CCAAT/Enhancer-binding Protein β (C/EBPβ) and C/EBPδ Contribute to Growth Hormone-regulated Transcription of c-fos

Growth Hormone Regulates Phosphorylation and Function of CCAAT/Enhancer-binding Protein β by Modulating Akt and Glycogen Synthase Kinase-3

“…Immunoblotting-Cell lysis and immunoblotting for C/EBP␤ were performed as previously described (14) and for Akt and GSK-3 as follows: confluent 3T3-F442A cells on 100-mm plates were washed with phosphate-buffered saline with vanadate (10 mM Tris (pH 7.4), 150 mM sodium phosphate, 1 mM sodium vanadate) and scraped in 0.3 ml of L-RIPA lysis buffer (50 mM Hepes, pH 7.0, 250 mM NaCl, 0.5% Triton X-100) containing 1 mM Na 3 VO 4 , 1 mM phenylmethylsulfonyl fluoride, and 10 g/ml each of aprotinin and leupeptin. Whole cell lysates (35-50 g) were analyzed by immunoblotting using antibodies against phospho-Akt or phospho-GSK-3.…”

“…GH increases the binding of C/⌭BP␤ and C/EBP␦ to the c-fos C/EBP site, which lies Ϫ295 to Ϫ303 bp relative to the transcription start site, and overlaps the c-fos serum response element (SRE) (14). GH promotes a rapid and transient dephosphorylation of LAP and LIP in 3T3-F442A cells, in addition to increasing binding of C/EBP␤ to the C/EBP site in c-fos (14).…”

“…GH promotes a rapid and transient dephosphorylation of LAP and LIP in 3T3-F442A cells, in addition to increasing binding of C/EBP␤ to the C/EBP site in c-fos (14). It is well established that modulation of target gene expression can be achieved through regulation of the phosphorylation state of transcription factors, which can positively or negatively alter their DNA binding affinity and/or their capacity to activate gene transcription (20 -25).…”

“…The N-terminal region of C/EBPs contains the transcription activation and inhibitory domains. C/EBP␤, the prominent GH-regulated form (14) is present in cells as three alternate translation products. The 32-and 35-kDa forms of C/EBP␤, also known as liver-activating proteins (LAP), are potent transactivators.…”

“…The CCAAT/Enhancer-Binding Proteins (C/EBPs) have recently been shown to participate in GH-regulated transcription of c-fos (14). C/EBPs, which include C/EBP␣, C/EBP␤, C/EBP␦, C/EBP⑀, and C/EBP/CHOP, belong to the bZIP family of transcription factors characterized by a C-terminal dimerization domain (leucine zipper) adjacent to a basic DNA binding domain.…”