Jennifer Y. Tan

Recently, a handful of intergenic long noncoding RNAs (lncRNAs) have been shown to compete with mRNAs for binding to miRNAs and to contribute to development and disease. Beyond these reports, little is yet known of the extent and functional consequences of miRNA-mediated regulation of mRNA levels by lncRNAs. To gain further insight into lncRNA-mRNA miRNA-mediated crosstalk, we reanalyzed transcriptome-wide changes induced by the targeted knockdown of over 100 lncRNA transcripts in mouse embryonic stem cells (mESCs). We predicted that, on average, almost one-fifth of the transcript level changes induced by lncRNAs are dependent on miRNAs that are highly abundant in mESCs. We validated these findings experimentally by temporally profiling transcriptome-wide changes in gene expression following the loss of miRNA biogenesis in mESCs. Following the depletion of miRNAs, we found that >50% of lncRNAs and their miRNA-dependent mRNA targets were up-regulated coordinately, consistent with their interaction being miRNA-mediated. These lncRNAs are preferentially located in the cytoplasm, and the response elements for miRNAs they share with their targets have been preserved in mammals by purifying selection. Lastly, miRNA-dependent mRNA targets of each lncRNA tended to share common biological functions. Post-transcriptional miRNA-mediated crosstalk between lncRNAs and mRNA, in mESCs, is thus surprisingly prevalent, conserved in mammals, and likely to contribute to critical developmental processes.

The MITF and SOX10 transcription factors regulate the expression of genes important for melanoma proliferation, invasion and metastasis. Despite growing evidence of the contribution of long noncoding RNAs (lncRNAs) in cancer, including melanoma, their functions within MITF-SOX10 transcriptional programmes remain poorly investigated. Here we identify 245 candidate melanoma associated lncRNAs whose loci are co-occupied by MITF-SOX10 and that are enriched at active enhancer-like regions. Our work suggests that one of these, Disrupted In Renal Carcinoma 3 (DIRC3), may be a clinically important MITF-SOX10 regulated tumour suppressor. DIRC3 depletion in human melanoma cells leads to increased anchorage-independent growth, a hallmark of malignant transformation, whilst melanoma patients classified by low DIRC3 expression have decreased survival. DIRC3 is a nuclear lncRNA that activates expression of its neighbouring IGFBP5 tumour suppressor through modulating chromatin structure and suppressing SOX10 binding to putative regulatory elements within the DIRC3 locus. In turn, DIRC3 dependent regulation of IGFBP5 impacts the expression of genes involved in cancer associated processes and is needed for DIRC3 control of anchorage-independent growth. Our work indicates that lncRNA components of MITF-SOX10 networks are an important new class of melanoma regulators and candidate therapeutic targets that can act not only as downstream mediators of MITF-SOX10 function but as feedback regulators of MITF-SOX10 activity.

Intergenic long noncoding RNAs (lincRNAs) are the largest class of transcripts in the human genome. Although many have recently been linked to complex human traits, the underlying mechanisms for most of these transcripts remain undetermined. We investigated the regulatory roles of a high-confidence and reproducible set of 69 trait-relevant lincRNAs (TR-lincRNAs) in human lymphoblastoid cells whose biological relevance is supported by their evolutionary conservation during recent human history and genetic interactions with other trait-associated loci. Their enrichment in enhancer-like chromatin signatures, interactions with nearby trait-relevant protein-coding loci, and preferential location at topologically associated domain (TAD) boundaries provide evidence that TR-lincRNAs likely regulate proximal trait-relevant gene expression in cis by modulating local chromosomal architecture. This is consistent with the positive and significant correlation found between TR-lincRNA abundance and intra-TAD DNA-DNA contacts. Our results provide insights into the molecular mode of action by which TR-lincRNAs contribute to complex human traits.

BACKGROUND: Recent reports have highlighted instances of mRNAs that, in addition to coding for protein, regulate the abundance of related transcripts by altering microRNA availability. These two mRNA roles - one mediated by RNA and the other by protein - are inter-dependent and hence cannot easily be separated. Whether the RNA-mediated role of transcripts is important, per se, or whether it is a relatively innocuous consequence of competition by different transcripts for microRNA binding remains unknown. RESULTS: Here we took advantage of 48 loci that encoded proteins in the earliest eutherian ancestor, but whose protein-coding capability has since been lost specifically during rodent evolution. Sixty-five percent of such loci, which we term 'unitary pseudogenes', have retained their expression in mouse and their transcripts exhibit conserved tissue expression profiles. The maintenance of these unitary pseudogenes' spatial expression profiles is associated with conservation of their microRNA response elements and these appear to preserve the post-transcriptional roles of their protein-coding ancestor. We used mouse Pbcas4, an exemplar of these transcribed unitary pseudogenes, to experimentally test our genome-wide predictions. We demonstrate that the role of Pbcas4 as a competitive endogenous RNA has been conserved and has outlived its ancestral gene's loss of protein-coding potential. CONCLUSIONS: These results show that post-transcriptional regulation by bifunctional mRNAs can persist over long evolutionary time periods even after their protein coding ability has been lost.