Comparative Transcriptomic and Epigenomic Analyses Reveal New Regulators of Murine Brown Adipogenesis

Abstract

Increasing energy expenditure through brown adipocyte recruitment is a promising approach to combat obesity. We report here the comprehensive profiling of the epigenome and transcriptome throughout the lineage commitment and differentiation of C3H10T1/2 mesenchymal stem cell line into brown adipocytes. Through direct comparison to datasets from differentiating white adipocytes, we systematically identify stage- and lineage-specific coding genes, lncRNAs and microRNAs. Utilizing chromatin state maps, we also define stage- and lineage-specific enhancers, including super-enhancers, and their associated transcription factor binding motifs and genes. Through these analyses, we found that in brown adipocytes, brown lineage-specific genes are pre-marked by both H3K4me1 and H3K27me3, and the removal of H3K27me3 at the late stage is necessary but not sufficient to promote brown gene expression, while the pre-deposition of H3K4me1 plays an essential role in poising the brown genes for expression in mature brown cells. Moreover, we identify SOX13 as part of a p38 MAPK dependent transcriptional response mediating early brown cell lineage commitment. We also identify and subsequently validate PIM1, SIX1 and RREB1 as novel regulators promoting brown adipogenesis. Finally, we show that SIX1 binds to adipogenic and brown marker genes and interacts with C/EBPα, C/EBPβ and EBF2, suggesting their functional cooperation during adipogenesis.

Regulation of white and brown adipocyte differentiation by RhoGAP DLC1

“…Currently the list of regulators of white, brown and beige adipogenic differentiation is still far from complete. And bioinformatics approaches are increasingly utilized to identify novel regulators of adipogenesis [ 14 – 17 ]. Among these approaches, the super-enhancer (SE) association analysis has emerged as a useful tool in the search of novel adipogenic factors.…”

“…SEs are also found to be associated with genes that define cell identity and this feature provides the mechanistic basis of the SE association analysis. Through this analysis, we identified RREB1 and PIM1 as new factors promoting brown adipogenesis [ 14 ], while KLF11 was identified as an essential regulator of human white fat browning [ 15 ].…”

miRNA-32 Drives Brown Fat Thermogenesis and Trans-activates Subcutaneous White Fat Browning in Mice

“…BAT and iWAT SVF cells were prepared as described previously (Brunmeir et al, 2016). Inhibition of miR-32 was performed using either 40 or 80 nM of mIRCURY locked nucleic acid (LNA) microRNA Inhibitors (Exiqon).…”

“…They can be defined by the occupancy of mediator and cell-type-specific transcription factors, as well as chromatin marks such as histone H3K27ac and H3K4me1 (Whyte et al, 2013). Through the super-enhancer association analysis, we have recently identified RREB1 and PIM1 as important factors promoting brown adipogenesis (Brunmeir et al, 2016), while KLF11 was previously identified as an essential regulator of human white fat browning (Loft et al, 2015). …”

Regulation of Thermogenic Adipocyte Differentiation and Adaptive Thermogenesis Through Histone Acetylation

“…Of the aforementioned epigenetics alterations, an increasing number of studies have pointed to the effects of histone modifications-in particular, histone acetylation-on adipocyte differentiation and adipogenesis (30,(36)(37)(38)(39)(40). Following which, in the recent years, the role of histone acetyltransferases (HATs) and histone deacetylases (HDACs) in the regulation of the development and function of thermogenic adipocytes have garnered remarkable interest from scientists and clinicians for their therapeutic potential in ameliorating obesity and its complications.…”