The vitamin D receptor (VDR) is expressed in human adipocytes and is transiently induced during early adipogenesis in mesenchymal progenitor cell models. VDR null mice exhibit enhanced energy expenditure and reduced adiposity even when fed high fat diets. Adipocyte-specific transgenic-expression of human VDR in mice enhances adipose tissue mass, indicating that VDR activation in adipocytes enhances lipid storage in vivo. In these studies, we conducted genomic profiling and differentiation assays in primary cultures of human adipose-derived mesenchymal progenitor cells to define the role of the VDR and its ligand 1,25-dihydroxyvitamin D3 (1,25D) in adipogenesis. In the presence of adipogenic media, 1,25D promoted lipid accumulation and enhanced the expression of FABP4, FASN, and PPARγ. Mesenchymal cells derived from 6-month old VDR null mice exhibited impaired adipogenesis ex vivo but differentiation was restored by stable expression of human VDR. STEAP4, a gene that encodes a metalloreductase linked to obesity, insulin sensitivity, metabolic homeostasis and inflammation, was highly induced in human adipose cells differentiated in the presence of 1,25D but was minimally affected by 1,25D in undifferentiated precursors. These studies provide a molecular basis for recent epidemiological associations between vitamin D status, body weight and insulin resistance which may have relevance for prevention or treatment of metabolic syndrome and obesity.

The Ty1 retrotransposons present in the genome of Saccharomyces cerevisiae belong to the large class of mobile genetic elements that replicate via an RNA intermediary and constitute a significant portion of most eukaryotic genomes. The retromobility of Ty1 is regulated by numerous host factors, including several subunits of the Mediator transcriptional co-activator complex. In spite of its known function in the nucleus, previous studies have implicated Mediator in the regulation of post-translational steps in Ty1 retromobility. To resolve this paradox, we systematically examined the effects of deleting non-essential Mediator subunits on the frequency of Ty1 retromobility and levels of retromobility intermediates. Our findings reveal that loss of distinct Mediator subunits alters Ty1 retromobility positively or negatively over a >10,000-fold range by regulating the ratio of an internal transcript, Ty1i, to the genomic Ty1 transcript. Ty1i RNA encodes a dominant negative inhibitor of Ty1 retromobility that blocks virus-like particle maturation and cDNA synthesis. These results resolve the conundrum of Mediator exerting sweeping control of Ty1 retromobility with only minor effects on the levels of Ty1 genomic RNA and the capsid protein, Gag. Since the majority of characterized intrinsic and extrinsic regulators of Ty1 retromobility do not appear to effect genomic Ty1 RNA levels, Mediator could play a central role in integrating signals that influence Ty1i expression to modulate retromobility.

The Ty1 retrotransposon family is maintained in a functional but dormant state by its host, Saccharomyces cerevisiae. Several hundred RHF and RTT genes encoding co-factors and restrictors of Ty1 retromobility, respectively, have been identified. Well-characterized examples include MED3 and MED15, encoding subunits of the Mediator transcriptional co-activator complex; control of retromobility by Med3 and Med15 requires the Ty1 promoter in the U3 region of the long terminal repeat. To characterize the U3-dependence of other Ty1 regulators, we screened a library of 188 known rhf and rtt mutants for altered retromobility of Ty1his3AI expressed from the strong, TATA-less TEF1 promoter or the weak, TATA-containing U3 promoter. Two classes of genes, each including both RHFs and RTTs, were identified. The first class comprising 82 genes that regulated Ty1his3AI retromobility independently of U3 is enriched for RHF genes that restrict the G1 phase of the cell cycle and those involved in transcriptional elongation and mRNA catabolism. The second class of 51 genes regulated retromobility of Ty1his3AI driven only from the U3 promoter. Nineteen U3-dependent regulators (U3DRs) also controlled retromobility of Ty1his3AI driven by the weak, TATA-less PSP2 promoter, suggesting reliance on the low activity of U3. Thirty-one U3DRs failed to modulate PPSP2-Ty1his3AI retromobility, suggesting dependence on the architecture of U3. To further investigate the U3-dependency of Ty1 regulators, we developed a novel fluorescence-based assay to monitor expression of p22-Gag, a restriction factor expressed from the internal Ty1i promoter. Many U3DRs had minimal effects on levels of Ty1 RNA, Ty1i RNA or p22-Gag. These findings uncover a role for the Ty1 promoter in integrating signals from diverse host factors to modulate Ty1 RNA biogenesis or fate.