β-Adrenergic Activation of p38 MAP Kinase in Adipocytes: cAMP INDUCTION OF THE UNCOUPLING PROTEIN 1 (UCP1) GENE REQUIRES p38 MAP KINASE

Wenhong Cao,Alexander V Medvedev,Kiefer W Daniel,Sheila Collins

doi:10.1074/jbc.m101049200

Abstract

Because of increasing evidence that G protein-coupled receptors activate multiple signaling pathways, it becomes important to determine the coordination of these pathways and their physiological significance. Here we show that the beta(3)-adrenergic receptor (beta(3)AR) stimulates p38 mitogen-activated protein kinase (p38 MAPK) via PKA in adipocytes and that cAMP-dependent transcription of the mitochondrial uncoupling protein 1 (UCP1) promoter by beta(3)AR requires p38 MAPK. The selective beta(3)AR agonist CL316,243 (CL) stimulates phosphorylation of MAP kinase kinase 3/6 and p38 MAPK in a time- and dose-dependent manner in both white and brown adipocytes. Isoproterenol and forskolin mimicked the effect of CL on p38 MAPK. In all cases activation was blocked by the specific p38 MAPK inhibitor SB202190 (SB; 1-10 microm). The involvement of PKA in beta(3)AR-dependent p38 MAPK activation was confirmed by the ability of the PKA inhibitors H89 (20 microm) and (R(p))-cAMP-S (1 mm) to block phosphorylation of p38 MAPK. Treatment of primary brown adipocytes with CL or forskolin induced the expression of UCP1 mRNA levels (6.8- +/- 0.8-fold), and this response was eliminated by PKA inhibitors and SB202190. A similar stimulation of a 3.7-kilobase UCP1 promoter by CL and forskolin was also completely inhibited by PKA inhibitors and SB202190, indicating that these effects on UCP1 expression are transcriptional. Moreover, the PKA-dependent transactivation of the UCP1 promoter, as well as its sensitivity to SB202190, was fully reproduced by a 220-nucleotide enhancer element from the UCP1 gene. We similarly observed that increased phosphorylation of ATF-2 by CL was sensitive to both H89 and SB202190, while phosphorylation of cAMP-response element-binding protein was inhibited only by H89. Together, these studies illustrate that p38 MAPK is an important downstream target of the beta-adrenergic/cAMP/PKA signaling pathway in adipocytes, and one of the functional consequences of this cascade is stimulation of UCP1 gene expression in brown adipocytes.

Highlights

Regulation of adipose tissue lipolysis and thermogenesis
Because of the powerful thermogenic response elicited by ␤3AR agonists in vivo and the cAMP/PKA-dependent stimulation of transcription factors suspected of being involved in the regulation of the uncoupling protein 1 (UCP1) gene, we tested the possibility that the activation of p38 MAPK by ␤3AR participated in the induction of the UCP1 gene in primary brown adipocytes
Through the use of selective kinase inhibitors and cAMP mimetics, our results clearly show that cAMP and PKA activity are required for the subsequent activation of p38 MAPK by ␤3AR in adipocytes

Summary

Introduction

Regulation of adipose tissue lipolysis and thermogenesis. Brown and white adipocytes contain all three ␤AR subtypes, each of which is coupled to G␣s to stimulate the cAMP/PKA pathway [1]. Brown and white adipocytes contain all three ␤AR subtypes, each of which is coupled to G␣s to stimulate the cAMP/PKA pathway [1]. We show that in both brown and white adipocytes the ␤3AR activates the p38 MAP kinase pathway as a downstream consequence of the generation of cAMP and PKA activity.

Results

Conclusion