PPARγ and the global map of adipogenesis and beyond

Abstract

Peroxisome proliferator-activated receptor γ (PPARγ) is a member of the nuclear receptor (NR) superfamily of ligand-dependent transcription factors (TFs) and function as a master regulator of adipocyte differentiation and metabolism. We review recent breakthroughs in the understanding of PPARγ gene regulation and function in the chromatin context. It is now clear that multiple TFs team up to induce PPARγ during adipogenesis, and that other TFs cooperate with PPARγ to ensure adipocyte-specific genomic binding and function. We discuss how this differs in other PPARγ-expressing cells such as macrophages and how these genome-wide mechanisms are preserved across species despite modest conservation of specific binding sites. These emerging considerations inform our understanding of PPARγ function as well as of adipocyte development and physiology.

Keywords: PPARγ; adipogenesis; chromatin; genome-wide analyses; transcriptional network.

Figure I for TEXT BOX 2. PPARγ domain structure and mechanism of binding to DNA

The main structural domains of the PPARγ protein are shown, including the N-terminal activation function 1 (AF1) domain, the DNA binding domain (DBD), and the C-terminal AF2 domain. PPARγ binds as a heterodimer with RXR to DNA sequences that conform to a consensus motif containing two imperfect direct repeats of the sequence AGGTCA, separated by a single nucleotide (DR1). The shown graphical representation of the PPARγ:RXR binding motif is based on the position weight matrix in the JASPAR database generated on PPARγ ChIP-seq data from 3T3-L1 cells [13]. Also shown is the conserved portion of the 5′ extension of the consensus motif. In the presence of agonist, the PPARγ AF2 domain facilitates agonist-dependent recruitment of coactivators and Mediator in exchange for corepressors, leading to increased expression of target genes. In the absence of agonist, the AF2 associates more strongly with corepressors. The AF2 domain is also responsible for agonist binding and heterodimerazation with RXR. The AF1 domain is mediates agonist-independent recruitment of coactivators and the Mediator complex.

Figure 1. Model for the establishment of the PPARγ transcriptional network during adipogenesis

Adipocyte differentiation proceeds through two waves of TF activation: factors in the first wave are induced by the adipogenic cocktail and collectively activate the second wave of TFs including PPARγ and C/EBPα. This process is associated with changes in chromatin accessibility such that a large number of sites become open (i.e. DNase I hypesensitive) in “hotspots” where TFs bind cooperatively. Such accessibility may be transient during early adipogenesis or persistent in mature adipocytes depending on the TFs that occupy the hotspots. Notably, C/EBPβ is able to bind to relatively ‘closed’ chromatin at the earliest stages of differentiation.

Figure 2. Model for the cell-type specific recruitment of PPARγ

The ability of PPARγ to access binding sites in the genome is limited within a given cell type, and may be defined by repressive mechanisms like chromatin silencing and active mechanisms such as cell type-specific recruitment of co-localizing TFs. For example, genes that are uniquely bound by PPARγ in macrophages (“macrophage genes”) contain features of chromatin silencing in adipocytes, such as histone 3 lysine 27 trimethylation (H3K27me3) and histone 3 lysine 9 dimethylation (H3K9me2), that make PPARγ binding sites in these regions inaccessible. In contrast, genes that are regulated by PPARγ in adipocytes (adipocyte genes) have greater DNA accessibility (DNase I Hypersensitivity) and signatures of active chromatin such as histone acetylation (H3K9ac/H3K27ac). The establishment of such putative enhancers may be partially due to binding of TFs that facilitate the recruitment of PPARγ such as C/EBPα/β in adipocytes and PU.1 and C/EBPα/β in macrophages, and ultimately collaborate in recruiting coactivators and chromatin remodelers.

Figure 3. Interspecies conservation of the PPARγ transcriptional network

Comparisons of PPARγ binding between mouse and human reveal extensive evolutionary conservation of regulatory networks but limited conservation of binding events. A binding event is “conserved” when a ChIP-seq peak is detected at orthologous sequences in both species, or “species-specific” when a ChIP-seq peak is detected only in one species. PPARγ binding sites with nearby C/EBPα binding are more likely to be conserved between species than sites only binding PPARγ. Gene regulation is considered “conserved” when a given gene is associated with PPARγ binding in both species, irrespective of whether binding sites are conserved or species-specific.