Kidney Week

Abstract: TH-OR062

A Live Cell In Vitro System to Assess Functional Rescue of Polycystin Missense Variants

Session Information

• Genetic Diseases with a Focus on ADPKD Mechanisms, Models, and Medicines
  October 22, 2026 | Location: Mile High Ballroom 4D, Convention Center
  Abstract Time: 05:40 PM - 05:50 PM

Category: Genetic Diseases of the Kidneys

• 1201 Genetic Diseases of the Kidneys: Cystic (Monogenic)

Authors

• Cai, Yiqiang, Yale University, New Haven, Connecticut, United States

Yiqiang Cai, MD, PhD

• Dong, Ke, Yale University, New Haven, Connecticut, United States

Ke Dong, MD

• Rehman, Michael, Yale University, New Haven, Connecticut, United States

Michael Rehman, PhD

• Cordido, Adrian, Yale University, New Haven, Connecticut, United States

Adrian Cordido, PhD

• Wei, Zemeng, Yale University, New Haven, Connecticut, United States

Zemeng Wei, PhD

• Tian, Xin, Yale University, New Haven, Connecticut, United States

Xin Tian, MD

• Meng, Xinmiao, Yale University, New Haven, Connecticut, United States

Xinmiao Meng, MPH

• Somlo, Stefan, Yale University, New Haven, Connecticut, United States

Stefan Somlo, MD

Group or Team Name

• Somlo group

Background

ADPKD is caused by mutations in genes encoding polycystin-1 (PC1) and polycystin-2 (PC2). The steady-state expression and ciliary trafficking of PC1 requires PC2, with PC1 as the rate limiting functional component. Some pathogenic mutations that disrupt PC2 trafficking to cilia can be rescued by alanine substitution at Ser829, but whether restoring ciliary trafficking of mutant PC2 also restores the functional PC1/PC2 complex remains unknown. Here, we developed a system to (1) visualize co-delivery of endogenous PC1 with rescued PC2 mutants in live renal epithelial cells and (2) assess functional rescue by the ability to suppress Glis2 upregulation.

Methods

We made constructs in which NeonGreen-tagged PC2 (wild-type or mutant) was linked via "PT2A" self-cleaving peptides to the ciliary marker AppleRed–NPHP3-NP200. Immortalized Pkd2-null kidney epithelial cells with endogenous Halo-tagged PC1 were transduced with PT2A constructs encoding PC2 or selected mutants alone or in combination with S829A. Live-cell confocal imaging of ciliated Halo-ligand–labeled cells assessed ciliary PC1/PC2 trafficking. Glis2 immunoblotting assessed functional rescue.

Results

Live-cell imaging showed that D511V, W414G, and C632R PC2 mutants with unmodified Ser829 failed to localize to cilia. PC1 was detectable in cilia with wild-type PC2 and S829A but not with the mutants. D511V combined with S829A trafficked to cilia and restored ciliary PC1, whereas W414G;S829A did not enter cilia and PC1 also remained absent from cilia. These findings support a model in which PC1 and PC2 traffic to cilia as an obligate complex and indicate that distinct mechanisms may be required to overcome impaired ciliary trafficking of different PC2 mutations. Glis2 was elevated in Pkd2-null cells; re-expression of either PC2 or S829A suppressed elevated Glis2 levels, whereas PC2 mutants that did not traffic to cilia did not. This supports the role of Glis2 as a downstream readout of PC1/PC2 function.

Conclusion

We have established an imaging platform for determining rescue of ciliary trafficking of PC1/PC2 in live cells and a biochemical assay for determining functional efficacy of mutant PC1/PC2 complexes localized in cilia by suppression of Glis2 upregulation. This platform provides a means by which to determine which missense mutations will restore PC1/PC2 function after correction of ciliary delivery defects.

Acknowledgment

This research is funded by NIH (SS).

Funding

• NIDDK Support