The DNA damage response (DDR) preserves genomic integrity. Small non-coding RNAs termed DDRNAs are generated at DNA double-strand breaks (DSBs) and are critical for DDR activation. Here we show that active DDRNAs specifically localize to their damaged homologous genomic sites in a transcription-dependent manner. Upon DNA damage, RNA polymerase II (RNAPII) binds to the MRE11/RAD50/NBS1 complex, is recruited to DSBs and synthesizes damage-induced long non-coding RNAs (dilncRNAs) from and towards DNA ends. DilncRNAs act both as DDRNA precursors and by recruiting DDRNAs through RNA:RNA pairing. Together dilncRNAs and DDRNAs fuel DDR focus formation and associate with 53BP1. Accordingly, inhibition of RNAPII prevents DDRNA recruitment, DDR activation and DNA repair. Antisense oligonucleotides matching dilncRNAs and DDRNAs impair site-specific DDR focus formation and DNA repair. We propose that DDR signalling sites, in addition to sharing a common pool of proteins, individually host a unique set of site-specific RNAs necessary for DDR activation.

a b s t r a c tOrgan size is controlled by the concerted action of biochemical and physical processes. Although mechanical forces are known to regulate cell and tissue behavior, as well as organogenesis, the precise molecular events that integrate mechanical and biochemical signals to control these processes are not fully known. The recently delineated Hippo-tumor suppressor network and its two nuclear effectors, YAP and TAZ, shed light on these mechanisms. YAP and TAZ are proto-oncogene proteins that respond to complex physical milieu represented by the rigidity of the extracellular matrix, cell geometry, cell density, cell polarity and the status of the actin cytoskeleton. Here, we review the current knowledge of how YAP and TAZ function as mechanosensors and mechanotransducers. We also suggest that by deciphering the mechanical and biochemical signals controlling YAP/TAZ function, we will gain insights into new strategies for cancer treatment and organ regeneration.

Physical forces in the form of substrate rigidity or geometrical constraints have been shown to alter gene expression profile and differentiation programs. However, the underlying mechanism of gene regulation by these mechanical cues is largely unknown. In this work, we use micropatterned substrates to alter cellular geometry (shape, aspect ratio, and size) and study the nuclear mechanotransduction to regulate gene expression. Genome-wide transcriptome analysis revealed cell geometry-dependent alterations in actin-related gene expression. Increase in cell size reinforced expression of matrix-related genes, whereas reduced cell-substrate contact resulted in up-regulation of genes involved in cellular homeostasis. We also show that large-scale changes in geneexpression profile mapped onto differential modulation of nuclear morphology, actomyosin contractility and histone acetylation. Interestingly, cytoplasmic-to-nuclear redistribution of histone deacetylase 3 modulated histone acetylation in an actomyosin-dependent manner. In addition, we show that geometric constraints altered the nuclear fraction of myocardin-related transcription factor. These fractions exhibited hindered diffusion time scale within the nucleus, correlated with enhanced serum-response element promoter activity. Furthermore, nuclear accumulation of myocardin-related transcription factor also modulated NF-κB activity. Taken together, our work provides modularity in switching gene-expression patterns by cell geometric constraints via actomyosin contractility.cell matrix interaction | substrate geometry | MRTF-A signaling | chromatin remodelling | transcription control C ells within the local tissue microenvironment acquire nonrandom geometrical organization by cell-matrix and cell-cell interaction. Cellular geometry has been shown to influence nuclear deformation, cytoskeleton reorganization, chromatin compaction, gene expression, growth, apoptosis, and cell division (1-7). Other physical cues such as substrate stretching, fluid flow, substrate rigidity, and cellular topography have also been shown to alter cellular morphology, nuclear architecture, and gene expression (8-11). Regulation of gene expression requires posttranslational modifications of histone tails (12), which alter higher-order chromatin assembly and, hence, the accessibility of gene-regulatory sites by transcriptional machinery (13). In addition, cytoplasmic to nuclear shuttling of transcription factors (TFs) and cofactors are key signaling intermediates rendering specificity. Some of these factors include NF-κB, STAT, and myocardin-related transcription factor (MRTF-A) (14-16). The transcription coactivator yes-associated protein (YAP)/transcription coactivator with PDZ binding domain (TAZ) and MRTF-A have been implicated in nuclear mechanotransduction (17)(18)(19). In a recent study, alterations in cell shape were shown to influence mesenchymal stem cell differentiation (20). However, the mechanisms underlying geometric control of gene expression by the modulation of cytoplasmi...

It is well established that cells sense chemical signals from their local microenvironment and transduce them to the nucleus to regulate gene expression programmes. Although a number of experiments have shown that mechanical cues can also modulate gene expression, the underlying mechanisms are far from clear. Nevertheless, we are now beginning to understand how mechanical cues are transduced to the nucleus and how they influence nuclear mechanics, genome organization and transcription. In particular, recent progress in super-resolution imaging, in genome-wide application of RNA sequencing, chromatin immunoprecipitation and chromosome conformation capture and in theoretical modelling of 3D genome organization enables the exploration of the relationship between cell mechanics, 3D chromatin configurations and transcription, thereby shedding new light on how mechanical forces regulate gene expression.

Cells sense mechanical signals from their microenvironment and transduce them to the nucleus to regulate gene expression programs. To elucidate the physical mechanisms involved in this regulation, we developed an active 3D chemomechanical model to describe the three-way feedback between the adhesions, the cytoskeleton, and the nucleus. The model shows local tensile stresses generated at the interface of the cell and the extracellular matrix regulate the properties of the nucleus, including nuclear morphology, levels of lamin A,C, and histone deacetylation, as these tensile stresses 1) are transmitted to the nucleus through cytoskeletal physical links and 2) trigger an actomyosin-dependent shuttling of epigenetic factors. We then show how cell geometric constraints affect the local tensile stresses and subsequently the three-way feedback and induce cytoskeleton-mediated alterations in the properties of the nucleus such as nuclear lamina softening, chromatin stiffening, nuclear lamina invaginations, increase in nuclear height, and shrinkage of nuclear volume. We predict a phase diagram that describes how the disruption of cytoskeletal components impacts the feedback and subsequently induce contractility-dependent alterations in the properties of the nucleus. Our simulations show that these changes in contractility levels can be also used as predictors of nucleocytoplasmic shuttling of transcription factors and the level of chromatin condensation. The predictions are experimentally validated by studying the properties of nuclei of fibroblasts on micropatterned substrates with different shapes and areas.