The Transcription Factor p8 Regulates Autophagy in Response to Palmitic Acid Stress via a Mammalian Target of Rapamycin (mTOR)-independent Signaling Pathway

Sheng-Nan Jia,Cheng Lin,Dian-Fu Chen,An-Qi Li,Li Dai,Li Zhang,Ling-Ling Zhao,Jin-Shu Yang,Fan Yang,Wei-Jun Yang

doi:10.1074/jbc.m115.675793

Abstract

Autophagy is an evolutionarily conserved degradative process that allows cells to maintain homoeostasis in numerous physiological situations. This process also functions as an essential protective response to endoplasmic reticulum (ER) stress, which promotes the removal and degradation of unfolded proteins. However, little is known regarding the mechanism by which autophagy is initiated and regulated in response to ER stress. In this study, different types of autophagy were identified in human gastric cancer MKN45 cells in response to the stress induced by nutrient starvation or lipotoxicity in which the regulation of these pathways is mammalian target of rapamycin (mTOR)-dependent or -independent, respectively. Interestingly, we found that p8, a stress-inducible transcription factor, was enhanced in MKN45 cells treated with palmitic acid to induce lipotoxicity. Furthermore, an increase in autophagy was observed in MKN45 cells stably overexpressing p8 using a lentivirus system, and autophagy induced by palmitic acid was blocked by p8 RNAi compared with the control. Western blotting analyses showed that autophagy was regulated by p8 or mTOR in response to the protein kinase-like endoplasmic reticulum kinase/activating transcription factor 6-mediated ER stress of lipotoxicity or the parkin-mediated mitochondrial stress of nutrient starvation, respectively. Furthermore, our results indicated that autophagy induced by palmitic acid is mTOR-independent, but this autophagy pathway was regulated by p8 via p53- and PKCα-mediated signaling in MKN45 cells. Our findings provide insights into the role of p8 in regulating autophagy induced by the lipotoxic effects of excess fat accumulation in cells.

Highlights

Implicated in the physiopathology of major diseases such as cancer, neurodegenerative disorders, and diabetes (4 – 8)
The results of Western blotting and immunofluorescence showed that cells cultured in the Earle’s balanced salt solution (EBSS) had a higher amount of lipidated LC3B than control cells cultured in complete medium, indicating that autophagy was increased in response to nutrient starvation for 6 h (Fig. 1, A and C)
Palmitic acid treatment or overexpression of p8 increased the levels of p-protein kinase-like endoplasmic reticulum kinase (PERK) and activating transcription factor 6 (ATF6) but had no effect on the level of parkin (Fig. 5, B and C). These results suggested that autophagy induced by palmitic acid was an endoplasmic reticulum (ER)-stress response regulated by the p8 signaling pathway, whereas autophagy induced by nutrient starvation was a mitochondrial-stress response controlled by the AMPK/mammalian target of rapamycin (mTOR) signaling pathway

Summary

Introduction

Implicated in the physiopathology of major diseases such as cancer, neurodegenerative disorders, and diabetes (4 – 8). The results demonstrated that autophagy induced by nutrient starvation or lipotoxicity was controlled by the mTOR or p8 signaling pathway, respectively. The results of Western blotting and immunofluorescence showed that cells cultured in the EBSS had a higher amount of lipidated LC3B than control cells cultured in complete medium, indicating that autophagy was increased in response to nutrient starvation for 6 h (Fig. 1, A and C).

Results

Conclusion