Faculty of 1000 evaluation for RAP2 mediates mechanoresponses of the Hippo pathway [dataset]

Guido Serini, Donatella Valdembri
2018 F1000 - Post-publication peer review of the biomedical literature   unpublished
Mammalian cells are surrounded by neighboring cells and extracellular matrix (ECM), which provide cells with structural support and mechanical cues that influence diverse biological processes 1 . The Hippo pathway effectors YAP and TAZ are regulated by mechanical cues and mediate cellular responses to ECM stiffness 2,3 . Here we identified Ras-related GTPase RAP2 as a critical intracellular signal transducer that relays ECM rigidity signals to control mechanosensitive cellular activities
more » ... YAP/TAZ. RAP2 is activated by low ECM stiffness, and RAP2 deletion blocks YAP/TAZ regulation by stiffness signals and promotes aberrant cell growth. Mechanistically, matrix stiffness acts through phospholipase Cγ1 (PLCγ1) to influence levels of phosphatidylinositol 4,5-bisphosphate (PIP2) and its product phosphatidic acid (PA), which activates RAP2 through PDZGEF1/2. At low stiffness, active RAP2 binds to and stimulates mitogen-activated protein kinase kinase kinase kinase 4/6/7 (MAP4K4/6/7) and Rho GTPase activating protein 29 (ARHGAP29), resulting in LATS1/2 activation and YAP/TAZ inhibition. RAP2 and YAP/TAZ play pivotal roles in mechano-regulated transcription, as YAP/TAZ deletion abolishes the ECM stiffness-responsive transcriptome. Our findings reveal RAP2 as a molecular switch in mechanotransduction, thereby defining a mechanosignaling pathway from ECM stiffness to the nucleus. YAP/TAZ function as essential effectors of mechanotransduction to regulate cell proliferation and differentiation [3] [4] [5] [6] [7] . When cells are shifted from stiff to soft matrices, YAP/TAZ translocate from the nucleus to the cytoplasm, and are thus inactivated. However, the signaling mechanism from ECM stiffness to the Hippo pathway is unclear. Because small GTPases function as molecular switches in many biological processes 8 , we screened for small GTPases that affect YAP/TAZ localization in cells seeded on soft (1 kPa) or stiff (40 kPa) matrices (Supplemental information). RAP2A was identified since its overexpression induced cytoplasmic translocation of YAP/TAZ even on a stiff matrix (Fig. 1a ). No other GTPases, including the closely related RAP1 and RAS, showed similar activity (Extended Data Fig. 1a ). At high stiffness, both wild-type (WT) and RAP2A/B/C-triple knockout (RAP2-KO) MCF10A cells showed nuclear localization of YAP/TAZ (Fig. 1b,c ). At low stiffness, WT cells exhibited mainly cytoplasmic YAP/TAZ, whereas RAP2-KO MCF10A cells retained YAP/TAZ in the nucleus (Fig. 1c ). RAP2 deletion in HEK293A cells also suppressed low stiffness-induced YAP/TAZ cytoplasmic translocation (Fig. 1d,e , Extended Data Fig. 1b ). YAP/TAZ target genes CTGF, CYR61, and ANKRD1 were repressed by low stiffness in WT cells, but not in the RAP2-KO cells (Fig. 1f ). Similar results were observed in human mesenchymal stem cells (Extended Data Fig. 1c-e ), in which RAP2 deletion suppressed their differentiation into adipocytes (Extended Data Fig. 1f,g ). In the luminal breast cancer MCF7 cells, ECM stiffness modulated YAP/TAZ localization in a RAP2-dependent manner, whereas the basal type MDA-MB-468 showed constitutively cytoplasmic YAP/TAZ localization regardless of stiffness (Extended Fig. 1h-l ). TWIST and β-catenin were reported to show nuclear-cytoplasmic shuttling in response to physical cues 9,10 . TWIST, but not β-Meng et al.
doi:10.3410/f.733846171.793552360 fatcat:nubd2ic3tnejfhv55c24ekgwtq