rac p21 is involved in insulin-induced membrane ruffling and rho p21 is involved in hepatocyte growth factor- and 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced membrane ruffling in KB cells.

Abstract

Insulin and hepatocyte growth factor (HGF) induced morphologically different membrane ruiflings in KB cells. Insulin-induced membrane ruffling was inhibited by microinjection of rho GDI, an inhibitory GDP/GTP exchange regulator for both rho p21 and rac p21 small GTP-binding proteins, but not inhibited by microinjection of botulinum exoenzyme C3, known to selectively ADP-ribosylate rho p21 and to impair its function. This rho GDI action was prevented by comicroinjection with guanosine 5'-(3-O-thio)triphosphate (GTPyS)-bound racl p21. In contrast, HGF-induced membrane ruffling was inhibited by microinjection of rho GDI or C3. This rho GDI action was prevented by comicroinjection with GTP-yS-bound rhoA p21, and this C3 action was prevented by comicroinjection with GTPyS-bound rhoAIe-41 p21, which is resistant to C3.Microinjection of either GTP-yS-bound racl p21 or rhoA p21 alone induced membrane ruffling in the absence of the growth factors. The racl p21-induced membrane ruffling was morphologically similar to the insulininduced kind, whereas rhoA p21-induced ruffling was apparently different from both the insulin-and HGF-induced kinds. Membrane ruffling was also induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), a protein kinase C-activating phorbol ester, but not by Ca2+ ionophore or microinjection of a dominant active Ki-ras p21 mutant (Ki-rasVaII2 p21). The phorbol ester-induced membrane ruffling was morphologically similar to the rhoA p21-induced kind and inhibited by microinjection of rho GDI or C3. These results indicate that rac p21 and rho GDI are involved in insulin-induced membrane ruffling and that rho p21 and rho GDI are involved in HGF-and phorbol ester-induced membrane rufflings.Membrane ruffling is rapid movement with irregular fluctuation of protrusion and withdrawal of the margin of the cell surface and is believed to be associated with spreading and locomotion in cultured cells (1, 17). The molecular events responsible for the ruffling formation are not known, but it has been suggested to be caused by a polymerization of actin at the inner surface of the plasma membrane (32). It is known that various growth factors induce morphologically different membrane rufflings in various types of cells (20,26,27,32,34,43 factor (PDGF) (32). These growth factors are known to induce a number of rapid biological responses, including an increase in receptor-associated tyrosine kinase activity, phosphoinositide turnover associated with production of diacylglycerol and inositol triphosphate, and intracellular Ca2+ mobilization (for a review, see reference 12). However, it is not known at present which, if any, of these receptor-mediated signals is involved in growth factor-induced membrane rufflings.It has recently been reported that racl p21 regulates membrane ruffling induced by PDGF, EGF, and insulin in Swiss 3T3 cells (43). racl p21 is a member of the rho family which belongs to the small GTP-binding protein (G protein) superfamily (15; for reviews, see references 23 and 52). The rho family consists of th...

RhoA/ROCK-mediated switching between Cdc42- and Rac1-dependent protrusion in MTLn3 carcinoma cells

“…Thus, RhoA activation is coincident with barbed end formation in MTLn3 cells in response to EGF stimulation [2]. The finding of activated RhoA in EGF-stimulated protrusions is consistent with recent reports implicating RhoA activity at the leading edge of moving cells [17,19,47,48].…”

Cell motility mediated by rho and rho-associated protein kinase plays a critical role in intrahepatic metastasis of human hepatocellular carcinoma

“…39 Rho activation is also involved in hepatocyte growth factor-and phorbol ester-induced membrane ruffling in epidermoid carcinoma cells. 40 Moreover, several reports have indicated that Rho is involved in the motility of several types of cells including neutrophils, 41 endothelial cells, 42 fibroblasts, 43 keratinocytes, 44 and some colon cancer cells (Sakamoto M, Hirohashi S, unpublished observation, 1996). However, it is still unclear how each member of the small GTPases is involved in the motility of different kinds of cells, or how cell adhesion and cytoskeletal systems are coordinated, especially in epithelial cells.…”

Differential Regulation of Focal Adhesion Kinase and Mitogen‐Activated Protein Kinase Tyrosine Phosphorylation During Insulin‐Like Growth Factor‐I‐Mediated Cytoskeletal Reorganization

“…In fibroblasts, membrane ruffling is stimulated by several growth factors, including IGF-I (Kotani et al, 1994;Wennström et al, 1994). In nonneuronal cells, activation of P1 3-K is necessary for membrane ruffling by acting as an upstream regulator of the small GTPase protein, Rac (Kotani et al, 1994;Nishiyama et al, 1994;Wennstrdm et al, 1994;Hawkins et al, 1995;Parker, 1995). We are currently investigating the role of P1 3-K activation of Rac in IGF-I-mediated changes in neuronal architecture (Kim et al, 1996).…”