PPARγ is a key regulator of glucose homeostasis and insulin sensitization. PPARγ must heterodimerize with its dimeric partner, the retinoid X receptor (RXR), to bind DNA and associated coactivators such as p160 family members or PGC-1α to regulate gene networks. To understand how coactivators are recognized by the functional heterodimer PPARγ/RXRα and to determine the topological organization of the complexes, we performed a structural study using small angle X-ray scattering of PPARγ/RXRα in complex with DNA from regulated gene and the TIF2 receptor interacting domain (RID). The solution structures reveal an asymmetry of the overall structure due to the crucial role of the DNA in positioning the heterodimer and indicate asymmetrical binding of TIF2 to the heterodimer. 1. Introduction PPARγ, a member of the nuclear receptor family, is a key regulator of adipocyte differentiation and is involved in glucose homeostasis and insulin sensitization (reviewed in [1, 2]). PPARγ together with the CCAAT/enhancer-binding proteins had been identified as key transcription factors of driving fat cell differentiation (reviewed in [3]). PPARγ is absolutely required for both white and brown fat cell development. Several PPARγ coregulators have also been shown to affect positively or negatively this differentiation (reviewed in [4]). PPARγ is activated through the binding of diverse ligands including natural fatty acid derivatives and nonsteroidal drugs and is the target of therapeutically active antidiabetics such as rosiglitazone (reviewed in [5]). Furthermore, cdk-5 phosphorylation of PPARγ leads to deregulation of some genes involved in metabolism [6]. The actions of PPARγ are mediated by 2 isoforms that result from alternative splicing. PPARγ2 is 28 amino acids longer at the N-terminal end (Figure 1(a)) and is mainly expressed in adipocyte cells, while PPARγ1 is ubiquitously expressed. Interestingly, PPARγ2 is ten times more active in ligand-independent transcriptional activation than PPARγ1 [7, 8]. Figure 1: Biophysical characterization of the stoichiometry of the TIF2 RID/PPAR γ/RXR complexes. (a) Structural organization of hPPAR γ1, hPPAR γ2, and hTIF2. (b) ESI mass spectra of TIF2 RID/PPAR γ/RXR LBDs recorded under nondenaturing conditions in 200？mM ammonium acetate at . The different charge states of the proteins are indicated above the peaks. The calculated molecular mass of the first peak corresponds to PPAR γ/RXR α LBDs and the second one to the complex containing one PPAR γ/RXR α LBDs dimer and one TIF2 RID. (c) Sedimentation equilibrium experiments. Best
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