BackgroundCells have developed many ways to cope with external stress. One distinctive feature in acute proteotoxic stresses, such as heat shock (HS), is rapid post-translational modification of proteins by SUMOs (small ubiquitin-like modifier proteins; SUMOylation). While many of the SUMO targets are chromatin proteins, there is scarce information on chromatin binding of SUMOylated proteins in HS and the role of chromatin SUMOylation in the regulation of transcription.ResultsWe mapped HS-induced genome-wide changes in chromatin occupancy of SUMO-2/3-modified proteins in K562 and VCaP cells using ChIP-seq. Chromatin SUMOylation was further correlated with HS-induced global changes in transcription using GRO-seq and RNA polymerase II (Pol2) ChIP-seq along with ENCODE data for K562 cells. HS induced a rapid and massive rearrangement of chromatin SUMOylation pattern: SUMOylation was gained at active promoters and enhancers associated with multiple transcription factors, including heat shock factor 1. Concomitant loss of SUMOylation occurred at inactive intergenic chromatin regions that were associated with CTCF-cohesin complex and SETDB1 methyltransferase complex. In addition, HS triggered a dynamic chromatin binding of SUMO ligase PIAS1, especially onto promoters. The HS-induced SUMOylation on chromatin was most notable at promoters of transcribed genes where it positively correlated with active transcription and Pol2 promoter-proximal pausing. Furthermore, silencing of SUMOylation machinery either by depletion of UBC9 or PIAS1 enhanced expression of HS-induced genes.ConclusionsHS-triggered SUMOylation targets promoters and enhancers of actively transcribed genes where it restricts the transcriptional activity of the HS-induced genes. PIAS1-mediated promoter SUMOylation is likely to regulate Pol2-associated factors in HS.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-015-0717-y) contains supplementary material, which is available to authorized users.

Androgen receptor (AR) is a ligand-activated transcription factor that plays a central role in the development and growth of prostate carcinoma. PIAS1 is an AR- and SUMO-interacting protein and a putative transcriptional coregulator overexpressed in prostate cancer. To study the importance of PIAS1 for the androgen-regulated transcriptome of VCaP prostate cancer cells, we silenced its expression by RNAi. Transcriptome analyses revealed that a subset of the AR-regulated genes is significantly influenced, either activated or repressed, by PIAS1 depletion. Interestingly, PIAS1 depletion also exposed a new set of genes to androgen regulation, suggesting that PIAS1 can mask distinct genomic loci from AR access. In keeping with gene expression data, silencing of PIAS1 attenuated VCaP cell proliferation. ChIP-seq analyses showed that PIAS1 interacts with AR at chromatin sites harboring also SUMO2/3 and surrounded by H3K4me2; androgen exposure increased the number of PIAS1-occupying sites, resulting in nearly complete overlap with AR chromatin binding events. PIAS1 interacted also with the pioneer factor FOXA1. Of note, PIAS1 depletion affected AR chromatin occupancy at binding sites enriched for HOXD13 and GATA motifs. Taken together, PIAS1 is a genuine chromatin-bound AR coregulator that functions in a target gene selective fashion to regulate prostate cancer cell growth.

The biologically most active vitamin D compound, 1α,25-dihydroxyvitamin D₃ (1α,25(OH)₂D₃), influences the status of inflammation by modulating the expression of several cytokine genes. In this study, we have examined the mechanism of transcriptional regulation of interleukin 10 (IL-10) by 1α,25(OH)₂D₃ in lipopolysaccharide (LPS)-treated human monocytes (THP-1). Quantitative PCR showed that IL-10 mRNA expression was significantly down-regulated (2.8-fold) during the first 8h of 1α,25(OH)₂D₃ treatment, while after 48 h it was up-regulated (3-fold). Gel shift and quantitative chromatin immunoprecipitation (ChIP) assays showed that the vitamin D receptor (VDR) binds in a cyclical fashion to a promoter region 1500-1700 bp upstream of the IL-10 transcription start site (TSS) containing two conserved VDR binding sites. Targeting of VDR binding sites by enhancer specific duplex RNAs revealed that only the more distal element is functional and chromosome conformation capture analysis suggested that this region loops 1α,25(OH)₂D₃-dependently to the TSS. Quantitative ChIP and micrococcal nuclease assays also revealed 1α,25(OH)₂D₃-dependent cyclical epigenetic changes and nucleosome remodeling at this promoter region. In conclusion, in LPS-treated THP-1 cells the primary effect of 1α,25(OH)₂D₃ on IL-10 expression is down-regulation, which is achieved via a cyclical recruitment of VDR to the promoter.

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Interleukin 12 (IL-12) is a heterodimeric, pro-inflammatory cytokine that plays a central role in activation and differentiation of CD4(+) T cells into interferon-γ secreting T-helper type 1 cells. IL-12B, a gene encoding the larger subunit of active IL-12, has been reported to be down-regulated by the nuclear hormone 1α,25-dihydroxyvitamin D(3) (1α,25(OH)(2)D(3)), but the mechanism of the regulation is unknown. In this study, we have examined the molecular mechanism of transcriptional regulation of the IL-12B gene by 1α,25(OH)(2)D(3) in lipopolysaccharide (LPS)-treated human monocytes (THP-1). Quantitative RT-PCR showed that IL-12B mRNA displays a cyclical expression profile and is down-regulated 2.8-fold during the first 8h and even 12.1-fold 24h after exposure to 1α,25(OH)(2)D(3). Gel shift and quantitative chromatin immunoprecipitation (ChIP) assays demonstrated vitamin D receptor (VDR) binding to genomic regions 480 and 6300bp upstream of the IL-12B transcription start site (TSS). Quantitative ChIP assays also revealed that together with VDR and its partner RXR the above regions recruited the co-repressor NCOR2/SMRT and histone deacetylase 3 leading to a decreased histone 4 acetylation and increased histone 3 trimethylation at the IL-12B promoter and its TSS. We suggest that these repressive epigenetic changes eventually cause down-regulation of IL-12 expression. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.