Networks of intergenic long-range enhancers and snpRNAs drive castration-resistant phenotype of prostate cancer and contribute to pathogenesis of multiple common human disorders

Abstract

Mechanistic relevance of intergenic disease-associated genetic loci (IDAGL) containing highly statistically significant disease-linked SNPs remains unknown. Here we present the experimental and clinical evidence supporting important role of IDAGL in human diseases. Targeted RT -PCR screen coupled with sequencing of purified PCR products detects widespread transcription at multiple IDAGL and identifies 96 small non-coding trans-regulatory RNAs of ∼100-300 nt in length containing SNPs (snpRNAs) associated with 21 common disorders. Multiple independent lines of experimental evidence support functionality of snpRNAs by documenting their cell-type-specific expression and evolutionary conservation of sequences, genomic coordinates and biological effects. Chromatin state signatures, expression profiling experiments and luciferase reporter assays demonstrate that many IDAGL are Polycomb-regulated long-range enhancers. Expression of snpRNAs in human and mouse cells markedly affects cellular behavior and induces allelespecific clinically-relevant phenotypic changes: NLRP1-locus snpRNAs rs2670660 exert regulatory effects on monocyte/ macrophage trans-differentiation, induce prostate cancer (PC) susceptibility snpRNAs and transform low-malignancy hormone-dependent human PC cells into highly malignant androgen-independent PC. Q-PCR analysis and luciferase reporter assays demonstrate that snpRNA sequences represent allele-specific "decoy" targets of microRNAs which function as SNP-allele-specific modifiers of microRNA expression and activity. We demonstrate that trans-acting RNA molecules facilitating resistance to androgen depletion (RAD) in vitro and castration-resistant phenotype (CRP) in vivo of PC contain intergenic 8q24-locus SNP variants (rs1447295; rs16901979; rs6983267) which were recently linked with increased risk of PC. Q-PCR analysis of clinical samples reveals markedly increased and highly concordant (r = 0.896; p < 0.0001) snpRNA expression levels in tumor tissues compared with the adjacent normal prostate [122-fold and 45- fold in Gleason 7 tumors (p = 0.03); 370-fold and 127-fold in Gleason 8 tumors (p = 0.0001); for NLRP1-locus and 8q24-locus snpRNAs, respectively]. Our experiments indicate that RAD and CR phenotype of human PC cells can be triggered by ncRNA molecules transcribed from the NLRP1-locus intergenic enhancer at 17p13 and down-stream activation of the 8q24-locus snpRNAs. Our results define the IDAGL at 17p13 and 8q24 as candidate regulatory loci of RAD and CR phenotypes of PC, reveal previously unknown molecular links between the innate immunity/inflammasome system and development of hormone-independent PC and identify novel molecular and genetic targets with diagnostic and therapeutic potentials exploration of which should be highly beneficial for personalized clinical management of PC. © 2011 Landes Bioscience.