Regulatory Effect of Methylation of the Porcine AQP3 Gene Promoter Region on Its Expression Level and Porcine Epidemic Diarrhea Virus Resistance

Abstract

As an important carrier for intestinal secretion and water absorption, aquaporin 3 (AQP3) is closely related to diarrhea. In this study, we investigated the mechanisms of AQP3 gene expression regulation in porcine epidemic diarrhea virus (PEDV)-induced diarrhea confirmed by PCR amplification and sequencing. Evaluation of intestinal pathology showed that diarrhea caused by PEDV infection destroyed the intestinal barrier of piglets. qPCR analysis showed that AQP3 expression in the small intestine of PEDV-infected piglets was extremely significantly decreased. qPCR and Bisulfite sequencing PCR revealed an increase in the methylation levels of both CpG islands in the AQP3 promoter region in the jejunum of PEDV-infected piglets. The methylation of mC-20 and mC-10 sites within the two CpG islands showed a significant negative correlation with AQP3 expression. Chromatin Co-Immunoprecipitation (ChIP)-PCR showed that the Sp1 transcription factor was bound to the AQP3 promoter region containing these two CpG sites. AQP3 expression was also extremely significantly reduced in Sp1-inhibited IPEC-J2 cells, indicating that abnormal methylation at the mC-20 site of CpG1 and the mC-10 site of CpG2 reduces its expression in PEDV-infected piglet jejunum by inhibiting the binding of Sp1 to the AQP3 promoter. These findings provide a theoretical basis for further functional studies of porcine AQP3.

Genome-Wide DNA Methylome and Transcriptome Analysis of Porcine Testicular Cells Infected With Transmissible Gastroenteritis Virus

“…Xiao et al (51) reported that, methylation of the mC-5 site in the MUC2 promoter inhibited the binding of Yin Yang 1 (YY1) to the promoter, down regulated the expression of MUC2 and increased the susceptibility of piglets to porcine epidemic diarrhea virus (PEDV). We also have confirmed abnormal methylation at the AQP3 promoter reduce its expression in PEDV-infected piglet jejunum (52). Zhang et al (53) suggested that porcine reproductive and respiratory syndrome virus (PRRSV) vaccination in sows induces DNA methylation changes in genes and DNA methylation changes occur through intergenerational transmission.…”

A Functional Variant in the Aquaporin-3 Promoter Modulates Its Expression and Correlates With Resistance to Porcine Epidemic Virus Infection in Porcine Intestinal Epithelial Cells

“…In addition, inhibition of AQP3 function in the colon results in diarrhea ( Ikarashi et al, 2012 ). Our previous work also showed that the expression of AQP3 was remarkably decreased in the intestinal segments of PEDV-induced piglets ( Wu et al, 2020 ), which indicates AQP3 was a potential player in modulating PEDV-induced diarrhea. However, the functions of AQP3 in cellular responses to PEDV infection and the regulatory mechanisms underlying the expression of AQP3 remain largely unknown.…”

Hypomethylated interferon regulatory factor 8 recruits activating protein-2α to attenuate porcine epidemic diarrhea virus infection in porcine jejunum

“…In this study, M gene amplification and intestinal morphology observations showed that PEDV bands could be amplified in piglets infected with PEDV, accompanied by symptoms, such as intestinal villous atrophy, epithelial cell necrosis, and shedding. Combined with the fact that no other typical diarrheal pathogens (TGEV and PoRV) were detected in these piglets ( 28 ), PEDV was determined to be the only pathogen of infection.…”