RNA editing mediates the functional switch of COPA in a novel mechanism of hepatocarcinogenesis

Abstract

Background & aims: RNA editing introduces nucleotide changes in RNA sequences. Recent studies have reported that aberrant adenosine-to-inosine RNA editing is implicated in cancers. Until now, very few functionally important protein-recoding editing targets have been discovered. Here, we investigated the role of a recently discovered protein-recoding editing target COPA (coatomer subunit α) in hepatocellular carcinoma (HCC).

Methods: Clinical implication of COPA editing was studied in a cohort of 125 HCC patients. CRISPR/Cas9-mediated knockout of the editing site complementary sequence (ECS) was used to delete edited COPA transcripts endogenously. COPA editing-mediated change in its transcript or protein stability was investigated upon actinomycin D or cycloheximide treatment, respectively. Functional difference in tumourigenesis between wild-type and edited COPA (COPA^WTvs. COPA^I164V) and the exact mechanisms were also studied in cell models and mice.

Results: ADAR2 binds to double-stranded RNA formed between edited exon 6 and the ECS at intron 6 of COPA pre-mRNA, causing an isoleucine-to-valine substitution at residue 164. Reduced editing of COPA is implicated in the pathogenesis of HCC, and more importantly, it may be involved in many cancer types. Upon editing, COPA^WT switches from a tumour-promoting gene to a tumour suppressor that has a dominant-negative effect. Moreover, COPA^I164V may undergo protein conformational change and therefore become less stable than COPA^WT. Mechanistically, COPA^I164V may deactivate the PI3K/AKT/mTOR pathway through downregulation of caveolin-1 (CAV1).

Conclusions: We uncover an RNA editing-associated mechanism of hepatocarcinogenesis by which downregulation of ADAR2 caused the loss of tumour suppressive COPA^I164V and concurrent accumulation of tumour-promoting COPA^WT in tumours; a rapid degradation of COPA^I164V protein and hyper-activation of the PI3K/AKT/mTOR pathway further promote tumourigenesis.

Lay summary: RNA editing is a process in which RNA is changed after it is made from DNA, resulting in an altered gene product. In this study, we found that RNA editing of a gene known as coatomer subunit α (COPA) is lower in tumour samples and discovered that this editing process changes COPA protein from a tumour-promoting form to a tumour-suppressive form. Loss of the edited COPA promotes the development of liver cancer.

Keywords: A-to-I; ADARs; COPA; HCC; Protein recoding; RNA editing.