Characterization and expression analysis of ATG4 paralogs in response to the palmitic acid induced-ER stress in Ctenopharyngodon idellus kidney cells.

Abstract

Autophagy is regulated by a variety of autophagy related genes (ATG), among them, ATG4 is a highly specific protease that is critical to autophagy. The purpose of this study was to investigate the role of the ATG4 response to Palmitic acid (PA) and their relationships with Endoplasmic reticulum (ER) stress in grass carp. In the present study, four ATG4 paralogs from grass carp were identified and analyzed. The open reading frames of ATG4A, ATG4B, ATG4C, and ATG4D were 1212, 1194, 1429, and 1482bp long, encoding 403, 394, 475, and 493 amino acids, respectively. Secondly, the mRNA levels of ATG4 paralogs transcripts in 13 tissues of grass carp were analyzed to determine the tissue distribution. The highest (p<0.05) expression of ATG4A, ATG4B, ATG4C, and ATG4D were found in the heart, white muscle, brain, and liver, respectively. Moreover, after stimulation with PA, the expression of the four ATG4 paralogs in Ctenopharyngodon idellus kidney (CIK) cells were significantly up-regulated (p<0.05). Inhibiting ER stress with 4 phenylbutyrate (4-PBA) significantly reduced PA-induced autophagy in CIK cells, while the expression of PA-induced ATG4 paralogs were significantly decreased (p<0.05), suggesting that ATG4 is involved in PA-induced autophagy. We also demonstrated that PA could induced inflammation-related genes in CIK cells, and the PA-induced inflammation were alleviated by 4-PBA. Moreover, overexpression ATG4C induced ER stress and decreased inflammation. Taken together, these results demonstrate for the first time that inhibition of ER stress could reduce PA-induced expression of ATG4 paralogs and some inflammation-related genes.