Recherche des partenaires moléculaires d’un long ARN non codant, régulé par l’hypoxie, dans les adénocarcinomes pulmonaires

Lung cancer is the leading cause of cancer death worldwide, with poor prognosis and a high rate of recurrence despite early surgical removal. It is therefore essential to identify new prognostic markers and new therapeutic targets. My team is interested in gene regulation related to hypoxia, a factor associated with relapse of lung adenocarcinomas (LUAD), the most represented form of lung cancer. While long non-coding RNAs (lncRNA) are increasingly recognized as major gene expression regulators through various mechanisms involving nucleic acid and protein partners, their roles in cancer development and hypoxic response are still largely unexplored. We previously identified a "hypoxaLinc", LincH3 that is significantly up-regulated both in LUAD tumor tissues and cell lines in response to hypoxia. This 10 kb nuclear transcript is regulated by NFB and may participate in the regulation of hypoxic or inflammatory responses.
My project focused on the characterization of LincH3 mode of action through the identification of its DNA-binding sites. My goal was to set up a novel technique, the ChIRP (Chromatin Isolation by RNA Purification), based on affinity capture of LncRNA: chromatin complexes by tiling antisense-oligos and mapping of its genomic binding sites at high resolution. Using WT and CRISPR/Cas9 LincH3-deleted LUAD cells, I performed ChIRP-Seq to identify 254 LincH3 binding sites on chromatin. Through transcriptome analysis I showed dysregulation of TGF, NRF2 and NFB signaling pathways in LincH3-deficient LUAD cells. Finally, the comparative analysis of these data identified 11 potential target genes of LincH3 including CTGF, a central regulator of TGFβ and angiogenic pathways.