Project description:Androgen signaling through the androgen receptor (AR) regulates multiple pathways in both normal and prostate cancer cells. Androgen regulates diverse aspects of the AR life cycle, including its post-translational modification, but understanding how specific modifications influence AR activity has been mostly elusive. Here, we show that androgen regulates AR through a pathway mediated by the mono-ADP ribosyltransferase, Parp7. We show that Parp7 ADP-ribosylates AR on multiple cysteines, and that a subset of these sites mediates agonist-specific recruitment of the E3 ligase Dtx3L/Parp9. Tandem macrodomains in Parp9 selectively recognize ADP ribosylated AR, and Dtx3L/Parp9 affects expression of a subset of AR-regulated genes. Parp7, ADP-ribosylation of AR, and AR-Dtx3L/Parp9 complex assembly are inhibited by 60 Olaparib, a compound used clinically to inhibit poly-ADP-ribosyltransferases Parp1/2. Our study reveals the components of a new androgen signaling axis that uses a writer and reader of ADP-ribosylation to modulate AR activity.
Project description:Androgen signaling through the androgen receptor (AR) regulates multiple pathways in both normal and prostate cancer cells. Androgen regulates diverse aspects of the AR life cycle, including its post-translational modification, but understanding how specific modifications influence AR activity has been mostly elusive. Here, we show that androgen regulates AR through a pathway mediated by the mono-ADP ribosyltransferase, Parp7. We show that Parp7 ADP-ribosylates AR on multiple cysteines, and that a subset of these sites mediates agonist-specific recruitment of the E3 ligase Dtx3L/Parp9. Tandem macrodomains in Parp9 selectively recognize ADP ribosylated AR, and Dtx3L/Parp9 affects expression of a subset of AR-regulated genes. Parp7, ADP-ribosylation of AR, and AR-Dtx3L/Parp9 complex assembly are inhibited by 60 Olaparib, a compound used clinically to inhibit poly-ADP-ribosyltransferases Parp1/2. Our study reveals the components of a new androgen signaling axis that uses a writer and reader of ADP-ribosylation to modulate AR activity.
Project description:We investigated the composition of chromatin protein network around endogenous androgen receptor (AR) in VCaP castration resistant prostate cancer cells using recently developed chromatin-directed proteomic approach called ChIP-SICAP . The androgen-induced AR chromatin protein network contained expected TFs, e.g. HOXB13, chromatin remodeling proteins, e.g. SMARCA4, and several novel candidates not previously associated with AR, e.g. prostate cancer biomarker SIM2. Based on these findings, the role of SMARCA4 and SIM2 was further characterized at AR chromatin domains . Silencing of SIM2 altered chromatin accessibility at a similar number of AR-binding sites as SMARCA4, an established ATPase subunit of the BAF chromatin remodeling complex, often aberrantly expressed in prostate cancer. Despite the wide co-occurrence on chromatin of SMARCA4 and AR, depletion of SMARCA4 influenced chromatin accessibility and expression of a restricted set of AR target genes, in particular those involved in cell morphogenetic changes in epithelial-mesenchymal transition. Silencing of SIM2, in turn, affected the expression of a much larger group of androgen-regulated genes, e.g. those involved in cellular responses to external stimuli and steroid hormone stimulus. The silencing also reduced proliferation of VCaP cells and tumor size in chick embryo chorioallantoic membrane assay, further suggesting the importance of SIM2 in the regulation prostate cancer cells.
