Abstract: PO0622
APOL1 Risk Variants Mediate Increased Oxidative Phosphorylation Complexes Biogenesis and Mitochondrial Dysfunction
Session Information
- CKD Mechanisms - 2
October 22, 2020 | Location: On-Demand
Abstract Time: 10:00 AM - 12:00 PM
Category: CKD (Non-Dialysis)
- 2103 CKD (Non-Dialysis): Mechanisms
Authors
- Ge, Mengyuan, Katz Family Division of Nephrology and Hypertension/ Drug Discovery Center, University of Miami, Miami, Florida, United States
- Ducasa, Gloria Michelle, Katz Family Division of Nephrology and Hypertension/ Drug Discovery Center, University of Miami, Miami, Florida, United States
- Mallela, Shamroop Kumar, Katz Family Division of Nephrology and Hypertension/ Drug Discovery Center, University of Miami, Miami, Florida, United States
- Liu, Shaoyi, Social Profit Network Research Lab, Alexandria LaunchLabs, New York, New York, United States
- Szeto, Hazel H., Social Profit Network Research Lab, Alexandria LaunchLabs, New York, New York, United States
- Kopp, Jeffrey B., Kidney Disease Section, NIDDK, NIH, Bethesda, Maryland, United States
- Fontanesi, Flavia, Department of Biochemistry and Molecular Biology, University of Miami, Miami, Florida, United States
- Merscher, Sandra M., Katz Family Division of Nephrology and Hypertension/ Drug Discovery Center, University of Miami, Miami, Florida, United States
- Fornoni, Alessia, Katz Family Division of Nephrology and Hypertension/ Drug Discovery Center, University of Miami, Miami, Florida, United States
Background
Susceptibility to focal segmental glomerulosclerosis (FSGS) in African Americans is associated with genetic variants of the Apolipoprotein L1 gene (APOL1) named G1 and G2. APOL1 risk variants (RV) are a major driver of mitochondrial dysfunction. The mitochondrial specific lipid cardiolipin (CL) interacts with oxidative phosphorylation (OXPHOS) complexes and plays an important role in the biogenesis of OXPHOS complexes. While APOL1 function was assessed in tagged and overexpressed systems, studies evaluating the functions of endogenous APOL1 protein are missing.
Methods
We studied mitochondrial function using human urinary podocyte-like epithelial cells (HUPECs) established from patients with FSGS carrying different APOL1 alleles. Protein and mRNA levels were measured by WB and quantitative PCR respectively. TEM was performed to study mitochondrial morphology. OXPHOS complexes were studied by BN-PAGE analysis followed by WB. To study how APOL1 RV contributes to mitochondrial dysfunction, we purified APOL1-6xHis protein using HeLa cells infected with lentivirus carrying the APOL1 G0, G1 under the CMV promoter, followed by protein-lipid overlay assay.
Results
The expression of endogenous APOL1 was decreased in HUPECs carrying RVs (G1/G2 HUPECs) when compared to G0/G0 carrying HUPECs. We observed reduced mitochondrial function in the presence of increased OXPHOS complexes in G1/G2 HUPECs. Using TEM, reduced mitochondrial matrix density and increased mitochondrial area were detected in G1/G2 HUPECs. Hyperbranched mitochondria in G1/G2 HUPECs were accompanied by a significant increase in the mRNA levels of mitochondrial fission and fusion proteins FIS1 and MFN1. The affinity of APOL1 G1 to CL was significantly higher than the affinity of APOL1 G0 to CL, when normalized to 6xHis tagged APOL1 expression. We found the mRNA level of cardiolipin synthase (CRLS1) was significantly increased in G1/G2 HUPECs, which is consistent with the overexpression of OXPHOS complexes in G1/G2 HUPECs.
Conclusion
Our findings indicate that endogenous APOL1 RV expression in human podocytes is associated with mitochondrial dysfunction in the presence of increased OXPHOS complexes, and that APOL1 RVs interact with CL thus interfering with CL function in mitochondria.
Funding
- NIDDK Support