Mechanotransduction at the nuclear envelope: the role of forces in facilitating embryonic stem cell fate decisions

While a large body of work has focused on the transcriptional regulation of cellular identity, the role of the mechanical properties of cells and the importance of their physical interactions with the local environment remains less well understood. In this project, we explored the impact of cytoskeleton-generated forces exerted on the nucleus in the context of early embryonic stem (ES) cell fate decisions. We chose to perturb force generating components in the cytoskeleton – notably the molecular motor non-muscle myosin II - and key structural and chromatin binding proteins in the nuclear envelope, notably, the lamins (LMNA), Lamin B receptor (LBR) and components of the LINC complex (nesprins/KASH). The structural proteins in the nuclear envelope regulate both the mechanical response of the nucleus to force and the stabilization of peripheral heterochromatin (repressed genes). Our hypothesis is that reducing forces transmitted directly to chromatin or increasing tethering of peripheral heterochromatin to the nuclear envelope would restrict access to lineage specific genes sequestered at the nuclear lamina and thereby either impair, or delay, differentiation. We found phenotypes in the capacity of mouse ES cells to specify to the neural lineage following our perturbations: overexpression of LMNA, LBR and KASH proteins resulted in a significant fraction of cells that did not express the neuroectoderm marker Sox1 after four days of differentiation, while inhibiting non-muscle myosin II delayed Sox1 expression in the entire population. Overexpression of LMNA and LBR did not affect the ability of the cells to exit the naive pluripotent state, which raises the possibility that the perturbations are halting the cells in a formative phase prior to lineage specification. Future work will focus on looking at genome-wide transcriptional changes accompanying differentiation combined with an analysis of spatial information of differentially regulated genes.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2017

Author dc:creator

• Wylde, George William

Advisor dc:contributor.advisor

• Chalut, Kevin J

• Biophysics
• Mechanotransduction
• Embryonic
• Stem cell
• Forces
• Fate
• Shape change
• Nucleus
• Lamin
• LINC complex
• Nuclear envelope
• Chromatin
• Fluctuation

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/2f865327-9880-46c3-990d-ebab74324c47/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

en

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.13845

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/267909