GSE99565
Retinoid X receptor activation alters the chromatin landscape ot commit mesenchymal stem cells to the adipose lineage
We set out to study the effects of the endocrine disrupting chemical tributyltin (TBT) on early lineage commitment of primary bone marrow-derived mesenchymal stem cells (MSCs) from the long bones of C57BL/6 mice. We previously showed that prenatal exposure to nanomolar levels of TBT results in increased adiposity in mice. TBT is an agonist of two nuclear receptors, peroxisome proliferator-activated receptor gamma (PPAR?) and retinoid X receptor (RXR), master regulators of adipogenesis. To test if TBT could influence early MSC lineage commitment we treated these cells with 50 nM TBT, 100 nM Rosiglitazone (ROSI, a pure PPAR? agonist), 100 nM AGN194204 (4204, a pure RXR agonist), and vehicle control (0.1% DMSO) for 48 hours prior to differentiation with a standard adipogenic cocktail. This experiment revealed an RXR-dependent commitment to the adipose lineage. To better understand how TBT induces adipose commimtment, we sequenced RNA from MSCs treated with DMSO (0.1%), ROSI (100 nM), 4204 (100 nM), and TBT (50 nM) for 48 hrs, prior to any differentiation. Unbiased hierarchical clustering analysis showed a clear separation between DMSO and ROSI replicates from 4204 and TBT, strengthening our conclusion that the observed phentype is RXR-dependent. Pathway analysis of differentially expressed genes revealed that TBT and 4204 de-repress targets of the repressive histone modifier Enhancer of zeste 2 (EZH2), the catalytic member of the Polycomb repressive complex 2 (PRC2), which deposits H3K27me3 on histones. We there for preformed ChIP-Seq analysis on MSCs treated with DMSO (0.1%) or TBT (50 nM) for 48 hrs, hypothesizing that TBT would reduce H3K27me3 in proximity to genes that regulate adipose lineage commitment. Overall design: We generated RNA-Seq data in undifferentiated MSCs treated with DMSO (0.1%), ROSI (100 nM), 4204 (100 nM), or TBT (50 nM) for 48 hrs. N=6 per treatment. We also generated H3K4me3 and H3K27me3 ChIP-Seq data in MSCs treated with DMSO (0.1%) or TBT (50 nM). N=4 per treatment, per histone mark, and N=1 input per treatment.