Androgen receptor requires an optimal level of condensation to assemble androgen-induced active enhancers [GRO-Seq]
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ABSTRACT:
PROVIDER: PRJNA889017 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR21853592.fastq.gz | Fastqsanger.gz | |||
| SRR21853593.fastq.gz | Fastqsanger.gz | |||
| SRR21853594.fastq.gz | Fastqsanger.gz | |||
| SRR25007861_1.fastq.gz | Fastqsanger.gz | |||
| SRR25007861_2.fastq.gz | Fastqsanger.gz |
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Project description:Transcription factors (TFs) activate enhancers to drive cell-specific gene expression in response to signals, but our understanding of enhancer assembly in signaling events is incomplete. Here we show that Androgen Receptor (AR), a steroid hormone signaling-regulated transcription factor, forms phase-separated condensates in response to androgen to activate enhancers. We demonstrate that the intrinsically disordered NTD of AR drives condensate formation and that NTD deletion or aromatic residue mutation reduces AR self-association and abolishes AR transcriptional activity. AR NTD can be substituted by some IDRs from other proteins for AR condensation capacity and transactivation function. Extending the polyQ tract within AR NTD strengthens AR phase separation and also leads to impaired transcriptional activity. PolyQ expansion does not affect AR binding on enhancers, but instead impairs enhancer assembly. These results suggest that AR phase separation mediates enhancer assembly, and an optimal level of AR condensation is required for its proper function in mediating AR-AR homotypic and AR-cofactor heterotypic interactions to regulate transcription in response to signals. Our study supports that alteration of the fine-tuned protein condensation might underlie AR-related human pathologies, therefore providing novel molecular insights for potential therapeutic strategies to treat prostate cancer and other AR-involved diseases by targeting AR condensation.
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Project description:Androgen receptor requires an optimal level of condensation to assemble androgen-induced active enhancers [ChIP-Seq]
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Project description:Transcription factors (TFs) activate enhancers to drive cell-specific gene expression in response to signals, but our understanding of enhancer assembly in signaling events is incomplete. Here we show that Androgen Receptor (AR), a steroid hormone signaling-regulated transcription factor, forms phase-separated condensates in response to androgen to activate enhancers. We demonstrate that the intrinsically disordered NTD of AR drives condensate formation and that NTD deletion or aromatic residue mutation reduces AR self-association and abolishes AR transcriptional activity. AR NTD can be substituted by some IDRs from other proteins for AR condensation capacity and transactivation function. Extending the polyQ tract within AR NTD strengthens AR phase separation and also leads to impaired transcriptional activity. PolyQ expansion does not affect AR binding on enhancers, but instead impairs enhancer assembly. These results suggest that AR phase separation mediates enhancer assembly, and an optimal level of AR condensation is required for its proper function in mediating AR-AR homotypic and AR-cofactor heterotypic interactions to regulate transcription in response to signals. Our study supports that alteration of the fine-tuned protein condensation might underlie AR-related human pathologies, therefore providing novel molecular insights for potential therapeutic strategies to treat prostate cancer and other AR-involved diseases by targeting AR condensation.
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Project description:Transcription factors (TFs) activate enhancers to drive cell-specific gene expression in response to signals, but our understanding of enhancer assembly in signaling events is incomplete. Here we show that Androgen Receptor (AR), a steroid hormone signaling-regulated transcription factor, forms phase-separated condensates in response to androgen to activate enhancers. We demonstrate that the intrinsically disordered NTD of AR drives condensate formation and that NTD deletion or aromatic residue mutation reduces AR self-association and abolishes AR transcriptional activity. AR NTD can be substituted by some IDRs from other proteins for AR condensation capacity and transactivation function. Extending the polyQ tract within AR NTD strengthens AR phase separation and also leads to impaired transcriptional activity. PolyQ expansion does not affect AR binding on enhancers, but instead impairs enhancer assembly. These results suggest that AR phase separation mediates enhancer assembly, and an optimal level of AR condensation is required for its proper function in mediating AR-AR homotypic and AR-cofactor heterotypic interactions to regulate transcription in response to signals. Our study supports that alteration of the fine-tuned protein condensation might underlie AR-related human pathologies, therefore providing novel molecular insights for potential therapeutic strategies to treat prostate cancer and other AR-involved diseases by targeting AR condensation.
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Project description:While thousands of long non-coding RNAs (lncRNAs) are expressed in higher eukaryotes, the potential regulatory roles of lncRNAs in regulated gene transcription programs remain rather poorly understood. Here, we report that two lncRNAs highly overexpressed in aggressive prostate cancer, PRNCR1 and PCGEM1, bind successively to the androgen receptor (AR) and strongly enhance both ligand-dependent and ligand-independent AR-mediated gene activation programs and proliferation in prostate cancer cells. Binding of PRNCR1 to the C-terminally acetylated AR on enhancers and its association with DOT1L appear to be required for recruitment of the second lncRNA, PCGEM, to the N-terminally methylated AR. Unexpectedly, recognition of the H3K4me3 promoter mark by the PHD finger-domain of Pygopus2, recruited by PCGEM1, proves to enhance selective looping of AR-bound enhancers to target gene promoters in these cells, revealing a novel aspect of ligand-induced enhancer-promoter interactions. In “resistant” prostate cancer cells, these overexpressed lncRNAs can interact with, and are required for the robust activation of both truncated and full length AR, causing DHT-independent activation of the AR transcriptional program and cell proliferation. Conditionally-expressed short hairpin RNA (shRNA)-mediated targeting of these lncRNAs in these resistant cancer cell lines strongly suppressed xenograft growth in vivo. Together, these results suggest that these overexpressed lncRNAs can potentially serve as a required component of castration-resistance in prostatic tumors. Global Run On (GRO) assay followed by high throughput sequencing (GRO-seq); after knocking-down lincRNAs PCGEM1 and PRNCR1. LNCaP cells were grown to 30-50% confluence and siRNA/ASO transfections were carried out using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions. Control samples were transfected with scramble ASO and control siRNA, respectively. On the following day of transfection, the cells were cultured in UltraCULTURE (Phenol red free) + 5% Charcoal Dextran Stripped (CDS) serum for 48 hours. For androgen induction, we treat cells with DHT from a 100 uM stock in 70% ethanol to a final concentration of 100 nM for 1 hour Scramble ASO, -DHT Scramble ASO, +DHT PRNCR1 ASO, -DHT PRNCR1 ASO, +DHT PCGEM1 ASO, -DHT PCGEM1 ASO, +DHT
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