Gαq signaling is required for Rho-dependent transcriptional activation of the cyclooxygenase-2 promoter in fibroblasts
✍ Scribed by Lee W. Slice; Sang-Kyou Han; Melvin I. Simon
- Publisher
- John Wiley and Sons
- Year
- 2003
- Tongue
- English
- Weight
- 419 KB
- Volume
- 194
- Category
- Article
- ISSN
- 0021-9541
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✦ Synopsis
Abstract
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.