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Abstract: PO1999

Mitochondrial Damage in FSGS due to ANLN Mutation

Session Information

  • Podocyte Biology
    October 22, 2020 | Location: On-Demand
    Abstract Time: 10:00 AM - 12:00 PM

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Lane, Brandon M., Department of Pediatrics, Duke University School of Medicine, Durham, North Carolina, United States
  • Wu, Guanghong, Duke Molecular Physiology Institute, Durham, North Carolina, United States
  • Chryst-Stangl, Megan, Duke Molecular Physiology Institute, Durham, North Carolina, United States
  • Hall, Gentzon, Department of Medicine, Duke University School of Medicine, Durham, North Carolina, United States
  • Gbadegesin, Rasheed A., Department of Pediatrics, Duke University School of Medicine, Durham, North Carolina, United States
Background

We previously identified ANLN R431C as a cause of focal segmental glomerulosclerosis (FSGS). In addition to defects in actin bundling, targeted evaluation of this variant in cultured human podocytes identified disruption of AKT/mTOR signaling as a cause of ER stress and reduced podocyte viability. Creation of the orthologous R431C point mutation in mice confirmed the increased podocyte ER stress and identified mitochondrial damage as another possible feature of disease. To gain an unbiased view of the molecular mechanisms driving ANLN R431C induced disease, we used transcriptomic analysis and automated live cell imaging to interrogate cultured human podocytes.

Methods

Conditionally immortalized human podocytes overexpressing wildtype ANLN or the R431C variant were evaluated by mRNA-Seq and smRNA-Seq analysis to identify differentially expressed genes and microRNAs, as well as the molecular pathways involved. Potential therapeutic strategies were examined by evaluating cultured podocyte cellular and organelle-specific viability using automated live cell imaging.

Results

The top differentially expressed genes encode molecules that interact with previously identified pathological mechanisms including F-actin bundling (SYNPO2L) and AKT/ mTOR signaling (CAVIN3, KIT), with mTOR signaling identified as a pathway likely to be affected by ANLN R431C. A common feature of the top differentially expressed gene and microRNA candidates is the potential to regulate mitochondrial viability. When evaluated for changes in mitochondrial membrane potential, ANLN R431C podocytes displayed increased susceptibility to mitochondrial damage that could be rescued by treatment with AKT/mTOR pathway inhibitors. Additionally, compounds targeting improved mitochondrial viability through increased bioenergetic function (AP39), reduced oxidative stress (MitoQ, MitoTEMPO) and prevention of pore opening (Olesoxime) could all rescue the increased susceptibility to apoptosis in ANLN R431C podocytes.

Conclusion

Unbiased transcriptomic analysis confirmed that ANLN R431C disrupts AKT/mTOR signaling and actin cytoskeletal dynamics, resulting in increased ER stress and mitochondrial damage that reduce podocyte viability. Targeting various aspects of mitochondrial regulation may present viable alternative treatment strategies for FSGS due to defects in ANLN gene.