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. 2005 Feb 15;19(4):453-61.
doi: 10.1101/gad.1263305. Epub 2005 Jan 28.

Corepressors selectively control the transcriptional activity of PPARgamma in adipocytes

Hong-Ping Guan  1 , Takahiro Ishizuka, Patricia C Chui, Michael Lehrke, Mitchell A Lazar
Affiliations

Corepressors selectively control the transcriptional activity of PPARgamma in adipocytes

Hong-Ping Guan et al. Genes Dev. .

Abstract

Peroxisome proliferator-activated receptor gamma (PPARgamma) is the master regulator of adipogenesis as well as the _target of thiazolidinedione (TZD) antidiabetic drugs. Many PPARgamma _target genes are induced during adipogenesis, but others, such as glycerol kinase (GyK), are expressed at low levels in adipocytes and dramatically up-regulated by TZDs. Here, we have explored the mechanism whereby an exogenous PPARgamma ligand is selectively required for adipocyte gene expression. The GyK gene contains a functional PPARgamma-response element to which endogenous PPARgamma is recruited in adipocytes. However, unlike the classic PPARgamma-_target gene aP2, which is constitutively associated with coactivators, the GyK gene is _targeted by nuclear receptor corepressors in adipocytes. TZDs trigger the dismissal of corepressor histone deacetylase (HDAC) complexes and the recruitment of coactivators to the GyK gene. TZDs also induce PPARgamma-Coactivator 1alpha (PGC-1alpha), whose recruitment to the GyK gene is sufficient to release the corepressors. Thus, selective modulation of adipocyte PPARgamma _target genes by TZDs involves the dissociation of corepressors by direct and indirect mechanisms.

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Figures

Figure 1.
Figure 1.
Identification of a functional PPRE in the GyK gene. (A) Luciferase reporter assays of transiently transfected 3T3-L1 adipocytes using different truncations of the GyK enhancer/promoter in the pGL2 plasmid vector treated with vehicle (DMSO) or rosiglitazone (rosi). (*) p < 0.01 vs. vehicle. (B) Alignment of mouse, rat, and human GyK, aP2, PEPCK, and perfect DR-1 PPRE region. (C) DNA mobility-shift assay using a 32P-labeled fragment from the GyK enhancer (–2041 to –1886 bp). Competition using cold oligonucleotides containing the GyK, aP2, or ideal DR-1 PPREs, or mutant forms, at 10- and 50-fold molar excess is shown at right. Mutation of the PPARγ/RXRα-binding site in the GyK gene abolishes rosiglitazone induction in adipocytes. (P/R) PPARγ/RXRα heterodimer complex; (FP) free probe. (D) Luciferase assay of the GyK reporter plasmid (–2009 in Fig. 2A) with (WT) or without (Mut1) the PPRE Mut 1 shown in Figure 2B. (*) p < 0.001 vs. vehicle; (#) p < 0.01 vs. wild type treated with vehicle. (RLU) Relative luciferase units.
Figure 2.
Figure 2.
Endogenous adipocyte PPARγ binds to the GyK PPRE in vitro and in living cells. (A) DNA mobility-shift assay using nuclear extracts from 3T3-L1 preadipocytes and adipocytes. The same oligonucleotides as in Figure 1B are used. (S) Supershift; (P/R) PPARγ/RXRα; (FP) free probe. (B) Sizes of sonicated DNA fragments in preadipocytes, adipocytes treated with DMSO, and rosiglitazone for ChIP analysis. (M) 100-bp marker ladder. (C) ChIP analysis of PPARγ association with GyK and aP2 genes. The location of the PPREs and ChIP primers are indicated. (1) Preadipocyte. (2) Adipocyte treated with DMSO for 48 h. (3) Adipocyte treated with rosiglitazone (1 μM for 48 h).
Figure 3.
Figure 3.
Exogenous ligand is differentially required for coactivator recruitment to PPARγ _target genes in adipocytes. ChIP analysis of factor binding and histone acetylation on adipocyte aP2 and GyK genes in preadipocytes and adipocytes ± rosiglitazone treatment. (A) aP2. (B) GyK.
Figure 4.
Figure 4.
Differential recruitment of corepressors to the aP2 and GyK genes in adipocytes. (A) ChIP assay of adipocytes transiently transfected with GyK reporter gene as in Figure 1D. (B) Effect of siRNA knockdown of N-CoR and SMRT on basal and rosiglitazone-stimulated expression of GyK-luciferase in 3T3-L1 adipocytes. Lamin siRNA was used as control. (C) ChIP assay of endogenous GyK and aP2 genes in adipocytes ± rosiglitazone treatment. (D) ChIP re-IP using the supernatants and eluates from ChIP performed for the endogenous GyK gene as in C.(E) Western blotting showing the protein levels of N-CoR, SMRT, and HDAC3 in adipocytes and rosiglitazone-treated adipocytes. RAN is used as loading control. (F) Quantitative real-time PCR measurement of GyK and aP2 mRNA expression in adipocytes treated with vehicle, trichostatin A (TSA, 100 nM) (Shang et al. 2002), and sodium butyrate (NaB, 5 mM) for 48 h. (*) p < 0.01 vs. vehicle.
Figure 5.
Figure 5.
TZD induces PGC-1α and PGC-1α induces GyK in adipocytes. (A) Rosiglitazone treatment induces PGC-1α mRNA significantly in adipocytes. (*) p < 0.001 vs. vehicle. (B) Immunoblot analysis of adenoviral Myc-PGC-1α expression in adipocytes. RAN is used as loading control. (C) Adenoviral expression of myc-tagged PGC-1α increases GyK expression, but not aP2, as measured by quantitative real-time PCR. (*) p < 0.001 vs. Ad-βGal/vehicle; (**) p < 0.01 vs. Adv-PGC-1α/vehicle; (#) p < 0.05 vs. Adv/vehicle.
Figure 6.
Figure 6.
PGC-1α expression facilitates corepressor dismissal on the GyK promoter in adipocytes. (A) ChIP analysis for PPARγ and corepressors on endogenous GyK gene in adipocytes infected with adenoviral Myc-PGC-1α or β-galactosidase (βGal) ± rosiglitazone. (B) ChIP analysis for coactivators on endogenous GyK and aP2 genes in adipocytes infected with adenoviral Myc-PGC-1α or βGal ± rosiglitazone.
Figure 7.
Figure 7.
Model of the molecular mechanisms underlying differential ligand requirements for PPARγ activation in adipocytes. In the absence of exogenous PPARγ ligand, PPARγ recruits corepressors N-CoR/SMRT or coactivators to the GyK (A) and aP2 (C) promoters, respectively. Thus, the expression of GyK is repressed and the basal level of GyK is very low, but aP2 expression is high in mature adipocytes. (B) Treatment of adipocytes with rosiglitazone induces GyK in adipocytes by two mechanisms. The direct mechanism involves triggering a conformational change in PPARγ, causing corepressor release and coactivator recruitment. The indirect mechanism involves induction of PGC-1α, which destabilizes the binding of corepressors.
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Comment in

