Post-transcriptional control of <i>KRAS</i> : functional roles of 5′UTR RNA G-quadruplexes, long noncoding RNA, and hnRNPA1

Previous studies have shown that human KRAS expression is regulated at the transcriptional level by G-quadruplex DNA structures within its promoter. Here we show an additional level of regulation involving a post-transcriptional mechanism centred on the 5'-untranslated region (5'UTR) of the messenger RNA (mRNA) characterized by G4 structures (rG4s). Long noncoding RNAs (lncRNAs) and the protein hnRNPA1 are also involved in this mechanism. RIP-seq confirmed the presence of rG4s in the 5'UTR. Deletion of the rG4 region using CRISPR/Cas9 resulted in a significant increase in KRAS mRNA levels, indicating the role of the 5'UTR in controlling mRNA levels. RIP shows that hnRNPA1 is recruited to the 5'UTR, where it unfolds the rG4 structures and potentially affects mRNA stability. In addition, lncRNAs transcribed from the LINC01750 locus can hybridize to the rG4 region of 5'UTR and form RNA duplexes leading to RNase III-assisted degradation of the targeted mRNA. Activation of the LINC01750 locus with dCas9-VP64 resulted in downregulation of KRAS mRNA, whereas its suppression with dCas9-KRAB led to upregulation of both KRAS mRNA and protein. Since lncRNA-mediated regulation of mRNA appears to be a crucial aspect of cellular homeostasis and its disruption contributes to various diseases, understanding these mechanisms may reveal promising new therapeutic targets.