Molecular regulation of PPAR[gamma]/RXR[alpha] signaling by the novel cofactor ZFP407

Cofactors interacting with PPAR[gamma] can regulate adipogenesis and adipocyte metabolism by modulating the transcriptional activity and selectivity of PPAR[gamma] signaling. ZFP407 was previously demonstrated to regulate PPAR[gamma] target genes such as GLUT4, and its overexpression improved glucose homeostasis in mice. Here, using a series of molecular assays, including protein-interaction studies, mutagenesis, and ChIP-seq, ZFP407 was found to interact with the PPAR[gamma]/RXR[alpha] protein complex in the nucleus of adipocytes. Consistent with this observation, ZFP407 ChIP-seq peaks significantly overlapped with PPAR[gamma] ChIP-seq peaks, with more than half of ZFP407 peaks overlapping with PPAR[gamma] peaks. Transcription factor binding motifs enriched in these overlapping sites included CTCF, RAR[alpha]/RXR[gamma], TP73, and ELK1, which regulate cellular development and function within adipocytes. Site-directed mutagenesis of frequent PPAR[gamma] phosphorylation or SUMOylation sites did not prevent its regulation by ZFP407, while mutagenesis of ZFP407 domains potentially necessary for RXR and PPAR[gamma] binding abrogated any impact of ZFP407 on PPAR[gamma] activity. These data suggest that ZFP407 controls the activity of PPAR[gamma], but does so independently of post-translational modifications, likely by direct binding, establishing ZFP407 as a newly identified PPAR[gamma] cofactor. In addition, ZFP407 ChIP-seq analyses identified regions that did not overlap with PPAR[gamma] peaks. These non-overlapping peaks were significantly enriched for the transcription factor binding motifs of TBX19, PAX8, HSF4, and ZKSCAN3, which may contribute to the PPAR[gamma]-independent functions of ZFP407 in adipocytes and other cell types.