Divergent Transcriptional Regulation of Suppressors of Cytokine Signaling Genes in Adipocytes

Abstract

The Janus Kinase - Signal Transducer and Activator of Transcription (JAK-STAT) signaling pathway transduces several signals crucial for development and homeostasis. Suppressors of cytokine signaling (SOCS) proteins control JAK-STAT signaling via a negative feedback loop. The transcription factor STAT5 is known to play a significant role in fat cell development and function, and several studies suggest that acetylation may affect STAT5 transcriptional activity. To test this hypothesis, we treated 3T3-L1 adipocytes with growth hormone (GH) to activate STAT5 in the presence or absence of histone deacetylase (HDAC) inhibitors. STAT5 acetylation levels were low in adipocytes and mostly unchanged by the inhibitors. Still, two STAT5 target genes from the SOCS family, Socs3 and Cish, were inversely regulated by general and specific HDAC inhibitors (Socs3 expression increased, while Cish levels decreased). Chromatin immunoprecipitation analyses revealed that changes in total and activated RNA polymerase II, but not STAT5A binding to Socs3 and Cish promoters highly correlated with changes in gene expression. Thus, we hypothesized that HDAC inhibitors were indirectly affecting another protein in the transcriptional complex. Members of the bromodomain and extra-terminal (BET) protein family bind acetylated histones and recruit transcription factors, thus playing a role in chromatin remodeling and transcription. Treatment with the BET inhibitor JQ1 produced the same divergent effects as HDAC inhibition on both Socs3 and Cish gene expression, as well as on RNA polymerase II binding. Moreover, BET proteins help drive productive elongation of mRNA by recruiting the positive transcription elongation factor (P-TEFb). We found that JQ1, but not the HDAC inhibitor LMK-235, could impact P-TEFb availability in a manner consistent with the Socs3 gene expression changes we observed. We propose a model in which GH-induced Cish transcription is dependent on the BET protein, BRD2, and susceptible to inhibition by JQ1 (and indirectly by HDAC inhibitors), whereas Socs3 mRNA elongation may involve recruitment of different factors, thus explaining the divergent effects of HDAC/BET inhibition on the two genes. Overall, our results demonstrate substantially different transcriptional regulation of Socs3 and Cish, suggesting distinct roles for these two related proteins in adipocytes.