Premium
Gα q signaling is required for Rho‐dependent transcriptional activation of the cyclooxygenase‐2 promoter in fibroblasts
Author(s) -
Slice Lee W.,
Han SangKyou,
Simon Melvin I.
Publication year - 2003
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.10195
Subject(s) - microbiology and biotechnology , transfection , gq alpha subunit , heterotrimeric g protein , luciferase , signal transduction , biology , cell culture , protein kinase c , thapsigargin , chemistry , g protein , endoplasmic reticulum , genetics
Previously, we demonstrated that the gastrin releasing peptide (GRP) induces cyclooxygenase‐2 (COX‐2) expression through a Rho‐dependent, protein kinase C (PKC)‐independent signaling pathway in fibroblasts (Slice et al., 1999, J Biol Chem 274:27562–27566). However, the specific role of heterotrimeric guanine nucleotide binding regulatory proteins (G‐proteins) that are coupled to the GRP receptor in Rho‐dependent COX‐2 expression has not been elucidated. In this report, we utilize embryonic fibroblasts from transgenic mice containing double gene knock‐outs (DKO) for Gα q/11 and Gα 12/13 to demonstrate that COX‐2 promoter activation by GRP requires Gα q . Furthermore, we show that GRP‐dependent COX‐2 gene expression, as assessed by a COX‐2 reporter luciferase assay, was induced in cells lacking Gα 12/13 but was blocked in cells that did not express Gα q/11 . GRP‐dependent COX‐2 promoter induction in Gα q/11 deficient cells was rescued by expression of wild type Gα q but blocked by inhibition of calcium signaling in calcium‐free media or in cells treated with 2‐aminoethoxydiphenylborate (2‐APB). Co‐stimulation of transfected Gα q/11 deficient cells with GRP and thapsigargin (TG) induced the COX‐2 promoter. Activation of endogenous Rho by expression of Onco‐lbc or expression of Rho A Q63L resulted in COX‐2 promoter activation in Gα q/11 deficient cells. Inhibition of Rho by Clostridium botulinum C3 toxin blocked COX‐2 promoter induction. Expression of Gα q Q209L in the well‐characterized fibroblast cell line, NIH3T3, induced the COX‐2 promoter which was blocked by expression of C3 toxin. These results demonstrate that calcium signaling mediated by Gα q and Rho play critical roles in GRP‐dependent COX‐2 expression in fibroblasts. © 2002 Wiley‐Liss, Inc.