PGC-1Î± gene regulation in adipocyte cell lines
The cold-inducible PPAR co-activator PGC-1α has been reported to play a pivotal role in the control of pre-adipocyte differentiation to brown adipose tissue (BAT) and mitochondriogenesis. The overall aim of this study was to investigate the transcriptional control of PGC-1α gene in brown and white adipose tissue (WAT) precursor cells. Fragments up to 1.8kb of the rat PGC-1α proximal promoter were cloned into a pGL3 luciferase reporter vector and transfected into 3T3-L1 (WAT pre-adipocyte cell line) and HIB-1B (BAT pre-adipocyte cell line). The phenotype of these cell lines was confirmed by histological and biochemical characterisation. When cells were induced to differentiate, even the shortest (264bp) reporter construct supported luciferase expression in HIB-1B, but not in 3T3-L1 cells. cAMP stimulation induced endogenous PGC-1α expression only in HIB-1B cells. Putative transcription factor binding sites were identified in the 264bp proximal promoter using both EMSA and promoter analysis software. Mutation of the cAMP response element at -183 to -175 abolished completely the cAMP response of the promoter in 3T3-L1 cells, but only attenuated this response in HIB-1B cells. this suggested that other indirect pathways were acting for the cAMP stimulation of this promoter in HIB-1B cells. Additional pathways through which differential control of the proximal PGC-1α promoter region could occur were next considered. A putative C/EBP site at -101 to -94 was also implicated in the transcriptional control of this promoter. Over-expression of C/EBPβ increased the activity of the promoter and rescued the response of PGC-1α in the 3T3-L1 cells. Similar to PGC-1α, C/EBPβ was cAMP induced only in HIB-1B cells.