FTO Regulates Apoptosis in CPB2-Treated IPEC-J2 Cells by Targeting Caspase 3 Apoptotic Protein

SIMPLE SUMMARY: Fat mass and obesity associated protein (FTO) is a key demethylase in the process of bacterial diarrhea in piglets. However, the involvement of FTO in infectious diarrhea caused by Clostridium perfringens type C is not known. This study demonstrated that FTO plays an indispensable role in this type of diarrhea; and that the absence of FTO promotes apoptosis and inflammation in the intestinal porcine epithelial cell line-J2 (IPEC-J2). FTO targets Caspase 3 to affect the apoptosis of IPEC-J2 cells. ABSTRACT: N6-methyladenosine (m6A) modification can accommodate mRNA processing, stability, and translation in mammals, and fat mass and obesity associated protein (FTO) is a vital demethylase in the m6A modification pathway. Clostridium perfringens type C (C. perfringens type C) causes diarrhea in piglets and has a serious impact on the pig industry. However, our understanding of the effect of m6A in the process of C. perfringens type C infectious piglet diarrhea (CPTCIPD) is limited. Here, an in vitro model of CPTCIPD was constructed by treating the intestinal porcine epithelial cell line-J2 (IPEC-J2) with Clostridium perfringens beta2 (CPB2) toxin, and the role of FTO was analyzed using quantitative real-time polymerase chain reaction, Western blotting, and flow cytometry. The results revealed that the overall RNA m6A contents at the tissue and cell levels were significantly up-regulated after C. perfringens infection (p < 0.05). FTO expression was significantly reduced in CPB2-treated IPEC-J2 cells. Functionally, FTO knockdown in the treated cells inhibited their proliferation and promoted apoptosis and the inflammation phenotype, whereas FTO overexpression had the opposite effects. Inhibiting FTO prolonged the half-life and up-regulated the expression of Caspase 3, leading to apoptosis. Therefore, this work explored the regulation of FTO in IPEC-J2 cells after CPB2 treatment and enhanced our understanding of the effect of the m6A modification in CPTCIPD.