networked co-laborative and permanent
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Marp is a markdown-based slide deck format.
marked down based slide deck format
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qmd is a good option: it's a local search engine for markdown files with hybrid BM25/vector search and LLM re-ranking, all on-device. I
to: https://hyp.is/P--XZIjWEfGSR5O212_JVg/github.com/tobi/qmd
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log.md is chronological. It's an append-only record of what happened and when
Flip that reverse chronologica prepend only
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Lint. Periodically, ask the LLM to health-check the wiki.
Flip that too
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. Obsidian is the IDE; the LLM is the programmer; the wiki is the codebase.
flip that
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curated collection of source documents. Articles, papers, images, data files.
In IPFS Desktop
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internal wiki maintained by LLMs, fed by Slack threads,
Flip that
Use LLM NoteBook or Gemini Notebook personal LLM
IndyWiki for scaling synthesis Organizing yOur experience of the Frontier of the Net
IndyGram you personal companion to Telegram Fed into your Gemini Notebook
WikiNizer
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github.com github.com
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while being all local
from: https://hyp.is/WfwHUIjWEfG6AhN882Hq6A/gist.github.com/karpathy/442a6bf555914893e9891c11519de94f
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muse-image.co muse-image.co
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A Five-Step Audit Trail for AI-Assisted Visual Assets
If you've ever handed off a design file and gotten the question "wait, is this AI-generated?" you know how awkward it is to reconstruct an answer after the fact. Prompt text gets lost in chat history, reference images get pulled from five different folders, and nobody remembers which revision fixed the extra finger. A lightweight audit habit, done at creation time, saves that scramble.
Here's a five-step version worth keeping next to your project files:
- Prompt text. Save the literal prompt used, not a paraphrase, in a plain text file alongside the output. Include the model or tool name and date.
- Reference-image provenance. Note where any reference images came from — licensed stock, your own photography, a client asset — and whether you had rights to use them as input.
- Revision intent. When you edit or regenerate, write one line on why: "changed lighting to match brand palette," not just "v2." This turns a folder of near-duplicates into a readable history.
- Output checks. Record what you verified before shipping — text legibility, anatomical errors, brand color accuracy, resolution for print versus web.
- Disclosure. Decide in advance whether the final asset needs an AI-assisted label for the audience it's going to, and note that decision.
None of this requires special software; a shared doc or spreadsheet works fine. The value is in doing it consistently, not in the tool.
If you're working in a platform built around iterative prompting and multi-reference composition, like Muse Image, the same five fields map cleanly onto its workflow — prompt history, reference inputs, and revision passes are already things you're generating, so the audit is mostly about capturing what you did rather than adding new work.
The limitation is that no checklist replaces judgment: you still have to actually look at the output, and you still have to decide what disclosure means for your context. But a five-minute habit beats a reconstructed memory every time.
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ipfs.indy0.net ipfs.indy0.net()1
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APPROACHES TO ARCHIVING PROFESSIONAL BLOGSHOSTED IN THE CLOUD
Brian Kelly and Marieke Guy UKOLN, University of Bath, Claverton Road
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www.google.com www.google.com
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www.researchgate.net www.researchgate.net
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Parallel Turing Machine, a Proposal
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en.wikipedia.org en.wikipedia.org
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Toggle the table of contents Contents move to sidebar hide (Top) 1 Introduction 2 Mathematical theory 3 Efficiency 4 Smallest machines 5 Machines with no internal states 6 Example of coding 7 See also 8 Notes 9 References Toggle References subsection 9.1 Footnotes 9.2 Original paper and correction 9.3 Other works cited 10 Further reading 11 External links Universal Turing machine
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54.241.53.176 54.241.53.176
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RRID:AB_477010
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RRID:AB_3678889
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SciCrunch record: RRID:AB_3678889
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SciCrunch record: RRID:AB_2050678
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SciCrunch record: RRID:AB_3678888
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RRID:AB_3711244
DOI: 10.1186/s40478-026-02240-y
Resource: RRID:AB_3711244
Curator: @scibot
SciCrunch record: RRID:AB_3711244
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RRID:AB_10013382
DOI: 10.1186/s40478-026-02240-y
Resource: (Agilent Cat# Z0334, RRID:AB_10013382)
Curator: @scibot
SciCrunch record: RRID:AB_10013382
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www.sciencedirect.com www.sciencedirect.com
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RRID:AB_777165
DOI: 10.1016/j.stem.2026.07.003
Resource: (Abcam Cat# ab32570, RRID:AB_777165)
Curator: @scibot
SciCrunch record: RRID:AB_777165
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RRID:AB_1523910
DOI: 10.1016/j.stem.2026.07.003
Resource: (Abcam Cat# ab64693, RRID:AB_1523910)
Curator: @scibot
SciCrunch record: RRID:AB_1523910
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RRID:AB_10975465
DOI: 10.1016/j.stem.2026.07.003
Resource: (Abcam Cat# ab125212, RRID:AB_10975465)
Curator: @scibot
SciCrunch record: RRID:AB_10975465
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54.241.53.176 54.241.53.176'-html.html13
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AB_330248
DOI: 10.1016/j.isci.2026.115716
Resource: (Cell Signaling Technology Cat# 3671, RRID:AB_330248)
Curator: @scibot
SciCrunch record: RRID:AB_330248
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AB_2534079
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Resource: (Thermo Fisher Scientific Cat# A-11012, RRID:AB_2534079)
Curator: @scibot
SciCrunch record: RRID:AB_2534079
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AB_2249358
DOI: 10.1016/j.isci.2026.115716
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Curator: @scibot
SciCrunch record: RRID:AB_2249358
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AB_561053
DOI: 10.1016/j.isci.2026.115716
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Curator: @scibot
SciCrunch record: RRID:AB_561053
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AB_2798136
DOI: 10.1016/j.isci.2026.115716
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SciCrunch record: RRID:AB_2798136
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AB_2800199
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Curator: @scibot
SciCrunch record: RRID:AB_2800199
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AB_2534069
DOI: 10.1016/j.isci.2026.115716
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Curator: @scibot
SciCrunch record: RRID:AB_2534069
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AB_10839118
DOI: 10.1016/j.isci.2026.115716
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Curator: @scibot
SciCrunch record: RRID:AB_10839118
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AB_10013641
DOI: 10.1016/j.isci.2026.115716
Resource: (Cell Signaling Technology Cat# 6943, RRID:AB_10013641)
Curator: @scibot
SciCrunch record: RRID:AB_10013641
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AB_2174466
DOI: 10.1016/j.isci.2026.115716
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Curator: @scibot
SciCrunch record: RRID:AB_2174466
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AB_2160882
DOI: 10.1016/j.isci.2026.115716
Resource: (Cell Signaling Technology Cat# 3528, RRID:AB_2160882)
Curator: @scibot
SciCrunch record: RRID:AB_2160882
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AB_477629
DOI: 10.1016/j.isci.2026.115716
Resource: (Sigma-Aldrich Cat# V9131, RRID:AB_477629)
Curator: @scibot
SciCrunch record: RRID:AB_477629
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AB_2291558
DOI: 10.1016/j.isci.2026.115716
Resource: RRID:AB_2291558
Curator: @scibot
SciCrunch record: RRID:AB_2291558
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