Induction of connective tissue growth factor by activation of heptahelicalreceptors - Modulation by Rho proteins and the actin cytoskeleton

Expression of connective tissue growth factor (CTGF) was induced in renal m esangial cells by activation of heptahelical receptors by serotonin (5-HT) and lysophosphatidic acid (LPA), Induction of CTGF mRNA was transient with maximal expression after I to 2 h, whereas induction of CTGF by transformin g growth factor beta (TGF-beta) increased over time, In contrast to the ind uction of other early response genes (Egr-1 and cyclooxygenase-2), LPA-medi ated induction of CTGF was pertussis toxin-insensitive and independent of p 42/44 MAP kinase activation. B-HT-mediated CTGF induction was due to activa tion of 5-HT2A receptors and likewise independent of p42/44 MAP kinase acti vation. Upon stimulation, enhanced levels of CTGF protein were detected in cellular homogenates, whereas no protein was detectable in cell culture sup ernatants, Inhibition of proteins of the Rho family by toxin B abrogated ba sal as well as CTGF expression stimulated by LPA, 5-HT, and TGF-beta. Inhib ition of the downstream mediator of RhoA, the Rho kinase by Y-27632 partial ly reduced induction of CTGF by LPA and TGF-beta. Toxin B not only affected gene expression, but disrupted the actin cytoskeleton similarly as observe d after treatment with cytochalasin D, Disassembly of actin stress fibers b y cytochalasin D partially reduced basal and stimulated CTGF expression. Th ese data indicate that an intact actin cytoskeleton is critical for the exp ression of CTGF, Elimination of the input of Rho proteins by toxin B, howev er, was significantly more effective and their effect on CTGF expression th us goes beyond disruption of the cytoskeleton. These findings thus establis h activation of heptahelical receptors coupled to pertussis toxin-insensiti ve G:proteins as a novel signaling pathway to induce CTGF. Proteins of the Rho family and an intact cytoskeleton were identified as critical determina nts of CTGF expression induced by LPA and 5-HT, and also by TGF-beta.