Cooperative regulation of endogenous cAMP-response element binding protein and CCAAT/enhancer-binding protein beta in GH-stimulated c-fos expression.
GH activates the c-fos promoter by regulating multiple transcription factors. This study adds to our understanding of GH-regulated transcription by demonstrating that GH regulates the c-fos cAMP-response element (CRE) and its binding protein, CREB. Activation of the c-fos promoter by GH is impaired by expression of dominant-negative A-CREB. GH stimulates rapid and transient phosphorylation of CREB at Ser 133 (P-CREB), a critical site for transactivation by CREB, in 3T3-F442A preadipocytes. Mutation of this residue impairs GH-induced c-fos expression, suggesting that phosphorylation of CREB at Ser 133 contributes to GH-induced c-fos activation. The MEK inhibitor UO126 impaired the phosphorylation of CREB and that of C/EBPbeta, suggesting that ERKs mediate the phosphorylation of both proteins. UO126, but not the protein kinase A inhibitor H89, blocked GH-induced c-fos mRNA expression. A combination of CREB and C/EBPbeta enhanced c-fos promoter activation, and mutation of the CRE impaired the enhancement, as well as GH-stimulated c-fos activation. GH treatment increased the occupancy of both endogenous phospho-CREB and phospho-C/EBPbeta on the c-fos promoter. The increases were impaired by UO126. The active P-CREB and P-C/EBPbeta are induced by GH to occupy the same c-fos promoter DNA, suggesting that they may participate in a GH-regulated complex on c-fos. These findings suggest that coordinated phosphorylation of CREB and C/EBPbeta in response to GH is mediated by ERK1/2, and that the phosphorylated proteins are part of a regulatory complex that occupies c-fos in vivo to regulate c-fos transcription cooperatively in response to GH.
Cui
Tracy Xiao
TX
Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, Michigan 48109-0622, USA.
Kwok
Roland
R
Schwartz
Jessica
J
D016475:3T3 Cells; D017667:Adipocytes / Q000378:metabolism; D000818:Animals; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000032:analysis / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D017362:Cyclic AMP Response Element-Binding Protein / Q000032:analysis / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D016762:Genes, fos / Q000187:drug effects / Q000235:genetics*; D013006:Growth Hormone / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012333:RNA, Messenger / Q000032:analysis; D011986:Receptors, Somatotropin / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000494:pharmacology
C508138:A-CREB protein; D022782:CCAAT-Enhancer-Binding Protein-beta; D017362:Cyclic AMP Response Element-Binding Protein; D012333:RNA, Messenger; D011986:Receptors, Somatotropin; D011993:Recombinant Fusion Proteins; D013006:Growth Hormone; D004247:DNA; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3