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 Twitter

Kidney Week

Abstract: FR-PO329

Characterization of the Human Podocin Short Isoform In Vivo Using a New Mouse Model

Session Information

Category: Genetic Diseases of the Kidneys

  • 1102 Genetic Diseases of the Kidneys: Non-Cystic

Authors

  • Butt, Linus, Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany
  • Unnersjö-Jess, David, Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany
  • Rinschen, Markus M., Aarhus Universitet Institut for Biomedicin, Aarhus, Denmark
  • Schermer, Bernhard, Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany
  • Benzing, Thomas, Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany
  • Höhne, Martin, Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital Cologne, Cologne, Germany
Background

To date, many mutations affecting podocyte-specific genes are known to cause steroid-resistant nephrotic syndrome, which usually progresses to end-stage renal disease due to a lack of effective therapeutic strategies. NPHS2, encoding the podocyte-specific protein podocin, is among the most frequently mutated genes in patients with hereditary steroid-resistant nephrotic syndromes. Previously, a short isoform of podocin lacking exon 5 was identified in human kidney tissue and characterized in vitro. However, its role in vivo remained elusive.

Methods

The short isoform of podocin is not found in any other organism except humans. To examine its functional relevance in vivo we generated a mouse line expressing the equivalent podocin isoform (podocinΔexon5) using CRISPR/Cas9 mediated genome editing. We characterized the phenotype of mice expressing podocinΔexon5 resembling the human short isoform. Targeted mass spectrometry and qPCR were used to compare protein and RNA levels of podocinwildtype and podocinΔexon5. STED microscopy following immunolabeling of slit diaphragm components was applied to visualize alterations of the podocytes’ foot process morphology.

Results

Homozygous podocinΔexon5 mice born heavily albuminuric, hardly produced any urine and did not survive past the first day after birth. Targeted mass spectrometry revealed decreased protein levels of podocinΔexon5 whereas RNA abundance was not different to the canonical form (podocinwildtype). STED microscopy revealed the complete absence of podocin at the podocytes’ slit diaphragm and severe morphological alterations of podocyte foot processes. Mice heterozygous for podocinΔexon5 were phenotypically unaffected but decreased podocin levels and altered foot process morphology could be detected.

Conclusion

The murine equivalent to the human short isoform of podocin is unable to stabilize the lipid-protein complex at the slit diaphragm. Reduction of podocin levels at the site of the slit diaphragm complex has a detrimental effect on podocyte morphology and decreases protein abundance of nephrin, the core protein of the extracellular slit diaphragm complex. In conclusion, our data support the hypothesis that podocin acts an orchestrator of the slit diaphragm complex, which allows for kidney ultrafiltration. The functional role of the short human isoform remains elusive.

Funding

  • Government Support – Non-U.S.