Genome-wide profiling of PPARγ:RXR binding and RNA polymerase II occupancy during adipogenesis

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Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor (NR) superfamily and important transcriptional regulators of metabolic pathways in several tissues including skeletal muscle, liver, and adipose tissue. PPARs bind to PPAR responsive elements (PPREs) in the genome as heterodimers with members of the retinoid X receptor (RXR) family. Binding of the PPAR/RXR heterodimer to a given PPRE is followed by coactivator recruitment and subsequent activation of transcription of the target gene controlled by that PPRE. One member of the family, PPARγ, is the master regulator of adipogenesis. The aim of this project is to decipher the molecular mechanisms underlying PPARγ action during adipogenesis and in mature adipocytes with special emphasis on the role of cofactors, specific histone modifications, and other transcription factors in transactivation of target genes by PPARγ.

To investigate PPARγ action in adipocytes, it is essential to know the genomic target sites of the PPARγ/RXR heterodimer, i.e. the sites in the genome, where the heterodimer can bind directly or indirectly to DNA. In collaboration with Prof. Henk Stunnenberg, Nijmegen, The Netherlands we have recently determined the genome-wide PPARγ/RXR binding site profile during differentiation of murine fibroblasts using a combination of chromatin immunoprecipitation (ChIP) and high throughput parallel sequencing (ChIP-seq). Interestingly, these data indicate that PPARγ cooperates with members of the CCAAT/enhancer binding protein (C/EBP) family, other key transcriptional regulators of adipogenesis, in the transcriptional regulation of several adipocyte specific genes. In this project I will investigate this putative cooperativity between PPARγ and C/EBPs in adipogenesis.

To investigate the molecular mechanisms underlying transcriptional regulation of target genes by PPARγ/RXR, and to understand what distinguishes this heterodimer from other PPAR/RXR heterodimers, it is necessary to know the identity of the transcriptional coregulators, which interact with and modulate the activity of PPARγ and RXR. Using a combination of optimized co-immunoprecipitation and high sensitivity mass spectrometry we aim to identify novel and PPARγ specific interaction partners in murine 3T3-L1 adipocytes. Preliminary results indicate that this experimental approach is ideal for identifying the components of transcriptionally active PPARγ/RXR complexes in adipocytes.

Periode14. sep. 2008
BegivenhedstitelGenome-wide profiling of PPARγ:RXR binding and RNA polymerase II occupancy during adipogenesis
ArrangørPhD skolen for molekylær metabolisme
PlaceringGl. Avernæs, Helnæsvej 9, 5631 Ebberup, DanmarkVis på kort