The long non-coding RNAGHSROSreprograms prostate cancer cell lines toward a more aggressive phenotype

Abstract

42It is now appreciated that long non-coding RNAs (lncRNAs) are important players in the 43 orchestration of cancer progression. In this study we characterized GHSROS, a human 44 lncRNA gene on the opposite DNA strand (antisense) to the ghrelin receptor gene, in prostate 45 cancer. The lncRNA was upregulated by prostate tumors from different clinical datasets. 46Consistently, transcriptome data revealed that GHSROS alters the expression of cancer-47 associated genes. Functional analyses in vitro showed that GHSROS mediates tumor growth, 48 migration, and survival and resistance to the cytotoxic drug docetaxel. Increased cellular 49 proliferation of GHSROS-overexpressing PC3, DU145, and LNCaP prostate cancer cell lines 50 in vitro was recapitulated in a subcutaneous xenograft model. Conversely, in vitro antisense 51 oligonucleotide inhibition of the lncRNA reciprocally regulated cell growth and migration, 52 and gene expression. Notably, GHSROS modulates the expression of PPP2R2C, the loss of 53 which may drive androgen receptor pathway-independent prostate tumor progression in a 54 subset of prostate cancers. Collectively, our findings suggest that GHSROS can reprogram 55 prostate cancer cells toward a more aggressive phenotype and that this lncRNA may 56 represent a potential therapeutic target. 57 58 Keywords: long non-coding RNA, lncRNA, prostate cancer 59 60 5 reprograms prostate cancer cells toward a more aggressive phenotype, possibly by repressing 86 the expression of the tumor suppressor PPP2R2C to allow androgen-independent growth. 87 88 RESULTS 89GHSROS is expressed in prostate cancer 90Microarrays and RNA-sequencing are commonly used to assess the expression of genes. 91LncRNAs are often expressed at orders of magnitude lower than protein-coding transcripts, 92 however, making them difficult to detect 5, 11,12,13,14,15 . Interrogation of exon arrays harboring 93 four different strand-specific probes against GHSROS demonstrated that the lncRNA is 94 actively transcribed, although expressed at very low levels in cancer cell lines and tissues 95 ( Supplementary Fig. S1), consistent with previous observations from Northern blotting and 96 RT-PCR experiments 3 . The low expression across the GHSROS and GHSR loci in RNA-seq 97 datasets is illustrated in Supplementary Fig. S2. Collectively, these data demonstrate that it is 98 not currently possible to detect GHSROS in public genome-wide gene expression datasets. 99 100We next evaluated GHSROS expression in a qRT-PCR tissue array of 18 cancers. This 101 analysis revealed particularly high GHSROS expression in lung tumors, as previously 102 reported 3 , and elevated expression in prostate tumors ( Fig. 1b). Analysis of additional 103 prostate tissue-derived cDNA arrays revealed that GHSROS could be detected in 104 approximately 41.7% of all normal prostate tissues (n=24), 55.7% of tumors (n=88), and 105 58.1% of other prostatic diseases (e.g. prostatitis; n=31) ( Supplementary Table S1). GHSROS 106 was highly expressed by a subset of prostate tumors (~11.4%; ...