Review coordinated by Life Science Editors Foundation Reviewed by: Dr. Angela Andersen, Life Science Editors Foundation & Life Science Editors. Potential Conflicts of Interest: None.

PUNCHLINE: Chromatin organization, orchestrated by the epigenetic reader MeCP2, governs nuclear stiffness in a concentration- and differentiation-dependent manner—providing a mechanistic link between heterochromatin compaction, mechanotransduction, and the severity of Rett syndrome phenotypes.

BACKGROUND: Nuclear mechanics are critical for how cells sense and respond to physical forces, yet most attention has focused on the cytoskeleton and lamin network. Chromatin, particularly heterochromatin, has been considered a secondary contributor, despite its dominant nuclear occupancy. MeCP2, a methyl-CpG-binding protein abundantly expressed in neurons and mutated in Rett syndrome, is known to cluster heterochromatin and modulate chromatin structure. Rett syndrome mutations impact MeCP2's binding and chromatin compaction abilities, but changes in gene expression do not strongly correlate with disease severity. This study proposes an alternative hypothesis: MeCP2 mutations impair the physical properties of the nucleus via disorganized chromatin architecture, offering a new framework to understand the mechanobiology of neuronal development and disease.

QUESTIONS ADDRESSED:

How does MeCP2 concentration influence nuclear stiffness, and is this linked to chromatin compaction?

Do Rett syndrome mutations disrupt MeCP2’s role in nuclear mechanics?

Is chromatin-mediated nuclear stiffness regulated independently of canonical mechanotransduction gene expression?

SUMMARY: Using atomic force microscopy (AFM) to directly measure nuclear stiffness in purified nuclei, the authors show that MeCP2 levels strongly correlate with increased nuclear stiffness. MeCP2 overexpression in myoblasts leads to heterochromatin clustering and ~15–20x increases in nuclear stiffness. During neural differentiation, wild-type cells exhibit dramatic stiffening of nuclei, which is largely abolished in MeCP2 knockout cells. Rett syndrome mutations, including R106W and T158M, differentially impair this function—with T158M inducing nuclear softening even below baseline. Importantly, these mechanical changes occur without global alterations in expression of mechanosensitive genes, implicating chromatin structure itself as a mechanical determinant.

KEY RESULTS

Chromatin Stiffness Is Cytoskeleton-Independent Nuclei purified from cells retain stiffness comparable to the nuclear region of intact cells, showing that chromatin contributes autonomously to nuclear mechanics. In the absence of cytoskeletal components, MeCP2-dependent changes remain robust.

MeCP2 Induces Heterochromatin Compaction and Increases Nuclear Stiffness MeCP2 clustering activity scales with concentration: untransfected myoblasts show 1.4 kPa stiffness, while MeCP2-overexpressing nuclei reach 23.5 kPa. Heterochromatin becomes fewer in number but larger in volume, indicating fusion and compaction.

MeCP2 Is Required for Nuclear Stiffening During Neural Differentiation Differentiation of ESCs into neurons leads to a ~10x increase in nuclear stiffness in wild-type cells, but not in MeCP2 knockouts. NSCs and neurons from KO mice show both impaired heterochromatin clustering and lower stiffness, especially at timepoints when MeCP2 expression peaks in wild-type neurons.

Rett Syndrome Mutations Impair MeCP2-Dependent Stiffening Of 9 Rett-linked mutations tested, several (e.g., R106W, T158M) failed to increase nuclear stiffness, clustering with untransfected controls. Other variants (e.g., A140V) retained or exaggerated MeCP2-like effects, correlating with milder phenotypes.

Mechanostiffness Is Not Driven by Mechanotransduction Gene Expression RNA-seq and qPCR reveal only minor changes in mechanotransduction-related genes (e.g., Tgfbr1, Notch2), and ChIP-seq does not show MeCP2 binding at these loci—supporting a model where stiffness arises from structural chromatin effects, not transcriptional changes.

STRENGTHS:

Direct mechanical measurements using AFM in purified nuclei across multiple cell states.

