A comprehensive evaluation of the effects of RNA-editing proteins ADAR and ADARB1 on the expression of the drug-metabolizing enzymes in HepaRG cells.

Two RNA editing proteins, the adenosine deaminase acting on RNA, ADAR, and ADARB1, broadly regulate gene expression in editing-dependent and editing-independent manners. Previous studies showed that the expression of the drug-metabolizing cytochrome P450s (CYPs) and UDP glucuronosyltransferases (UGTs) changes upon knock-down (KD) of ADAR or ADARB1 in different hepatic cell lines. To systematically survey the effects of these two ADARs on the expression of CYPs and UGTs, we used siRNA in HepaRG cells and tested the association between the expression of the CYPs and ADARs in a liver sample cohort (n=246). KD of ADAR increased the expression of the CYP3As and CYP2C9 and reduced the expression of the others, while KD of ADARB1 reduced the expression of nearly all genes tested. ADAR KD also suppressed the induction of most CYPs, while ADARB1 KD had mixed effects depending on the inducer/gene combination. CYP expression was positively associated with both ADARs in liver samples, consistent with the KD results. However, after adjusting for the expression of transcription factors (TFs) known to regulate CYP expression, the associations disappeared, indicating that the effects of ADAR or ADARB1 primarily occur through TFs. Moreover, we found that the expression of normally-spliced CYP3A5 transcripts is increased in both KDs, indicating a direct effect of the ADARs on promoting the usage of the cryptic splice site generated by CYP3A5*3. Taken together, our results revealed the non-overlapping regulatory effects of ADAR and ADARB1 and supported their broad roles in controlling the expression of drug-metabolizing enzymes in the liver. Significance Statement Here we systematically surveyed the roles of ADAR and ADARB1 in both basal and induced expression of drug-metabolizing enzymes and assessed their co-expression in liver samples. Our results support that ADAR and ADARB1 regulate the expression of the drug-metabolizing enzymes in the liver, suggesting that factors affecting ADAR expression also have the potential to impact drug metabolism.