The peroxisome proliferator-activated receptor-gamma regulates murine pyruvate carboxylase gene expression in vivo and in vitro.
Pyruvate carboxylase (PC) plays a crucial role in various metabolic pathways, including gluconeogenesis, lipogenesis, and glucose-induced insulin secretion. Here we showed for the first time that the PC gene is transcriptionally regulated by peroxisome proliferator-activated receptor-gamma (PPARgamma) in vitro and in vivo in white and brown adipose tissue. PC mRNA and protein are markedly increased during differentiation of 3T3-L1 cells and HIB-1B, in parallel with the expression of the adipogenic transcription factors, CCAAT-enhancer binding protein alpha, PPARgamma1, and PPARgamma2. Tumor necrosis factor-alpha, a cytokine that blocks differentiation of 3T3-L1 cells, suppressed PC expression. Co-transfection studies in 3T3-L1 preadipocytes or HEK293T cells with a 2.3-kb promoter fragment of mouse PC gene linked to a luciferase reporter construct and with plasmids overexpressing retinoid X receptor alpha/PPARgamma1 or retinoid X receptor alpha/PPARgamma2 showed a 6-8-fold increase above the basal promoter activity. Furthermore, treatment of these transfected cells with the PPARgamma agonist doubled the promoter activity. Mutation of the putative PPAR-response element-(-386/-374) of this 2.3-kb PC promoter fragment abolished the PPARgamma response. Gel shift and chromatin immunoprecipitation assays demonstrated that endogenous PPARgamma binds to this functional PPAR-response element of the PC promoter. Mice with targeted disruption of the PPARgamma2 gene displayed approximately 50-60% reduction of PC mRNA and protein in white adipose tissue. Similarly, in brown adipose tissue of PPARgamma2-deficient mice subjected to cold exposure, PC mRNA was 40% lower than that of wild type mice. Impaired in vitro differentiation of white adipocytes of PPARgamma2 knock-out mice was also associated with a marked reduction of PC mRNA. Our findings identified PC as a PPARgamma-regulated gene and suggested a role for PPARgamma regulating intermediary metabolism.
Jitrapakdee
Sarawut
S
Cambridge Institute of Diabetes, Endocrinology, and Metabolism and the Department of Clinical Biochemistry, University of Cambridge, Addenbrooke's Hospital, Cambridge, CB2 2QR, United Kingdom. scsji@mahidol.ac.th
Slawik
Marc
M
Medina-Gomez
Gema
G
Campbell
Mark
M
Wallace
John C
JC
Sethi
Jaswinder K
JK
O'rahilly
Stephen
S
Vidal-Puig
Antonio J
AJ
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D047495:PPAR gamma / Q000502:physiology*; D011401:Promoter Regions, Genetic; D011766:Pyruvate Carboxylase / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D047488:Retinoid X Receptors / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection
D047495:PPAR gamma; D012333:RNA, Messenger; D047488:Retinoid X Receptors; D011766:Pyruvate Carboxylase