The role of RhoA in the regulation of cell morphology and motility

Members of the Rho family of small GTPases are key regulators of the actin cytoskeleton, particularly in relation to the cell shape changes and the adhesion dynamic that drive cell migration. Here, we report the effect of activation or inhibition of the function of RhoA on cell motility and morphology. Both in the presence and the absence of serum, expression of constitutively active RhoA dramatically inhibited L929 fibroblasts' cell motility, and induced a rounding of the cells and a decrease in the number of processes per cell. In contrast, expression of a dominant negative mutant of RhoA had no effect on cell motility or morphology in steady-state conditions with or without serum in the medium. Inhibition of p160ROCK, a kinase effector of RhoA, only partially inhibited cell migration. Conversely, when cells were submitted to a period of serum deprivation followed by addition of serum, inhibition of endogenous RhoA by expression of the dominant negative mutant of RhoA impeded cell motility after serum stimulation. Thus, RhoA activity is required for stimulation of cell locomotion by serum factors. It was also observed that the addition of serum factors to quiescent L929 and NR6wtEGFR fibroblasts resulted in a delayed motility response of several hours compared to the immediately induced morphological changes, indicating the absence of a previously assumed direct correlation between changes in cell motility and cell morphology in response to serum addition. The motility response of L929 and NR6wtEGFR fibroblasts to serum stimulation required protein synthesis.