ASN's Mission

To create a world without kidney diseases, the ASN Alliance for Kidney Health elevates care by educating and informing, driving breakthroughs and innovation, and advocating for policies that create transformative changes in kidney medicine throughout the world.

learn more

Contact ASN

1401 H St, NW, Ste 900, Washington, DC 20005

email@asn-online.org

202-640-4660

The Latest on X

Kidney Week

Please note that you are viewing an archived section from 2021 and some content may be unavailable. To unlock all content for 2021, please visit the archives.

Abstract: PO1338

Discovery of Podocyte-Specific Interaction Partners of the Nephrotic Syndrome-Associated Protein NOS1AP

Session Information

Category: Genetic Diseases of the Kidneys

  • 1002 Genetic Diseases of the Kidneys: Non-Cystic

Authors

  • Ball, David A., Boston Children's Hospital Department of Pediatrics, Boston, Massachusetts, United States
  • Buerger, Florian, Boston Children's Hospital Department of Pediatrics, Boston, Massachusetts, United States
  • Hildebrandt, Friedhelm, Boston Children's Hospital Department of Pediatrics, Boston, Massachusetts, United States
  • Majmundar, Amar J., Boston Children's Hospital Department of Pediatrics, Boston, Massachusetts, United States
Background

Recessive NOS1AP mutations are a novel Mendelian cause of nephrotic syndrome (NS) in humans and mice (Majmundar Sci Adv 2021). The NS patient mutation p.C143Y destabilizes the predicted structure of the NOS1AP phosphotyrosine-binding domain, impairs NOS1AP-dependent actin remodeling in immortalized podocytes, and causes abnormal kidney organoid glomerulogenesis. Here, we aimed to discover podocyte-specific NOS1AP interaction partners, which may mediate NOS1AP functions in the podocyte and whose binding is abrogated by the patient mutation p.C143Y.

Methods

Protein interaction data from candidate immunoprecipitation, mass spectrometry, and yeast two-hybrid studies were queried from the literature and public datasets (Orchard NAR 2014; Oughtred Protein Sci 2020; Szklarczyk NAR 2019). Gene expression was queried from kidney single cell mRNA sequencing (scRNAseq) datasets (Karaiskos JASN 2018; Menon Development 2018; Wang Cell Rep 2018; Wu JASN 2018). Protein interactions were validated by co-immunoprecipitation studies using tagged cDNA constructs.

Results

85 putative NOS1AP-interacting proteins were identified from candidate interaction and proteomics studies. Six interacting proteins (of 85) demonstrated co-expression with NOS1AP in podocyte clusters from at least three out of four kidney scRNAseq datasets (% cell expression z-score > 1). Four of six candidates (FYN, GSN, SNTA1, HSPA12A) were cloned into expression vectors for interaction studies. SNTA1 and HSPA12A exhibited bi-directional co-immunoprecipitation with wildtype NOS1AP upon co-overexpression in a podocyte cell line. Co-immunoprecipitation was, similarly, observed with the NS patient mutant NOS1APC143Y.

Conclusion

Our results suggest NOS1AP is co-expressed with and can physically interact with SNTA1 and HSPA12A in podocytes. These proteins may mediate NOS1AP functions in podocyte biology.

Funding

  • Other NIH Support