"Studies of cellular differentiation in simulated microgravity reveal an important role for β-actin in mechanosensing
Authors: Oscar Sapkota, Tomas Venit and Piergiorgio Percipalle
Science Division, Biology Program, New York University Abu Dhabi, Abu Dhabi, UAE
Mechanotransduction is mediated by the actin cytoskeleton and it is important in helping stem cells determine their fate. Differentiating cells assess changes in mechanical stressors in the extracellular matrix (ECM) and transduce them into a reaction cascade to regulate genes involved in cellular lineage specification. During differentiation, the nuclear β-actin pool plays an important role in regulating gene expression as part of the chromatin remodeling complex BAF and bound to all three eukaryotic RNA polymerases. Based on these considerations, the aim of this study is to investigate whether the nuclear β-actin pool is involved in regulating gene expression that controls mechanosensory pathways in response to mechanical unloading in the ECM.
Methodology and results
Wild-type (WT) and β-actin knock-out (KO) mouse embryonic fibroblasts (MEFs) were subjected to simulated microgravity (sµG) for 72 hours using a Random Positioning Machine (RPM) to study changes in their global epigenetic landscape. This was achieved by staining MEFs with antibodies against active, repressive, and enhancer-associated histone marks and analyzing their intensity and distribution inside cell nuclei using the Cellomics CX7 Laser High Content Phenotypic Platform. Results show an increase in repressive chromatin marks (H3K9Me3, Hp1a) and dysregulated levels of enhancer-specific epigenetic marks (H3K27Ac, H3K4Me) in the β-actin KO but not in the WT cultured in sµG. This confirms a role of β-actin in regulating heterochromatin and hints to a novel role for the nuclear beta actin pool in the regulation of enhancers to transduce microgravity-induced changes. To examine this further, total RNA was extracted from WT and β-actin KO MEFs cultured in sµG for 24 and 72 hours and subjected to RNAseq. Analysis of the transcriptomic data obtained shows upregulation of genes involved in focal adhesion, osteogenesis, and axon guidance in the WT under sµG, but not in the KO.
Considering the observed epigenetic and transcriptomic changes observed upon β-actin depletion, we propose that β-actin regulates expression of genes involved in mechanosensing. We speculate that β-actin performs this novel function by controlling heterochromatin levels and potentially impacting on enhancers’ activity."