The landscape of RNA polymerase II–associated chromatin interactions in prostate cancer
Susmita G. Ramanand, Yong Chen, Jiapei Yuan, Kelly Daescu, Maryou B.K. Lambros, Kathleen E. Houlahan, Suzanne Carreira, Wei Yuan, GuemHee Baek, Adam Sharp, Alec Paschalis, Mohammed Kanchwala, Yunpeng Gao, Adam Aslam, Nida Safdar, Xiaowei Zhan, Ganesh V. Raj, Chao Xing, Paul C. Boutros, Johann de Bono, Michael Q. Zhang, Ram S. Mani
Susmita G. Ramanand, Yong Chen, Jiapei Yuan, Kelly Daescu, Maryou B.K. Lambros, Kathleen E. Houlahan, Suzanne Carreira, Wei Yuan, GuemHee Baek, Adam Sharp, Alec Paschalis, Mohammed Kanchwala, Yunpeng Gao, Adam Aslam, Nida Safdar, Xiaowei Zhan, Ganesh V. Raj, Chao Xing, Paul C. Boutros, Johann de Bono, Michael Q. Zhang, Ram S. Mani
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Research Article Genetics Oncology

The landscape of RNA polymerase II–associated chromatin interactions in prostate cancer

Abstract

Transcriptional dysregulation is a hallmark of prostate cancer (PCa). We mapped the RNA polymerase II–associated (RNA Pol II–associated) chromatin interactions in normal prostate cells and PCa cells. We discovered thousands of enhancer-promoter, enhancer-enhancer, as well as promoter-promoter chromatin interactions. These transcriptional hubs operate within the framework set by structural proteins — CTCF and cohesins — and are regulated by the cooperative action of master transcription factors, such as the androgen receptor (AR) and FOXA1. By combining analyses from metastatic castration-resistant PCa (mCRPC) specimens, we show that AR locus amplification contributes to the transcriptional upregulation of the AR gene by increasing the total number of chromatin interaction modules comprising the AR gene and its distal enhancer. We deconvoluted the transcription control modules of several PCa genes, notably the biomarker KLK3, lineage-restricted genes (KRT8, KRT18, HOXB13, FOXA1, ZBTB16), the drug target EZH2, and the oncogene MYC. By integrating clinical PCa data, we defined a germline-somatic interplay between the PCa risk allele rs684232 and the somatically acquired TMPRSS2-ERG gene fusion in the transcriptional regulation of multiple target genes — VPS53, FAM57A, and GEMIN4. Our studies implicate changes in genome organization as a critical determinant of aberrant transcriptional regulation in PCa.

Authors

Susmita G. Ramanand, Yong Chen, Jiapei Yuan, Kelly Daescu, Maryou B.K. Lambros, Kathleen E. Houlahan, Suzanne Carreira, Wei Yuan, GuemHee Baek, Adam Sharp, Alec Paschalis, Mohammed Kanchwala, Yunpeng Gao, Adam Aslam, Nida Safdar, Xiaowei Zhan, Ganesh V. Raj, Chao Xing, Paul C. Boutros, Johann de Bono, Michael Q. Zhang, Ram S. Mani

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Figure 8

Transcriptional regulation by the PCa risk SNP rs684232.

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Transcriptional regulation by the PCa risk SNP rs684232. (A) Integrated ...
(A) Integrated genome view of RNA Pol II–associated chromatin interactions in the genomic region harboring the PCa risk SNP rs684232. (B) rs684232 was significantly associated with mRNA abundance of FAM57A, VPS53, and GEMIN4 in the CPC-GENE, TCGA, and Porto cohorts. Box plots represent the median and the 0.25 and 0.75 quantiles, with whiskers at 1.5 times the IQR. mRNA abundance was measured in FPKM. The numbers below the genotypes indicate the number of samples in each group. P values and effect size are from a linear model. (C) Epigenetic features of the PCa risk SNP rs684232 locus in VCaP cells are described using ChIP-Seq analysis. (D) rs684232 falls in an active enhancer region, and the alternative allele was found to be significantly associated with decreased H3K27ac binding. Heatmap shows H3K27ac ChIP-Seq signal within chr17:614900-622900 (x axis) for 92 patients (y axis). Color indicates ChIP-Seq signal intensity, and the black bar in the covariate along the top indicates the location of rs684232. Box plot shows H3K27ac signal intensity stratified by genotype in the Porto cohort (Mann-Whitney U test for the recessive model). The y axis represents the number of H3K27ac ChIP-Seq read counts mapped to the SNP rs684232 region, which were normalized by the trimmed mean of M values (TMM). (E) Box plots show H3K27ac signal intensity in the promoter regions of FAM57A and GEMIN4 stratified by genotype in the Porto cohort (Mann-Whitney U test for the recessive model). P values and effect size are from a linear model. (F) Sequence analysis to confirm the cloning of the WT and risk (rs684232) alleles in the pGL2 luciferase reporter plasmid. (G) Luciferase reporter assays in LNCaP and VCaP cells. Cells were cotransfected with pSV-Renilla and the luciferase reporter encoding the WT or risk (rs684232) allele and processed 48 hours after transfection. Firefly Luc/Renilla Luc activity was determined (mean ± SD, n = 6; ****P < 0.0001, by 2-tailed Student’s t test).