Dissects MeCP2 function independently of its transcriptional effects.

Uses Rett syndrome mutants to connect biophysics to disease severity.

Establishes chromatin structure as an autonomous determinant of nuclear stiffness.

Integrates epigenetics, mechanics, and disease in a novel conceptual framework.

FUTURE WORK:

Can modulating MeCP2 levels or chromatin compaction rescue mechanical defects in Rett models?

Do neurons use MeCP2-mediated stiffness to regulate mechanosensitive gene expression or signaling?

Are similar chromatin-stiffness mechanisms active in other cell types or diseases?

Could small molecules targeting chromatin modifiers restore nuclear mechanics in disease?

FINAL TAKEAWAY: This study redefines the role of chromatin—particularly MeCP2-organized heterochromatin—as a critical regulator of nuclear stiffness during neuronal differentiation. By decoupling mechanical properties from transcriptional changes, it provides a mechanistic explanation for how MeCP2 mutations contribute to the pathophysiology of Rett syndrome. These findings suggest that chromatin organization is not merely a regulator of gene expression but also a physical architect of the cell’s mechanical identity.

Mixing does occur between chromosome regions. Not as distinct as first thought.

Subnuclear structure: nuclear bodies * typically spherical<br /> * PML bodies (promyelocytic leukemia protein), involved in transcriptional regulation/cell division * Cleavage bodies * Cajal bodies (CB) * antibody against coilin * Gems * antibody against SMN * CB and gems co-localise with exposure/differentiation to RA. Vary with cell type. * Nuclei of SMA afflicted foetuses lose gems.

Subnuclear structure: Nuclear speckles, not a blanket term. * Also called interchromatin granule clusters * Splicing factor storage (for splicing factors (mostly) not in use) * Proteins may move out of or enter nuclear speckles.

FRAP as a method to test dynamism, movement. Diffusion vs no diffusion of bleached protein. * Mobile fraction * Immobile fraction

The fibrillar centre is where genes for rRNA are transcribed. The dense fibrillar component where rRNA is processed, chemically modified. The granular component is where protein components are combined with rRNA. Generates preribosomal molecules that are close to being exported to cytoplasm.

mRNA needs to be assisted across the NPC. Like protein, also classed as facilitated diffusion. * mRNP exporter combines with mRNA with poly A tail, by interacting with FG repeats * mRNA moves through NPC * Dpb5 is an RNA helicase * Straightens the mRNA secondary structure and allows passage, removes proteins on the strand (NXT1, NXF1) * mRNP exporter proteins dissociate from the mRNA. * mRNA is now in cytoplasm

Was the type specimen for Cassidula nucleus (Gmelin, 1791) collected by James Cook (1728-1779)? The type specimen with catalog number NHMD-155242 is collected at Tahiti and James Cook visited Tahiti during all of his three voyages (1768-1771; 1772-1775; 1776-1779).

Cassidula nucleus (Gmelin, 1791) https://www.gbif.org/species/7932657 https://en.wikipedia.org/wiki/Cassidula_nucleus http://www.wikidata.org/entity/Q3695551 http://www.marinespecies.org/aphia.php?p=taxdetails&id=549377 http://www.marinespecies.org/aphia.php?p=taxdetails&id=882010

original description (of Helix nucleus Gmelin, 1791) Gmelin J.F. (1791). Vermes. In: Gmelin J.F. (Ed.) Caroli a Linnaei Systema Naturae per Regna Tria Naturae, Ed. 13. Tome 1(6). G.E. Beer, Lipsiae [Leipzig]. pp. 3021-3910. , available online at http://www.biodiversitylibrary.org/item/83098#5 page(s): 3193

Original description of Helix nucleus Gmelin, 1791: "Nucleus. 255. H. tefta imperforata ovata glauca transverfim ftriata: cingulis atris, apertura finuola. Martin univ. Conch. 2. t. 67. fig. exter. Habitat in Tahiti."

