Combinatorial transcription factor regulation of the cyclic AMP-response element on the Pgc-1alpha promoter in white 3T3-L1 and brown HIB-1B preadipocytes

Cold stress in rodents increases the expression of UCP1 and PGC-1alpha in brown and white adipose tissue. We have previously reported that C/EBPbeta specifically binds to the CRE on the proximal Pgc-1alpha promoter and increases forskolin-sensitive Pgc-1alpha and Ucp1 expression in white 3T3-L1 preadipocytes. Here we show that in mice exposed to a cold environment for 24 h, Pgc-1alpha, Ucp1, and C/ebpbeta but not C/ebpalpha or C/ebpdelta expression were increased in BAT. Conversely, expression of the C/EBP dominant negative Chop10 was increased in WAT but not BAT during cold exposure. Reacclimatization of cold-exposed mice to a warm environment for 24 h completely reversed these changes in gene expression. In HIB-1B, brown preadipocytes, forskolin increased expression of Pgc-1alpha, Ucp1, and C/ebpbeta early in differentiation and inhibited Chop10 expression. Employing chromatin immunoprecipitation, we demonstrate that C/EBPbeta, CREB, ATF-2, and CHOP10 are bound to the Pgc-1alpha proximal CRE, but CHOP10 does not bind in HIB-1B cell lysates. Forskolin stimulation and C/EBPbeta overexpression in 3T3-L1 cells increased C/EBPbeta and CREB but displaced ATF-2 and CHOP10 binding to the Pgc-1alpha proximal CRE. Overexpression of ATF-2 and CHOP10 in 3T3-L1 cells decreased Pgc-1alpha transcription. Knockdown of Chop10 in 3T3-L1 cells using siRNA increased Pgc-1alpha transcription, whereas siRNA against C/ebpbeta in HIB-1B cells decreased Pgc-1alpha and Ucp1 expression. We conclude that the increased cAMP stimulation of Pgc-1alpha expression is regulated by the combinatorial effect of transcription factors acting at the CRE on the proximal Pgc-1alpha promoter.