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Kidney Week

Abstract: FR-PO377

Oligomerization of APOL1 Risk Variants After Mitochondrial Translocation

Session Information

  • CKD: Mechanisms - II
    November 08, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
    Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

  • 2103 CKD (Non-Dialysis): Mechanisms


  • Shah, Shrijal, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Lannon, Herbert F., Beth Israel Deaconess Medical Institute, Boston, Massachusetts, United States
  • Pollak, Martin R., Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, Massachusetts, United States
  • Friedman, David J., Beth Israel Deaconess Medical Center/Harvard Medical School, Boston, Massachusetts, United States

Inheriting two copies of APOL1 risk variants (G1 and G2) increases susceptibility to chronic kidney disease in African Americans. G1 and G2 are toxic, gain-of-function variants despite their recessive mode of inheritance. APOL1 multimerization has been proposed as an explanation for recessive gain-of-function.


Immunoprecipitation experiments were performed in HEK293 cells with co-transfection of constructs expressing APOL1 (G0, G1, or G2) tagged with FLAG or MYC. Oligomerization of untagged APOL1 was assessed in tetracycline inducible APOL1-expressing HEK293 TRex cells using blue native PAGE. Mitochondrial-enriched and cytosolic fractions were prepared by differential centrifugation. TOMM20 was knocked down with siRNA 48 hours prior to induction of APOL1 expression. APOL1-induced cytotoxicity was determined using the Muti-Tox Fluor Mutliplex cytotoxicity assay kit from Promega.


After co-transfection of FLAG- and MYC-tagged APOL1, immunoprecipitation of FLAG followed by Western blot for MYC demonstrated APOL1-APOL1 binding. Blue native PAGE of APOL1 expressing cell lines revealed that G1 and G2 tend to form large oligomers whereas G0 remains mostly monomeric. When we fractionated cells via differential centrifugation, we found that the oligomers were present mostly in the mitochondrial-enriched fractions. Knockdown of the mitochondrial outer membrane protein TOMM20 blocked APOL1 mitochondrial import and eliminated both APOL1 oligomer formation and APOL1-induced cytotoxicity.


APOL1 molecules can interact with other APOL1 molecules and risk variants have a greater tendency to form large oligomers. The oligomers are mostly located in the mitochondrial-enriched cell fraction and inhibiting mitochondrial import of APOL1 dramatically reduces the formation of oligomers, suggesting that mitochondrial import of APOL1 is necessary for subsequent oligomerization of G1 and G2. Whether these APOL1 oligomers directly cause cytotoxicity remains to be answered.


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