RASSF1A–LATS1 signalling stabilizes replication forks by restricting CDK2-mediated phosphorylation of BRCA2

Genomic instability is a key hallmark of cancer leading to tumour heterogeneity and therapeutic resistance. ​BRCA2 has a fundamental role in error-free DNA repair but also sustains genome integrity by promoting ​RAD51 nucleofilament formation at stalled replication forks. ​CDK2 phosphorylates ​BRCA2 (pS3291-​BRCA2) to limit stabilizing contacts with polymerized ​RAD51; however, how replication stress modulates ​CDK2 activity and whether loss of pS3291-​BRCA2 regulation results in genomic instability of tumours are not known. Here we demonstrate that the Hippo pathway kinase ​LATS1 interacts with ​CDK2 in response to genotoxic stress to constrain pS3291-​BRCA2 and support ​RAD51 nucleofilaments, thereby maintaining genomic fidelity during replication stalling. We also show that ​LATS1 forms part of an ​ATR-mediated response to replication stress that requires the tumour suppressor ​RASSF1A. Importantly, perturbation of the ​ATR–​RASSF1A–​LATS1 signalling axis leads to genomic defects associated with loss of ​BRCA2 function and contributes to genomic instability and ‘BRCA-ness’ in lung cancers

RASSF1A inhibits PDGFB-driven malignant phenotypes of nasopharyngeal carcinoma cells in a YAP1-dependent manner.

Nasopharyngeal carcinoma (NPC) is a highly aggressive tumor characterized by distant metastasis. Deletion or down-regulation of the tumor suppressor protein ras-association domain family protein1 isoform A (RASSF1A) has been confirmed to be a key event in NPC progression; however, little is known about the effects or underlying mechanism of RASSF1A on the malignant phenotype. In the present study, we observed that RASSF1A expression inhibited the malignant phenotypes of NPC cells. Stable silencing of RASSF1A in NPC cell lines induced self-renewal properties and tumorigenicity in vivo/in vitro and the acquisition of an invasive phenotype in vitro. Mechanistically, RASSF1A inactivated Yes-associated Protein 1 (YAP1), a transcriptional coactivator, through actin remodeling, which further contributed to Platelet Derived Growth Factor Subunit B (PDGFB) transcription inhibition. Treatment with ectopic PDGFB partially increased the malignancy of NPC cells with transient knockdown of YAP1. Collectively, these findings suggest that RASSF1A inhibits malignant phenotypes by repressing PDGFB expression in a YAP1-dependent manner. PDGFB may serve as a potential interest of therapeutic regulators in patients with metastatic NPC

Integrating the DNA damage and protein stress responses during cancer development and treatment

During evolution, cells have developed a wide spectrum of stress response modules to ensure homeostasis. The genome and proteome damage response pathways constitute the pillars of this interwoven ‘defensive’ network. Consequently, the deregulation of these pathways correlates with ageing and various pathophysiological states, including cancer. In the present review, we highlight: (1) the structure of the genome and proteome damage response pathways; (2) their functional crosstalk; and (3) the conditions under which they predispose to cancer. Within this context, we emphasize the role of oncogene-induced DNA damage as a driving force that shapes the cellular landscape for the emergence of the various hallmarks of cancer. We also discuss potential means to exploit key cancer-related alterations of the genome and proteome damage response pathways in order to develop novel efficient therapeutic modalities. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

MST2 kinase suppresses rDNA transcription in response to DNA damage by phosphorylating nucleolar histone H2B

The heavily transcribed rDNA repeats that give rise to the ribosomal RNA are clustered in a unique chromatin structure, the nucleolus. Due to its highly repetitive nature and transcriptional activity, the nucleolus is considered a hotspot of genomic instability. Breaks in rDNA induce a transient transcriptional shut down to conserve energy and promote rDNA repair; however, how nucleolar chromatin is modified and impacts on rDNA repair is unknown. Here, we uncover that phosphorylation of serine 14 on histone H2B marks transcriptionally inactive nucleolar chromatin in response to DNA damage. We identified that the MST2 kinase localises at the nucleoli and targets phosphorylation of H2BS14p in an ATM-dependent manner. We show that establishment of H2BS14p is necessary for damage-induced rDNA transcriptional shut down and maintenance of genomic integrity. Ablation of MST2 kinase, or upstream activators, results in defective establishment of nucleolar H2BS14p, perturbed DNA damage repair, sensitisation to rDNA damage and increased cell lethality. We highlight the impact of chromatin regulation in the rDNA damage response and targeting of the nucleolus as an emerging cancer therapeutic approach

RASSF1A is required for the maintenance of nuclear actin levels

Nuclear actin participates in many essential cellular processes including gene transcription, chromatin remodelling and mRNA processing. Actin shuttles into and out the nucleus through the action of dedicated transport receptors importin‐9 and exportin‐6, but how this transport is regulated remains unclear. Here, we show that RASSF1A is a novel regulator of actin nucleocytoplasmic trafficking and is required for the active maintenance of nuclear actin levels through supporting binding of exportin‐6 (XPO6) to RAN GTPase. RASSF1A (Ras association domain family 1 isoform A) is a tumour suppressor gene frequently silenced by promoter hypermethylation in all major solid cancers. Specifically, we demonstrate that endogenous RASSF1A localises to the nuclear envelope (NE) and is required for nucleocytoplasmic actin transport and the concomitant regulation of myocardin‐related transcription factor A (MRTF‐A), a co‐activator of the transcription factor serum response factor (SRF). The RASSF1A/RAN/XPO6/nuclear actin pathway is aberrant in cancer cells where RASSF1A expression is lost and correlates with reduced MRTF‐A/SRF activity leading to cell adhesion defects. Taken together, we have identified a previously unknown mechanism by which the nuclear actin pool is regulated and uncovered a previously unknown link of RASSF1A and MRTF‐A/SRF in tumour suppression

