Phase separation of YAP reorganizes genome topology for long-term YAP target gene expression

Abstract

Yes-associated Protein (YAP) is a transcriptional co-activator that regulates cell proliferation and survival by binding to a select set of enhancers for target gene activation. How YAP coordinates these transcriptional responses is unknown. Here, we demonstrate that YAP forms liquid-like condensates in the nucleus. Formed within seconds of hyperosmotic stress, YAP condensates compartmentalized YAP’s transcription factor TEAD1 and other YAP-related co-activators, including TAZ, and subsequently induced transcription of YAP-specific proliferation genes. Super-resolution imaging using Assay for Transposase Accessible Chromatin with photoactivated localization microscopy (ATAC-PALM) revealed that YAP nuclear condensates were areas enriched in accessible chromatin domains organized as super-enhancers. Initially devoid of RNA Polymerase II (RNAPII), the accessible chromatin domains later acquired RNAPII, transcribing RNA. Removal of YAP’s intrinsically-disordered transcription activation domain (TAD) prevented YAP condensate formation and diminished downstream YAP signaling. Thus, dynamic changes in genome organization and gene activation during YAP reprogramming is mediated by liquid-liquid phase separation.

Multivalent proteins rapidly and reversibly phase-separate upon osmotic cell volume change

“…Perhaps the most striking aspect of HOPS is its rapid onset, which is faster than the speed of canonical stress responses . This feature is similar to recent reports of rapid nuclear condensation of DEAD-box RNA helicase DDX4 in response to environmental stress (Nott et al, 2015), and of transcriptional co-activator YAP in response to hyperosmotic stress (Cai et al, 2018). Notably, prolonged exposure to hyperosmotic conditions, similar to other environmental stressors, triggers the ISR and subsequent assembly of SGs, often localized adjacently to pre-formed DCP1A condensates or PBs (Figure 1) (Kedersha et al, 2005).…”

“…Both rapid and prolonged HOPS are reversible ( Figure 1) and can mediate widespread effects, including impairment of transcription termination ( Figure 6) (Vilborg et al, 2015), YAP-programmed transcription initiation (Cai et al, 2018), inhibition of ribosomal translocation (Wu et al, 2019), and modulation of RNA silencing (Pitchiaya et al, 2019). While other mechanisms may be also at play, protein sequestration away from the site of their function provides a straightforward biophysical explanation for many of these effects ( Figure 6).…”

Mechanical memory in cells emerges from mechanotransduction with transcriptional feedback and epigenetic plasticity

“…Cytoskeletal structure and forces are now known to regulate nuclear transport of mechanosensitive transcriptional activators such as YAP and MRTF-A (23,44,45). Previously, YAP has been shown to undergo phase-mediated reorganization within nucleus and cytoplasm, which leads to upregulation of YAP-related transcription factors, such as TEAD1 (46). Upregulated YAP levels have also been shown to predict higher contractility in cancer associated fibroblasts, leading to matrix stiffening and cancer cell invasion (40).…”

Mask family proteins ANKHD1 and ANKRD17 regulate YAP nuclear import, stability and phase separation

“…These results suggest that Mask proteins can cluster/polymerise YAP to drive colloidal phase separation -a classic mechanism of cellular compartmentalisation (Hardy, 1899;Iborra, 2007;Walter and Brooks, 1995;Wilson, 1899). Recent work has shown that cells expressing GFP-tagged YAP can exhibit colloidal phase separation and formation of YAP liquid droplets in both nucleus and cytoplasm after treatment with 25% PEG to induce macromolecular crowding (Cai et al, 2018). Our findings indicate that endogenous YAP can also phase separate when it becomes stabilised and concentrated by clustering with Mask family proteins.…”

“…An interesting consequence of the role of Mask1/2 in stabilising YAP protein is that overexpression of Mask proteins is sufficient to dramatically raise the concentration of YAP within the cytoplasm, even causing formation of colloidal YAP liquid droplets in human cells. Similar YAP liquid droplets have recently been shown to occur by phase separation after treatment of cells expressing GFP-tagged YAP with 25% PEG to drive macromolecular crowding (Cai et al, 2018). Whether phase-separation of YAP has a physiological role is still unknown, but the possibility is supported by the fact that endogenous YAP can also be observed to phase separate when in a complex with Mask1 at high concentrations.…”