  • 'No, really, how do they work?'.
    Moore DD. Moore DD. Genes Dev. 2005 Feb 15;19(4):413-4. doi: 10.1101/gad.1294105. Genes Dev. 2005. PMID: 15713837 No abstract available.

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References

    1. Adams M., Reginato, M.J., Shao, D., Lazar, M.A., and Chatterjee, V.K. 1997. Transcriptional activation by PPARγ is inhibited by phosphorylation at a consensus mitogen-activated protein kinase site. J. Biol. Chem. 272: 5128–5132. - PubMed
    1. Armoni M., Kritz, N., Harel, C., Bar-Yoseph, F., Chen, H., Quon, M.J., and Karnieli, E. 2003. Peroxisome proliferator-activated receptor-γ represses GLUT4 promoter activity in primary adipocytes, and rosiglitazone alleviates this effect. J. Biol. Chem. 278: 30614–30623. - PubMed
    1. Camp H.S., Chaudhry, A., and Leff, T. 2001. A novel potent antagonist of peroxisome proliferator-activated receptor γ blocks adipocyte differentiation but does not revert the phenotype of terminally differentiated adipocytes. Endocrinology 142: 3207–3213. - PubMed
    1. Castillo G., Brun, R.P., Rosenfield, J.K., Hauser, S., Park, C.W., Troy, A.E., Wright, M.E., and Spiegelman, B.M. 1999. An adipogenic cofactor bound by the differentiation domain of PPARγ. EMBO J. 18: 3676–3687. - PMC - PubMed
    1. Chawla A., Schwarz, E.J., Dimaculangan, D.D., and Lazar, M.A. 1994. Peroxisome proliferator-activated receptor γ (PPARγ): Adipose predominant expression and induction early in adipocyte differentiation. Endocrinology 135: 798–800. - PubMed

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