TGF-β targets the hippo pathway scaffold RASSF1A to facilitate YAP/SMAD2 nuclear translocation

Epigenetic inactivation of the Hippo pathway scaffold RASSF1A is associated with poor prognosis in a wide range of sporadic human cancers. Loss of expression reduces tumor suppressor activity and promotes genomic instability, but how this pleiotropic biomarker is regulated at the protein level is unknown. Here we show that TGF-β is the physiological signal that stimulates RASSF1A degradation by the ubiquitin-proteasome pathway. In response to TGF-β, RASSF1A is recruited to TGF-β receptor I and targeted for degradation by the co-recruited E3 ubiquitin ligase ITCH. RASSF1A degradation is necessary to permit Hippo pathway effector YAP1 association with SMADs and subsequent nuclear translocation of receptor-activated SMAD2. We find that RASSF1A expression regulates TGF-β-induced YAP1/SMAD2 interaction and leads to SMAD2 cytoplasmic retention and inefficient transcription of TGF-β targets genes. Moreover, RASSF1A limits TGF-β induced invasion, offering a new framework on how RASSF1A affects YAP1 transcriptional output and elicits its tumor-suppressive function

RASSF1A is required for the maintenance of nuclear actin levels

Nuclear actin participates in many essential cellular processes including gene transcription, chromatin remodelling and mRNA processing. Actin shuttles into and out the nucleus through the action of dedicated transport receptors importin‐9 and exportin‐6, but how this transport is regulated remains unclear. Here, we show that RASSF1A is a novel regulator of actin nucleocytoplasmic trafficking and is required for the active maintenance of nuclear actin levels through supporting binding of exportin‐6 (XPO6) to RAN GTPase. RASSF1A (Ras association domain family 1 isoform A) is a tumour suppressor gene frequently silenced by promoter hypermethylation in all major solid cancers. Specifically, we demonstrate that endogenous RASSF1A localises to the nuclear envelope (NE) and is required for nucleocytoplasmic actin transport and the concomitant regulation of myocardin‐related transcription factor A (MRTF‐A), a co‐activator of the transcription factor serum response factor (SRF). The RASSF1A/RAN/XPO6/nuclear actin pathway is aberrant in cancer cells where RASSF1A expression is lost and correlates with reduced MRTF‐A/SRF activity leading to cell adhesion defects. Taken together, we have identified a previously unknown mechanism by which the nuclear actin pool is regulated and uncovered a previously unknown link of RASSF1A and MRTF‐A/SRF in tumour suppression

RASSF1A is required for the maintenance of nuclear actin levels

Nuclear actin participates in many essential cellular processes including gene transcription, chromatin remodelling and mRNA processing. Actin shuttles into and out the nucleus through the action of dedicated transport receptors importin-9 and exportin-6, but how this transport is regulated remains unclear. Here, we show that RASSF1A is a novel regulator of actin nucleocytoplasmic trafficking and is required for the active maintenance of nuclear actin levels through supporting binding of exportin-6 (XPO6) to RAN GTPase. RASSF1A (Ras association domain family 1 isoform A) is a tumour suppressor gene frequently silenced by promoter hypermethylation in all major solid cancers. Specifically, we demonstrate that endogenous RASSF1A localises to the nuclear envelope (NE) and is required for nucleocytoplasmic actin transport and the concomitant regulation of myocardin-related transcription factor A (MRTF-A), a co-activator of the transcription factor serum response factor (SRF). The RASSF1A/RAN/XPO6/nuclear actin pathway is aberrant in cancer cells where RASSF1A expression is lost and correlates with reduced MRTF-A/SRF activity leading to cell adhesion defects. Taken together, we have identified a previously unknown mechanism by which the nuclear actin pool is regulated and uncovered a previously unknown link of RASSF1A and MRTF-A/SRF in tumour suppression. © 2019 The Authors. Published under the terms of the CC BY 4.0 licens

RASSF1A uncouples Wnt from Hippo signalling and promotes YAP mediated differentiation via p73.

Transition from pluripotency to differentiation is a pivotal yet poorly understood developmental step. Here, we show that the tumour suppressor RASSF1A is a key player driving the early specification of cell fate. RASSF1A acts as a natural barrier to stem cell self-renewal and iPS cell generation, by switching YAP from an integral component in the β-catenin-TCF pluripotency network to a key factor that promotes differentiation. We demonstrate that epigenetic regulation of the Rassf1A promoter maintains stemness by allowing a quaternary association of YAP-TEAD and β-catenin-TCF3 complexes on the Oct4 distal enhancer. However, during differentiation, promoter demethylation allows GATA1-mediated RASSF1A expression which prevents YAP from contributing to the TEAD/β-catenin-TCF3 complex. Simultaneously, we find that RASSF1A promotes a YAP-p73 transcriptional programme that enables differentiation. Together, our findings demonstrate that RASSF1A mediates transcription factor selection of YAP in stem cells, thereby acting as a functional "switch" between pluripotency and initiation of differentiation