m6A modification impacts hepatic drug and lipid metabolism properties by regulating carboxylesterase 2

Methylation of adenosine at the N-6 position to form N-6-methyladenosine (m(6)A) is the most prevalent epitran-scriptomic modification of mammalian mRNA. This modification is catalyzed by a methyltransferase-like 3 (METTL3)-METTL14 complex and is erased by demethylases such as fat mass and obesity-associated protein (FTO) or AlkB homolog 5 (ALKBH5). m(6)A modification regulates mRNA stability, nuclear export, splicing, and/or protein translation via recognition by reader proteins such as members of YT521-B homology (YTH) family. Carboxylesterase 2 (CES2) is a serine esterase responsible for the hydrolysis of drugs and endogenous substrates, such as triglycerides and diacylglycerides. Here, we examined the potential regulation of human CES2 expression by m(6)A modification. CES2 mRNA level was significantly increased by double knockdown of METTL3 and METTL14 but was decreased by knockdown of FTO or ALKBH5 in HepaRG and HepG2 cells, leading to changes in its protein level and hydrolase activity for 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11), suggesting that m(6)A modification negatively regulates CES2 expression. Consistent with the changes in CES2 expression, lipid accumulation in the cells was decreased by double knockdown of METTL3 and METTL14 but was increased by knockdown of FTO or ALKBH5. RNA immunoprecipitation assays using an anti-m(6)A antibody showed that adenosines in the 5'-untranslated region (UTR) and the last exon of CES2 are methylated. Luciferase assays revealed that YTHDC2, which degrades m(6)A-containing mRNA, downregulates CES2 expression by recognition of m(6)A in the 5'-UTR of CES2. Collectively, we demonstrated that m(6)A modification has a great impact on the regulation of CES2, affecting pharmacokinetics, drug response and lipid metabolism.

Automated summary

The study reveals that m(6)A modification negatively regulates CES2 expression by impacting its protein level, hydrolase activity, and lipid accumulation, thereby affecting pharmacokinetics and lipid metabolism. The regulation of CES2 by m(6)A modification highlights the potential significance of epitranscriptomic modifications in drug response and metabolic processes.