Kidney Week

Abstract: TH-PO468

Therapeutic Strategies for PodR138Q Nephrotic Syndrome

Session Information

• Genetic Diseases: Glomerulopathies - I
  November 02, 2023 | Location: Exhibit Hall, Pennsylvania Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Genetic Diseases of the Kidneys

• 1202 Genetic Diseases of the Kidneys: Non-Cystic

Authors

• Lu, Pei-Chen, University of Bristol, Bristol, United Kingdom

Pei-Chen Lu,

• Welsh, Gavin Iain, University of Bristol, Bristol, Bristol, United Kingdom

Gavin Iain Welsh,

• Saleem, Moin, University of Bristol, Bristol, Bristol, United Kingdom

Moin Saleem,

Background

The most common missense mutation in podocin, Pod^R138Q, results in steroid-resistant nephrotic syndrome. Pod^R138Q causes the podocin to be trafficked incorrectly resulting in it being trapped in ER and degraded leading to aberrant function of the slit diaphragm. Understanding the mechanism of this mis-trafficking and degradation may lead to new therapeutic strategies for treating this disease.

Methods

Both Pod^WT and Pod^R138Q were overexpressed in immortalized human podocytes. These cells were treated with MG132 (proteosome inhibitor), bafilomycin A1 (lysosome inhibitor), and kifunensine (Kif) (ER mannosidase I inhibitor). Podocin localization/tyrafficking was analyzed by western blotting and immunofluorescence.

Results

Pod^R138Q had higher co-localization coefficients with ER than Pod^WT (Fig 1). Pod^WT was mainly degraded by the lysosome, and Pod^R138Q degraded only by the proteosome (Fig 2A, 2B). Blocking podocin proteosomal degradation with MG132 resulted in Pod^R138Q being localized to plasma membrane lipid rafts (Fig 2C). Kif leads to the accumulation of high mannose N-glycan podocin (Fig 2B). High mannose N-glycan Pod^R138Q had affinity to calnexin (Fig 2D), and this implies that Pod^R138Q is degraded via ER-associated degradation (ERAD).

Conclusion

Pod^WT is mainly degraded by the lysosome whereas Pod^R138Q undergoes ERAD. By using a proteasome inhibitor, Pod^R138Q can be rescued correctly trafficked to lipid rafts. This finding could suggest a new therapeutic target for medical intervention for Pod^R138Q nephrotic syndrome.

The colocalization coefficients between podocin and ER.

Podocin treated by bafilomycin A1, MG132, and Kif (2A, 2B). Sucrose gradient centrifugation showed MG132 treated Pod^R138Q in lipid rafts (2C). Co-IP showed high mannose Pod^R138Q interacting with calnexin (2D).

Funding

• Private Foundation Support