Project description:The spliced variant forms of androgen receptor (AR-Vs) have been identified recently in castration-resistant prostate cancer (CRPC) cell lines and clinical samples. Here we identified the cistrome and transcriptome landscape of AR-Vs in CRPC cell lines and determine the clinical significance of AR variants regulated gene.The AR variants binding sites can be identified in 22Rv1 cell line in the absence of androgen. Knocking down full-length AR (AR-FL) doesn't affect AR-Vs binding sites in genome-wide. A set of genes were identified to be regulated uniquely by AR-Vs, but not by AR-FL in androgen-depleted condition. Integrated analysis showed that some genes may be modulated by AR-Vs directly. Unsupervised clustering analysis demonstrated that AR variants gene signature can separate not only the benign and malignant prostate tissue, but also the localized prostate cancer and metastatic CRPC specimens. Some genes modulated uniquely by AR variants were also identified to correlate with the Gleason Pattern of prostate cancer and PSA failure. We conclude that AR spliced variants bind to DNA independent of full-length AR, and can modulate a unique set of genes which is not regulated by full-length AR in the absence of androgen. AR variants gene signature correlate with CRPC and prostate cnacer disease progress. Androgen receptor (AR) binding sites in human prostate cancer 22Rv1 cell lines were studied using ChIP-seq. ChIP enriched and input DNA were sequenced using Illumina HiSeq 2000.
Project description:Prostate cancer is dependent on androgen receptor (AR) signaling at all stages of the disease and cyclin D1 has been shown to negatively modulate the expression of the AR-dependent gene prostate specific antigen (KLK3/PSA). Unbiased gene expression profiling was performed to determine the androgen-regulated geneset that is sensitive to cyclin D1. Androgen-sensitive LNCaP cells were straved of androgen, then transduced with control green fluorescent protein (GFP) or cyclin D1 (CCND1) adenovirus, then subsequently stimulated with control (Ethanol) or dihydrotestosterone (DHT).
Project description:Prostate cancer is dependent on androgen receptor (AR) signaling at all stages of the disease and cyclin D1 has been shown to negatively modulate the expression of the AR-dependent gene prostate specific antigen (KLK3/PSA). Unbiased gene expression profiling was performed to determine the androgen-regulated geneset that is sensitive to cyclin D1.
Project description:The spliced variant forms of androgen receptor (AR-Vs) have been identified recently in castration-resistant prostate cancer (CRPC) cell lines and clinical samples. Here we identified the cistrome and transcriptome landscape of AR-Vs in CRPC cell lines and determine the clinical significance of AR variants regulated gene.The AR variants binding sites can be identified in 22Rv1 cell line in the absence of androgen. Knocking down full-length AR (AR-FL) doesn't affect AR-Vs binding sites in genome-wide. A set of genes were identified to be regulated uniquely by AR-Vs, but not by AR-FL in androgen-depleted condition. Integrated analysis showed that some genes may be modulated by AR-Vs directly. Unsupervised clustering analysis demonstrated that AR variants gene signature can separate not only the benign and malignant prostate tissue, but also the localized prostate cancer and metastatic CRPC specimens. Some genes modulated uniquely by AR variants were also identified to correlate with the Gleason Pattern of prostate cancer and PSA failure. We conclude that AR spliced variants bind to DNA independent of full-length AR, and can modulate a unique set of genes which is not regulated by full-length AR in the absence of androgen. AR variants gene signature correlate with CRPC and prostate cnacer disease progress.
Project description:Androgen receptor (AR) orchestrates an intricate transcriptional regulatory network that governs prostate cancer initiation, development and progression. To understand this network in detail, we generated genome-wide maps of AR occupancy by ChIP-seq in LNCaP cells. We found NKX3-1, an androgen-dependent homeobox protein well-characterized for its role in prostate development and differentiation, being recruited to AR binding sites (ARBS) in response to androgen signaling. We identified 6,359 NKX3-1 binding sites, most of which overlapped with AR. In addition to its novel collaborative transcriptional role at well-known prostate cancer model genes, our binding and knockdown studies further suggested that NKX3-1 potentially regulates AR in a feed-forward manner. Integrative analysis of Oncomine molecular concepts showed that these androgen-regulated AR and NKX3-1 associated genes are significantly overexpressed in prostate carcinoma as well as advanced and recurrent prostate tumors. From our transcriptomic profiling and Gene Ontology analysis, we observed that AR and NKX3-1 co-regulate genes involved in "protein trafficking" processes, which are mandatory events in the integration of oncogenic signaling pathways leading to prostate cancer development and progression. Interestingly, we found that AR and NKX3-1 co-regulate several members of the RAB GTPase family of secretory/trafficking proteins via the involvement of FoxA1 in a ternary complex and we believe that these AR/NKX3-1/FoxA1 co-regulated RAB genes could serve as expression signatures in prostate carcinogenesis. More specifically, through functional analyses, we showed that NKX3-1, together with AR and FoxA1, could promote prostate cancer cell survival through activation of RAB3B expression. Collectively, our study has provided important insights into the hierarchical transcriptional regulatory network established between AR and NKX3-1 and sought to elucidate the important genetic-molecular-phenotypic paradigm in androgen-dependent prostate cancer. Genome-wide binding analyses of AR, NKX3-1 and FoxA1 in LNCaP with or without DHT (5alpha-dihydrotestosterone) stimulation using ChIP-Seq.
