CCK-A receptor activates RhoA through Gα12/13 in NIH3T3 cells

Sophie L. Le Page, Yan Bi, John A. Williams

Research output: Contribution to journalArticle

30 Scopus citations

Abstract

Cholecystokinin (CCK) is a major regulator of pancreatic acinar cells and was shown previously to be capable of inducing cytoskeletal changes in these cells. In the present study, using NIH3T3 cells stably transfected with CCK-A receptors as a model cell, we demonstrate that CCK can induce actin stress fibers through a G13- and RhoA-dependent mechanism. CCK induced stress fibers within minutes similar to those induced by lysophosphatidic acid (LPA), the active component of serum. The effects of CCK were mimicked by active RhoV14 and blocked by dominant-negative RhoN19, Clostridium botulinum C3 transferase, and the Rho-kinase inhibitor Y-27632. CCK rapidly induced active Rho in cells as shown with a pull-down assay using the Rho binding domain of rhotekin and by a serum response element (SRE)-luciferase reporter assay. To evaluate the G protein mediating the action of CCK, cells were transfected with active α-subunits; Gα13 and Gα12 but not Gαq induced stress fibers and in some cases cell rounding. A p115 Rho guanine nucleotide exchange factor (GEF) regulator of G protein signaling (RGS) domain known to interact with G12/13 inhibited active α12/13- and CCK-induced stress fibers, whereas RGS2 and RGS4, which are known to inhibit Gq, had no effect. Cotransfection with plasmids coding for the G protein α-subunit carboxy-terminal peptide from α13 and, to a lesser extent α12, also inhibited the effect of CCK, whereas the peptide from αq did not. These results show that in NIH3T3 cells bearing CCK-A receptors, CCK activates Rho primarily through G13, leading to rearrangement of the actin cytoskeleton.

Original languageEnglish (US)
Pages (from-to)C1197-C1206
JournalAmerican Journal of Physiology - Cell Physiology
Volume285
Issue number5 54-5
DOIs
StatePublished - Nov 2003

Keywords

  • Actin
  • Cholecystokinin
  • Rho
  • Rho-kinase
  • Stress fibers

ASJC Scopus subject areas

  • Physiology
  • Cell Biology

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