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Abstract: TH-PO758

Exploring the Pathogenicity of an Anillin Coding Variant in Proteinuric Kidney Disease

Session Information

Category: Glomerular Diseases

  • 1403 Podocyte Biology

Authors

  • Fischer, Matthew, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • Craig, Zie, University of Michigan Department of Molecular Cellular and Developmental Biology, Ann Arbor, Michigan, United States
  • Fermin, Damian, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • Berthier, Celine C., University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • El Saghir, Jamal, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • Larkina, Maria, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • Eddy, Sean, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • Kretzler, Matthias, University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
  • Miller, Ann L., University of Michigan Department of Molecular Cellular and Developmental Biology, Ann Arbor, Michigan, United States
  • Harder, Jennifer L., University of Michigan Michigan Medicine, Ann Arbor, Michigan, United States
Background

Mutations in slit diaphragm proteins and regulators of actin dynamics are implicated in the pathogenesis of proteinuric kidney disease (protKD). Coding variants of Anillin (ANLN), a scaffolding protein found at slit diaphragms, have been associated with FSGS, but mechanisms are unknown. The aim of this study is to investigate the pathogenicity of ANLN coding variants identified in individuals with protKD.

Methods

Expression of ANLN relative to proteinuria was assessed in bulkRNAseq profiles of microdissected kidney tissues from NEPTUNE (n=238 glom, n=316 tub) and living donor (n=8, 10) cohorts. WGS data in NEPTUNE (n=620) were interrogated for ANLN coding variants; allele frequencies were compared to gnomeAD. Kidney organoids (hKO) were generated from an iPSC line from a NEPTUNE participant harboring an ANLN coding variant and scRNA-seq (Sc) performed. Cell junction integrity and cytoskeletal dynamics were assessed by IF and live imaging in Xenopus laevis embryonic epithelial cells following morpholino knockdown of ANLN and replacement with ANLN WT or coding variant.

Results

Kidney tissue ANLN mRNA expression correlated with UPCR and negatively with eGFR. Ten previously uncharacterized variants of ANLN were found in 16 NEPTUNE individuals; 6 harbored one missense mutation (rs141247770) resulting in an I1109V substitution in the pleckstrin homology (PH) domain, which binds phospholipids. Sc profiling of hKO ANLN I1109V versus WT epithelial cells revealed perturbation of YAP and MAPK8 signaling pathways. Live imaging of X. laevis embryos expressing the ANLN variant revealed disrupted tight junction (ZO-1) integrity plus abnormalities in junctional F-actin and cell size and shape.

Conclusion

Our protKD cohort data are consistent with prior animal models showing that perturbed ANLN expression is associated with kidney disease. Functional studies in hKO and Xenopus models harboring the ANLN variant within the PH domain revealed disrupted cell-cell junction integrity, cytoskeletal dynamics, cytokinesis, and several dysregulated signaling pathways. Together, our findings reveal that alterations in both ANLN expression levels and function are critical in the pathogenicity of proteinuric kidney disease.

Funding

  • NIDDK Support