PPARγ and Wnts are master positive and negative regulators of adipogenesis, respectively. The H3K4 methylation regulator PTIP promotes PPARγ expression while the H3K27 methyltransferase Ezh2 represses Wnt expression during adipogenesis (). However, the epigenetic factors that repress PPARγ but promote Wnt expression in preadipocytes have not been identified. Analyzing the enrichment and/or the dynamic changes of histone methylation patterns on PPARγ and Wnt promoters in preadipocytes and in the early phase of adipogenesis may provide clues to solving this issue.
It has been reported that knockdown of histone demethylase LSD1 increases histone H3K9 dimethylation (H3K9me2) on C/EBPα
promoter and leads to decreased adipogenesis. Conversely, knockdown of histone H3K9 methyltransferase SetDB1 (also known as ESET) decreases H3K9me2 on C/EBPα
promoter and leads to increased adipogenesis. These results suggest opposing roles of LSD1 and SetDB1 in regulating C/EBPα
expression and adipogenesis [42
]. However, LSD1 mainly demethylates H3K4me1/2 and SetDB1 mainly performs trimethylation on histone H3K9 [7
]. The H3K9me2 level in cells is predominantly controlled by the euchromatin-associated H3K9 methyltransferase G9a [44
]. Thus, it will be important to determine whether G9a-mediated H3K9me2 plays any role in regulation of PPARγ
expression and adipogenesis.
Within hours of initiation of adipogenesis, cAMP-induced pCREB and Dex-bound GR rapidly induce expression of early adipogenic transcription factors C/EBPβ, KLF4, Krox20 and C/EBPδ, which are essential for induction of PPARγ and C/EBPα expression and for adipogenesis (). How histone methylations regulate the rapid induction of these early adipogenic transcription factors is completely unknown.
Another important question is how H3K4 or H3K27 methylation targets PPARγ or Wnt genes, respectively. In other words, how H3K4 methyltransferase complexes are recruited to PPARγ genes and how H3K27 methyltransferase complex PRC2 is recruited to Wnt genes are unclear. These histone modifying complexes may be recruited by sequence-specific transcription factors that directly bind to PPARγ or Wnt gene loci. Alternatively, sequence-specific non-coding RNAs may directly recruit these HMT complexes to PPARγ or Wnt genes. Since subunits of these histone modifying complexes contain multiple histone modification-binding domains, it is possible that these HMT complexes are recruited to PPARγ or Wnt genes by recognizing pre-existing histone modifications on target genes, i.e. through cross-talk between histone modifications.
PPARγ is a nuclear receptor and thus a ligand-activated transcription factor. Correlating with ligand-induced nuclear receptor target gene activation, ligand induces sequential enrichment of H3K18/27ac, RNA Pol II, and several histone methylations including H3K4me3, H3K36me3 and H3K79me2 on nuclear receptor target genes [5
]. While the exact roles of these gene activation-associated histone methylations remain to be determined, gene repression-associated histone methylations have been implicated in regulating nuclear receptor target gene expression [45
]. One good example is the role of histone methyltransferase SetDB1, which is activated by non-canonical Wnt signaling to methylate histone H3K9 to repress target gene activation by PPARγ [46
]. The contributions of other site-specific histone methylations to the transcriptional activation or repression of PPARγ target genes important for adipogenesis remain to be defined.
The majority of histone methyltransferases and demethylases were identified in the 21st
century and their biological functions are poorly understood. Adipogenesis provides an excellent model system to study the roles of histone methyltransferases and demethylases, and the dynamics of site-specific histone methylations, in regulation of gene expression and cell differentiation. In addition to histone H3K4 and H3K27 methylations described above, it will be important to understand the roles of methylations on histone H3K9, H3K36 and H3K79, and the related histone methyltransferases and demethylases, in regulation of adipogenesis. Straightforward knockout (KO) of these enzymes usually leads to embryonic lethality. Fortunately, conditional KO mouse strains (floxed mice) for many of these histone modifying enzymes and associated factors are becoming available. Preadipocytes carrying conditional KO of histone modifying enzymes can be easily isolated from these mice, thus providing an excellent model system to study epigenetic regulation of cell differentiation in vitro
]. The results should be verified in vivo
by crossing floxed mice with tissue-specific Cre mice to specifically delete the gene-of-interest in adipose tissue [47