CRNDE - A Long Non-Coding RNA Activated in Colorectal Cancer

The CRNDE lncRNA was recently identified as being over expressed in >90% colorectal cancers (CRC) and adenomas suggesting CRNDE is either a key player in or is activated in response to early colorectal oncogenic events. CRNDE undergoes alternate splicing with at least 10 different isoforms identified from cDNA cloning. It is proposed to interact with chromatin modifying complexes PRC2 and CoREST and therefore may play a key role in epigenetic remodeling in cancer. The overall aim of my thesis was to define the transcriptional complexity of CRNDE and its role in potential epigenetic deregulation in CRC. First, I characterised the expression and structure of CRNDE transcripts in CRC using quantitative PCR (qPCR) and NimbleGen custom tiling arrays and confirmed the expected 5’ and 3’ end of transcripts, dominance of the 5 major exons and notably presence of transcripts retaining intronic regions, particularly a conserved region in intron 4 (gVC-In4). Intron retaining transcripts localised to the nucleus while fully spliced transcripts localised to the cytoplasm. Second, I investigated chromatin structure across CRNDE and flanking regions and found no evidence for alternate start sites within CRNDE or an indepenant role of gVC-In4 as an enhancer. The expected transcription start site (TSS) of CRNDE was clearly marked with the active promoter mark H3K4me3 and H3K27ac, however there was no evidence for concordant enrichment of H3K4me3 or H3K27ac on genes in an extended region around CRNDE, indicating that CRNDE is not part of a long range activated region in CRC. Finally ,I conducted preliminary experiments using siRNA targeted to gVC-In4 of CRNDE in HCT116 to address the role of the unspliced nuclear transcript in chromatin remodelling. I found local changes in promoter associated H3K4me3 within CRNDE and neighbouring gene IRX5 and at distant FTO and metallothioneins genes. Therefore I propose CRNDE may be acting in cis as well as in trans through deregulation of H3K4me3 (possibly via CoREST) to elicit downstream chromatin remodeling effects. This preliminary data needs further extension and validation, however, supports the hypothesis that the lncRNA CRNDE may act to mediate epigenetic changes in CRC