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The peroxisome proliferator-activated receptor-γ regulates murine pyruvate carboxylase gene expression in vivo and in vitro

The peroxisome proliferator-activated receptor-γ regulates murine pyruvate carboxylase gene expression in vivo and in vitro
The peroxisome proliferator-activated receptor-γ 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-γ (PPARγ) 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 α, PPARγ1, and PPARγ2. Tumor necrosis factor-α, 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 α/PPARγ1 or retinoid X receptor α/PPARγ2 showed a 6-8-fold increase above the basal promoter activity. Furthermore, treatment of these transfected cells with the PPARγ agonist doubled the promoter activity. Mutation of the putative PPAR-response element-(-386/-374) of this 2.3-kb PC promoter fragment abolished the PPARγ response. Gel shift and chromatin immunoprecipitation assays demonstrated that endogenous PPARy binds to this functional PPAR-response element of the PC promoter. Mice with targeted disruption of the PPARγ2 gene displayed ∼50-60% reduction of PC mRNA and protein in white adipose tissue. Similarly, in brown adipose tissue of PPARγ2-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 PPARγ2 knock-out mice was also associated with a marked reduction of PC mRNA. Our findings identified PC as a PPARγ-regulated gene and suggested a role for PPARγ regulating intermediary metabolism.

0021-9258
27466-27476
Jitrapakdee, Sarawut
65ab65c0-ec85-4e57-86ee-e0619ecce1b5
Slawik, Marc
2fa28a50-9447-4875-bdd7-4b2be433f0c7
Medina-Gomez, Gema
9e8167bb-cfac-4bec-8e25-da1b42120e26
Campbell, Mark
ed49d284-9484-44c4-9bb0-5a661309e60f
Wallace, John C.
d34fbc59-5a77-47fb-b916-2a2b67fda5ab
Sethi, Jaswinder K.
923f1a81-91e4-46cd-8853-bb4a979f5a85
O'Rahilly, Stephen
e5c7869f-10d1-4fdc-a564-e4907be415a3
Vidal-Puig, Antonio J.
bd6ec045-f9b8-48ff-ae86-a86a6a513435
Jitrapakdee, Sarawut
65ab65c0-ec85-4e57-86ee-e0619ecce1b5
Slawik, Marc
2fa28a50-9447-4875-bdd7-4b2be433f0c7
Medina-Gomez, Gema
9e8167bb-cfac-4bec-8e25-da1b42120e26
Campbell, Mark
ed49d284-9484-44c4-9bb0-5a661309e60f
Wallace, John C.
d34fbc59-5a77-47fb-b916-2a2b67fda5ab
Sethi, Jaswinder K.
923f1a81-91e4-46cd-8853-bb4a979f5a85
O'Rahilly, Stephen
e5c7869f-10d1-4fdc-a564-e4907be415a3
Vidal-Puig, Antonio J.
bd6ec045-f9b8-48ff-ae86-a86a6a513435

Jitrapakdee, Sarawut, Slawik, Marc, Medina-Gomez, Gema, Campbell, Mark, Wallace, John C., Sethi, Jaswinder K., O'Rahilly, Stephen and Vidal-Puig, Antonio J. (2005) The peroxisome proliferator-activated receptor-γ regulates murine pyruvate carboxylase gene expression in vivo and in vitro. The Journal of Biological Chemistry, 280 (29), 27466-27476. (doi:10.1074/jbc.M503836200).

Record type: Article

Abstract

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-γ (PPARγ) 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 α, PPARγ1, and PPARγ2. Tumor necrosis factor-α, 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 α/PPARγ1 or retinoid X receptor α/PPARγ2 showed a 6-8-fold increase above the basal promoter activity. Furthermore, treatment of these transfected cells with the PPARγ agonist doubled the promoter activity. Mutation of the putative PPAR-response element-(-386/-374) of this 2.3-kb PC promoter fragment abolished the PPARγ response. Gel shift and chromatin immunoprecipitation assays demonstrated that endogenous PPARy binds to this functional PPAR-response element of the PC promoter. Mice with targeted disruption of the PPARγ2 gene displayed ∼50-60% reduction of PC mRNA and protein in white adipose tissue. Similarly, in brown adipose tissue of PPARγ2-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 PPARγ2 knock-out mice was also associated with a marked reduction of PC mRNA. Our findings identified PC as a PPARγ-regulated gene and suggested a role for PPARγ regulating intermediary metabolism.

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e-pub ahead of print date: 25 May 2005
Published date: 22 July 2005

Identifiers

Local EPrints ID: 415402
URI: http://eprints.soton.ac.uk/id/eprint/415402
ISSN: 0021-9258
PURE UUID: 083db56d-e424-4bbd-b10d-519dec5e870d
ORCID for Jaswinder K. Sethi: ORCID iD orcid.org/0000-0003-4157-0475

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Date deposited: 09 Nov 2017 17:30
Last modified: 16 Mar 2024 04:31

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Contributors

Author: Sarawut Jitrapakdee
Author: Marc Slawik
Author: Gema Medina-Gomez
Author: Mark Campbell
Author: John C. Wallace
Author: Stephen O'Rahilly
Author: Antonio J. Vidal-Puig

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