Endoplasmic reticulum–associated degradation is required for nephrin maturation and kidney glomerular filtration function
Sei Yoshida, Xiaoqiong Wei, Gensheng Zhang, Christopher L. O’Connor, Mauricio Torres, Zhangsen Zhou, Liangguang Lin, Rajasree Menon, Xiaoxi Xu, Wenyue Zheng, Yi Xiong, Edgar Otto, Chih-Hang Anthony Tang, Rui Hua, Rakesh Verma, Hiroyuki Mori, Yang Zhang, Chih-Chi Andrew Hu, Ming Liu, Puneet Garg, Jeffrey B. Hodgin, Shengyi Sun, Markus Bitzer, Ling Qi
Sei Yoshida, Xiaoqiong Wei, Gensheng Zhang, Christopher L. O’Connor, Mauricio Torres, Zhangsen Zhou, Liangguang Lin, Rajasree Menon, Xiaoxi Xu, Wenyue Zheng, Yi Xiong, Edgar Otto, Chih-Hang Anthony Tang, Rui Hua, Rakesh Verma, Hiroyuki Mori, Yang Zhang, Chih-Chi Andrew Hu, Ming Liu, Puneet Garg, Jeffrey B. Hodgin, Shengyi Sun, Markus Bitzer, Ling Qi
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Research Article Cell biology Nephrology

Endoplasmic reticulum–associated degradation is required for nephrin maturation and kidney glomerular filtration function

Abstract

Podocytes are key to the glomerular filtration barrier by forming a slit diaphragm between interdigitating foot processes; however, the molecular details and functional importance of protein folding and degradation in the ER remain unknown. Here, we show that the SEL1L-HRD1 protein complex of ER-associated degradation (ERAD) is required for slit diaphragm formation and glomerular filtration function. SEL1L-HRD1 ERAD is highly expressed in podocytes of both mouse and human kidneys. Mice with podocyte-specific Sel1L deficiency develop podocytopathy and severe congenital nephrotic syndrome with an impaired slit diaphragm shortly after weaning and die prematurely, with a median lifespan of approximately 3 months. We show mechanistically that nephrin, a type 1 membrane protein causally linked to congenital nephrotic syndrome, is an endogenous ERAD substrate. ERAD deficiency attenuated the maturation of nascent nephrin, leading to its retention in the ER. We also show that various autosomal-recessive nephrin disease mutants were highly unstable and broken down by SEL1L-HRD1 ERAD, which attenuated the pathogenicity of the mutants toward the WT allele. This study uncovers a critical role of SEL1L-HRD1 ERAD in glomerular filtration barrier function and provides insights into the pathogenesis associated with autosomal-recessive disease mutants.

Authors

Sei Yoshida, Xiaoqiong Wei, Gensheng Zhang, Christopher L. O’Connor, Mauricio Torres, Zhangsen Zhou, Liangguang Lin, Rajasree Menon, Xiaoxi Xu, Wenyue Zheng, Yi Xiong, Edgar Otto, Chih-Hang Anthony Tang, Rui Hua, Rakesh Verma, Hiroyuki Mori, Yang Zhang, Chih-Chi Andrew Hu, Ming Liu, Puneet Garg, Jeffrey B. Hodgin, Shengyi Sun, Markus Bitzer, Ling Qi

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Figure 7

Nephrin disease mutants are unstable, retained in the ER, and targeted for proteasomal degradation by ERAD.

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Nephrin disease mutants are unstable, retained in the ER, and targeted f...
(A) Structural modeling of human nephrin showing the domains (Ig and FN) and location of 6 pathogenic mutations. EvoEF2 (ΔΔG) for each mutant is indicated in blue. (B and C) The predicted local structures of WT, I171N (B), and G270C (C). WT and mutated residues are shown in green and magenta, respectively. Distances between the indicated residues are shown. Arrows indicate disulfide bonds. (D and E) Western blot analysis of WT and mutant nephrin proteins in transfected HEK293T cells, showing nephrin mutants running as the b form on SDS-PAGE. Quantitation is shown in E. (F) Western blot analysis following EndoH digestion in HEK293T cells transfected with WT or mutant nephrin. (G and H) Western blot analysis of WT and mutant nephrin in transfected HEK293T cells treated with CHX for 4 hours, with quantitation shown in H. Values represent the mean ± SEM. Data are representative of at least 2 independent experiments. *P < 0.05, by 2-tailed Student’s t test.