The first voyage (1768–1771) of James Cook arrived at Tahiti on 13 April 1769. The second voyage (1772–1775) of James Cook also landed at Tahiti to resupply in 1774. And again during his third voyage (1776–1779). https://en.wikipedia.org/wiki/James_Cook

Systema Naturae was originally published in 1735. But does not include Helix nucleus. The 10th edition 1758 does not include Helix nucleus. The 12 edition (1766-68) and last edited by Carl Linnaeus (1707-1778) does not include Helix nucleus. While the 13th edition edited by Johann Friedrich Gmelin between 1788 and 1793 does include Helix nucleus in volume 1 part 1 published in July 1788. https://en.wikipedia.org/wiki/Systema_Naturae

The type specimen held at Zoological museum of the the Natural History Museum of Denmark with catalog number NHMD-155242 is indicated as collected by James Cook at Tahiti. As the type specimen for Helix nucleus Gmelin, 1791, it must have been collected before 1791.

https://www.gbif.org/occurrence/2012930732 https://www.gbif.org/occurrence/2012930732#annotations:_zXnVoS2EeuTD5vLxRG34Q

Was the type specimen for Cassidula nucleus (Gmelin, 1791) collected by James Cook (1728-1779)? The type specimen with catalog number NHMD-155242 is collected at Tahiti and James Cook visited Tahiti during all of his three voyages (1768-1771; 1772-1775; 1776-1779).

Cassidula nucleus (Gmelin, 1791) https://www.gbif.org/species/7932657 https://en.wikipedia.org/wiki/Cassidula_nucleus http://www.wikidata.org/entity/Q3695551 http://www.marinespecies.org/aphia.php?p=taxdetails&id=549377 http://www.marinespecies.org/aphia.php?p=taxdetails&id=882010

original description (of Helix nucleus Gmelin, 1791) Gmelin J.F. (1791). Vermes. In: Gmelin J.F. (Ed.) Caroli a Linnaei Systema Naturae per Regna Tria Naturae, Ed. 13. Tome 1(6). G.E. Beer, Lipsiae [Leipzig]. pp. 3021-3910. , available online at http://www.biodiversitylibrary.org/item/83098#5 page(s): 3193

Original description of Helix nucleus Gmelin, 1791: "Nucleus. 255. H. tefta imperforata ovata glauca transverfim ftriata: cingulis atris, apertura finuola. Martin univ. Conch. 2. t. 67. fig. exter. Habitat in Tahiti."

The first voyage (1768–1771) of James Cook arrived at Tahiti on 13 April 1769. The second voyage (1772–1775) of James Cook also landed at Tahiti to resupply in 1774. And again during his third voyage (1776–1779). https://en.wikipedia.org/wiki/James_Cook

Systema Naturae was originally published in 1735. But does not include Helix nucleus. The 10th edition 1758 does not include Helix nucleus. The 12 edition (1766-68) and last edited by Carl Linnaeus (1707-1778) does not include Helix nucleus. While the 13th edition edited by Johann Friedrich Gmelin between 1788 and 1793 does include Helix nucleus in volume 1 part 1 published in July 1788. https://en.wikipedia.org/wiki/Systema_Naturae

The type specimen held at Zoological museum of the the Natural History Museum of Denmark with catalog number NHMD-155242 is indicated as collected by James Cook at Tahiti. As the type specimen for Helix nucleus Gmelin, 1791, it must have been collected before 1791.

https://www.gbif.org/occurrence/2012930732 https://www.gbif.org/occurrence/2012930732#annotations:_zXnVoS2EeuTD5vLxRG34Q

This is the process of a complementary mRNA copy of a single gene on the DNA that is created in the nucleus. The mRNA is smaller than the DNA so it can carry the genetic code into the ribosome and into the cytoplasm that enables the protein creation.

This is a single strand on an RNA molecule that leaves the the nucleus of a cell in order to relocate to the cytoplasm. This is where the mRNA can help create the protein for the cell in a process known as protein synthesis. The mRNA takes in information passed into it by DNA and decode it for the ribosomes to make more protein for the cell to live on.