Kidney Week

Abstract: FR-PO0760

Phosphorylative State of Podocin Affects Podocyte Morphology and Function

Session Information

• Glomerular Diseases: Cell Homeostasis and Novel Injury Mechanisms
  November 07, 2025 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Glomerular Diseases

• 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology

Authors

• Butt, Linus, Universitatsklinikum Koln, Cologne, NRW, Germany

Linus Butt, MD

• Unnersjö-Jess, David, SciLifeLab, Stockholm, Stockholm County, Sweden

David Unnersjö-Jess, PhD

• Höhne, Martin, Universitatsklinikum Koln, Cologne, NRW, Germany

Martin Höhne, PhD

• Reilly, Dervla, Universitatsklinikum Koln, Cologne, NRW, Germany

Dervla Reilly

• Hahnfeldt, Robert, Universitatsklinikum Koln, Cologne, NRW, Germany

Robert Hahnfeldt

• Bozek, Kasia, University of Cologne Center for Molecular Medicine Cologne, Cologne, NRW, Germany

Kasia Bozek, PhD

• Schermer, Bernhard, Universitatsklinikum Koln, Cologne, NRW, Germany

Bernhard Schermer, MD

• Benzing, Thomas, Universitatsklinikum Koln, Cologne, NRW, Germany

Thomas Benzing, MD, PhD, FASN

Background

NPHS2 is one the most frequently mutated genes in hereditary Nephrotic Syndrome. Its gene product podocin is central to the multiprotein complex at the slit diaphragm, acts as a scaffolding protein and facilitates signaling events. Multiple lines of evidence suggest mechanosensory properties of the slit diaphragm complex that depend on podocin. Firstly, our laboratory previously demonstrated that the podocin homologue in C. elegans, MEC-2, recruits ion channel MEC-4/MEC-10 to form a mechanosensor required for touch sensation. Secondly, podocin forms a complex with ion channel TRPC6, similar to its orthologue MEC-2. Thirdly, there is a preferential alignment of the slit diaphragm along the axis of the highest physical strain, which is lost in podocin mutants. Podocin belongs to the evolutionary conserved SPFH protein family. Recently, the structure of several SPFH-proteins was resolved using cryo-EM. These data show that SPFH-domain proteins oligomerize to form cone-shape cage-like structures. This aligns with our observation that podocin oligomerizes to form megadalton protein-lipid complexes. We and others showed that multimerization of podocin and further SPFH-proteins is affected by post-translational phosphorylation. There are multiple evolutionarily conserved phosphorylation sites on podocin, one of which was affected in a patient mutation (p.T232I) and one was shown to be involved in binding to nephrin (p.S380). Structural predictions indicate a direct effect of dephosphorylation of these site on the conformation of podocin multimers.

Methods

To investigate the effect of phosphorylation in vivo, we generated two novel mouse lines using CRISPR/Cas9. As a first step, we have comprehensively characterized podocyte morphology using our workflow of STED microscopy with automated analysis of foot process morphology.

Results

Homozygous Nphs2^T234I and Nphs2^S382A mice develop a mild pattern of foot process effacement during ageing. In addition, crossing these lines to a mouse line carrying a Nphs2 null allele (Nphs2^Dexon5) results in a full-blown picture of FSGS in Nphs2^T234I/Dexon5 and Nphs2^T234I/Dexon5 mice, whereas Nphs2^{wildtype/Dexon5} remain healthy.

Conclusion

In summary, our in vivo data indicate a direct functional effect of podocin phosphorylation, possibly through affecting podocin multimerization.

Digital Object Identifier (DOI)

• doi: 10.1681/ASN.2025pg9k5dhz