The angiotensin II type 1 receptor induces membrane blebbing by coupling to Rho A, Rho kinase, and myosin light chain kinase.
The angiotensin II type 1 receptor (AT(1)R) is a G alpha(q/11)-coupled G protein-coupled receptor that is widely expressed in multiple tissues, including vascular smooth muscle cells, brain, and kidney. Activation of the AT(1)R in vascular smooth muscle cells leads to alterations in actin-based membrane protrusions such as lamellipodia, filopodia, and membrane blebs that ultimately lead to cell migration, which is important for the regulation of vascular tone. In the present study, we examine the role of small G proteins in mediating AT(1)R-induced alterations in membrane dynamics in human embryonic kidney 293 cells. We find that the activation of the AT(1)R with 100 nM angiotensin II results in the rapid formation of membrane blebs at early time points of agonist stimulation that cease within 40 min of agonist stimulation. AT(1)R-stimulated membrane bleb formation is independent of RalA, RalB, Rac1, cdc42, Arf6, and Ras, but it involves RhoA. Furthermore, membrane blebbing activated by the AT(1)R is attenuated in the presence of the beta-arrestin amino-terminal domain, Ral GDP dissociation stimulator (RalGDS) beta-arrestin binding domain, and short interfering RNA (siRNA) depletion of beta-arrestin2. However, siRNA depletion of RalGDS protein did not affect membrane blebbing in response to AT(1)R activation. The inhibition of the downstream RhoA effectors Rho kinase (ROCK) and myosin light chain kinase (MLCK) effectively attenuated AT(1)R-mediated membrane blebbing. Thus, we show that membrane blebbing in response to AT(1)R signaling is dependent on beta-arrestin2 and is mediated by a RhoA/ROCK/MLCK-dependent pathway.
Godin
Christina M
CM
J. Allyn Taylor Centre for Cell Biology, Robarts Research Institute, University of Western Ontario, 100 Perth Drive, London, ON, Canada.
Ferguson
Stephen S G
SS
D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D006801:Humans; D018613:Microscopy, Confocal; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D034741:RNA, Small Interfering; D044140:Receptor, Angiotensin, Type 1 / Q000502:physiology*; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D047428:Protein Kinase Inhibitors; D034741:RNA, Small Interfering; D044140:Receptor, Angiotensin, Type 1; D054460:rho-Associated Kinases; D009219:Myosin-Light-Chain Kinase