Integrin alpha2beta1 modulates EGF stimulation of Rho GTPase-dependent morphological changes in adherent human rhabdomyosarcoma RD cells

The ability of cells to undergo shape changes is essential for diverse cellular functions including cell growth, differentiation, and movement. The present study examines how an integration of the function of alpha2beta1 integrin with that of the receptor for epidermal growth factor (EGFR) modulates EGF-stimulated morphological changes in human rhabdomyosarcoma RD transfectant cells. Upon EGF stimulation, RD transfectant cells that lacked alpha2beta1 integrin expression (RDpF) underwent contraction; in contrast, expression of alpha2beta1 on RD cells (RDX2C2) resulted in transient cell spreading. Integrin alpha2 cytoplasmic domain played a critical role in the observed alpha2beta1-mediated conversion from a cell rounding to a cell spreading phenotype. Thus, the expression of an alpha2 cytoplasmic domain deletion variant (X2C0) or a chimeric alpha2beta1 containing the cytoplasmic domain of alpha4 (X2C4) or alpha5 (X2C5), instead of alpha2, failed to mediate spreading upon EGF stimulation. Using dominant negative (DN) mutants of RhoGTPases, results revealed that RhoA activation was required for both EGF-stimulated responses of cell rounding and spreading, Cdc42 functioned in the re-spreading of cells after undergoing EGF-stimulated contraction, and Rac1 was required in alpha2beta1-mediated RD cell spreading. Therefore, alpha2beta1 integrin function can switch the Rho GTPase-dependent cell shape changes in RD cells from an EGF-stimulated cell contraction to a spreading morphology. Together, results show that integrin alpha2 cytoplasmic domain plays an indispensable role in the ability of integrin alpha2beta1 to modulate EGF stimulation of Rho-GTPase-dependent morphological changes in RD cells.