Steroid-sensitive nephrotic syndrome candidate gene CLVS1 regulates podocyte oxidative stress and endocytosis.

Brandon M Lane,Guanghong Wu,Mohamed Shalaby,Alejandro Ochoa,Rasheed Gbadegesin,Megan Chryst-Stangl,Kinsie Huggins,Amika Sood,David N Howell,So Young Kim,Sherif El Desoky,Claire C Middleton,Sara E Miller,Andrew F Malone,Ricardo Vancini,Gentzon Hall,Jameela A Kari

doi:10.1172/jci.insight.152102

Abstract

We performed next-generation sequencing in patients with familial steroid-sensitive nephrotic syndrome (SSNS) and identified a homozygous segregating variant (p.H310Y) in the gene encoding clavesin-1 (CLVS1) in a consanguineous family with 3 affected individuals. Knockdown of the clavesin gene in zebrafish (clvs2) produced edema phenotypes due to disruption of podocyte structure and loss of glomerular filtration barrier integrity that could be rescued by WT CLVS1 but not the p.H310Y variant. Analysis of cultured human podocytes with CRISPR/Cas9-mediated CLVS1 knockout or homozygous H310Y knockin revealed deficits in clathrin-mediated endocytosis and increased susceptibility to apoptosis that could be rescued with corticosteroid treatment, mimicking the steroid responsiveness observed in patients with SSNS. The p.H310Y variant also disrupted binding of clavesin-1 to α-tocopherol transfer protein, resulting in increased reactive oxygen species (ROS) accumulation in CLVS1-deficient podocytes. Treatment of CLVS1-knockout or homozygous H310Y-knockin podocytes with pharmacological ROS inhibitors restored viability to control levels. Taken together, these data identify CLVS1 as a candidate gene for SSNS, provide insight into therapeutic effects of corticosteroids on podocyte cellular dynamics, and add to the growing evidence of the importance of endocytosis and oxidative stress regulation to podocyte function.

Highlights

Childhood nephrotic syndrome (NS) is a common pediatric kidney disease estimated to affect 16/100,000 children worldwide.[1]
We confirmed the requirement for CLVS1 in human podocyte homeostasis in vitro by demonstrating reduced viability and endocytosis in podocytes that are deficient in CLVS1, we showed that this phenotype could be rescued with corticosteroid treatment
In the present study, we showed that CLVS1 encodes an essential component of podocyte clathrin-mediated endocytosis (CME) and that a rare homozygous variant in this gene (p.H310Y) is a potential cause of corticosteroidresponsive NS

Summary

Introduction

Childhood nephrotic syndrome (NS) is a common pediatric kidney disease estimated to affect 16/100,000 children worldwide.[1]. The p.H310Y variant is shown to be deleterious to clavesin-1 function in vitro and in vivo and affects binding to the ligand, alpha-tocopherol transfer protein. This reduced affinity for a transporter of the antioxidant alpha-tocopherol, causes an increased accumulation of reactive oxygen species (ROS) in CLVS1 knockout and homozygous H310Y knockin podocytes. Pharmacological inhibition of ROS accumulation is sufficient to rescue the aberrant viability phenotype in podocytes with reduced functional clavesin-1, revealing new possible therapeutic strategies for patients with NS due to CLVS1 defects and possible adjunct therapy for children with the more common idiopathic NS

Methods

Results

Conclusion