Project description:Androgen receptor (AR) orchestrates an intricate transcriptional regulatory network that governs prostate cancer initiation, development and progression. To understand this network in detail, we generated genome-wide maps of AR occupancy by ChIP-seq in LNCaP cells. We found NKX3-1, an androgen-dependent homeobox protein well-characterized for its role in prostate development and differentiation, being recruited to AR binding sites (ARBS) in response to androgen signaling. We identified 6,359 NKX3-1 binding sites, most of which overlapped with AR. In addition to its novel collaborative transcriptional role at well-known prostate cancer model genes, our binding and knockdown studies further suggested that NKX3-1 potentially regulates AR in a feed-forward manner. Integrative analysis of Oncomine molecular concepts showed that these androgen-regulated AR and NKX3-1 associated genes are significantly overexpressed in prostate carcinoma as well as advanced and recurrent prostate tumors. From our transcriptomic profiling and Gene Ontology analysis, we observed that AR and NKX3-1 co-regulate genes involved in ‘protein trafficking’ processes, which are mandatory events in the integration of oncogenic signaling pathways leading to prostate cancer development and progression. Interestingly, we found that AR and NKX3-1 co-regulate several members of the RAB GTPase family of secretory/trafficking proteins via the involvement of FoxA1 in a ternary complex and we believe that these AR/NKX3-1/FoxA1 co-regulated RAB genes could serve as expression signatures in prostate carcinogenesis. More specifically, through functional analyses, we showed that NKX3-1, together with AR and FoxA1, could promote prostate cancer cell survival through activation of RAB3B expression. Collectively, our study has provided important insights into the hierarchical transcriptional regulatory network established between AR and NKX3-1 and sought to elucidate the important genetic-molecular-phenotypic paradigm in androgen-dependent prostate cancer. Gene expression profiling of LNCaP in response to ETOH (vehicle) or DHT (5α-dihydrotestosterone) stimulation across different treatment time-points using microarray.
Project description:In castration-resistant prostate cancer (CRPC), clinical response to androgen receptor (AR) antagonists is limited mainly due to AR-variants expression and restored AR signaling. The metabolite spermine is most abundant in prostate and it decreases as prostate cancer progresses, but its functions remain poorly understood. Here, we show spermine inhibits full-length androgen receptor (AR-FL) and androgen receptor splice variant 7 (AR-V7) signaling and suppresses CRPC cell proliferation by directly binding and inhibiting protein arginine methyltransferase PRMT1. Spermine reduces H4R3me2a modification at the AR locus and suppresses AR binding as well as H3K27ac modification levels at AR target genes. Spermine supplementation restrains CRPC growth in vivo. PRMT1 inhibition also suppresses AR-FL and AR-V7 signaling and reduces CRPC growth. Collectively, we demonstrate spermine as an anticancer metabolite by inhibiting PRMT1 to transcriptionally inhibit AR-FL and AR-V7 signaling in CRPC, and we indicate spermine and PRMT1 inhibition as powerful strategies overcoming limitations of current AR-based therapies in CRPC.