Ishikawa, Y. | Search bioRxiv

bioRxiv2019

Membrane Protrusion Formation Mediated by Rho/ROCK Signalling and Modulation of Chloride Flux

Membrane protrusion is an important structural property associated with various cellular functions. The pentaspan membrane protein Prominin-1 (Prom1/CD133) is known to be localised to the protrusions and plays a pivotal role in migration and the determination of cellular morphology; however, the underlying mechanisms have been elusive. Here, we demonstrate that Prom1 is sufficient to trigger membrane protrusion formation. Overexpression of Prom1 in the RPE-1 cells triggers multiple long cholesterol-enriched protrusions, independently from actin and tubulin polymerisation. For this protrusion formation, the five amino acid stretch located at the carboxyl cytosolic region is essential. Moreover, the small GTPase Rho and its effector kinase ROCK are essential for this protrusion formation, and the intersection point of active Rho and Prom1 is where the protrusion formation initiates. Importantly, Prom1 causes the chloride ion efflux induced by calcium ion uptake, and protrusion formation is closely associated with the chloride efflux activity. Altogether, this study has elucidated that Prom1 plays critical roles for the membrane morphology and chloride ion flux.

cell biology↗

bioRxiv2018

Pharmacological inhibition of PRMT7 links arginine monomethylation to the cellular stress response

Protein arginine methyltransferases (PRMTs) regulate diverse biological processes and are increasingly being recognized for their potential as drug targets. Here we report the discovery of a potent, selective and cell active chemical probe for PRMT7. SGC3027 is a cell permeable prodrug, which in cells, is converted to SGC8158, a potent, SAM-competitive PRMT7 inhibitor. Inhibition or knockout of cellular PRMT7 resulted in drastically reduced levels of arginine monomethylation of HSP70 family members and other stress-associated proteins. Structural and biochemical analysis revealed that PRMT7-driven in vitro methylation of HSP70 at R469 requires an ATP-bound, open conformation of HSP70. In cells, SGC3027 inhibited methylation of both constitutive and inducible forms of HSP70, and led to decreased tolerance for perturbations of proteostasis including heat shock and proteasome inhibitors. These results demonstrate a role for PRMT7 and arginine methylation in stress response.

biochemistry↗