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ATF6 pathway of unfolded protein response mediates advanced oxidation protein product-induced hypertrophy and epithelial-to-mesenchymal transition in HK-2 cells

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Abstract

Advanced oxidation protein products (AOPPs) accelerate the progression of chronic kidney disease. We previously demonstrated that AOPPs induce hypertrophy and epithelial-to-mesenchymal transition (EMT) in human proximal tubular cells (HK-2 cells) through induction of endoplasmic reticulum (ER) stress. However, which pathway of unfolded protein response (UPR) induced by ER stress plays crucial roles in this process remains unclear. In this study, we investigated the roles of the protein kinase RNA-like ER kinase (PERK), activating transcription factor 6 (ATF6), and inositol-requiring enzyme 1 (IRE1) pathways of UPR in this process in HK-2 cells. AOPP treatment induced the overexpression of cleaved ATF6 and spliced form of X-box binding protein-1, and induced the phosphorylation of PERK, eukaryotic translation initiation factor 2α and IRE1. Furthermore, silencing of ATF6 increased E-cadherin and zonula occludens-1 expression, lowered the expression of vimentin, and downregulated total protein content, whereas knockdown of PERK or IRE1 resulted in no difference compared with the scramble siRNA-transfected cells. AOPP-induced phosphorylation of Src, which was reproduced by thapsigargin, an inducer of ER stress, was partly reversed by salubrinal, an inhibitor of ER stress. Furthermore, the Src inhibitor saracatinib effectively blocked AOPP-induced phosphorylation of Src, activation of ER stress, hypertrophy, and EMT in HK-2 cells. Collectively, our results indicate that AOPPs induce the PERK, ATF6, and IRE1 pathways of UPR, and the ATF6 pathway rather than the other two pathways mediates AOPP-induced HK-2-cell hypertrophy and EMT. We also suggest that the ER stress involved in this process is likely mediated by the activation of Src kinase.

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Acknowledgments

This work was supported by the Natural Science Foundation of Guangdong Province (Grant Nos. 10151051501000030, S2011010004053, S2011040003566), the Science and Technical Plan of Guangzhou, Guangdong, China (Grant No. 11C22120703), the Science and Technical Plan of Guangdong, China (Grant No. 2013B021800149), and the National Natural Science Foundation of China (Grant No. 81202842).

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    Authors and Affiliations

    1. Department of Nephrology, Zhujiang Hospital, Southern Medical University, No. 253, Gongye Road, Haizhu District, Guangzhou, 510280, Guangdong, China

      Xun Tang, Xiujie Liang, Tingting Guo, Na Duan, Yue Wang, Lei Yang & Jun Zhang

    2. Southern Medical University, Guangzhou, Guangdong, China

      Minhui Li

    3. Department of Nephrology, The Third People’s Hospital, Nanhai District, Foshan, Guangdong, China

      Minhui Li

    4. Department of Nephrology, Shilong People’s Hosiptal, Southern Medical University, Dongguan, Guangdong, China

      Guang Rong

    5. College of Arts and Science, Syracuse University, Syracuse, USA

      Shaojie Zhang

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    Xun Tang and Xiujie Liang have contributed equally to this work.

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    Tang, X., Liang, X., Li, M. et al. ATF6 pathway of unfolded protein response mediates advanced oxidation protein product-induced hypertrophy and epithelial-to-mesenchymal transition in HK-2 cells. Mol Cell Biochem 407, 197–207 (2015). https://doi.org/10.1007/s11010-015-2469-0

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