Investigation of cellular functions of ADCK4 and its relation to nephrotic syndrome

Abstract

Mutations in ADCK4 (aarF domain containing kinase 4, also known as COQ8B) usually manifest as steroid-resistant nephrotic syndrome (SRNS) and sometimes accompany medullary nephrocalcinosis or extrarenal symptoms including seizure. ADCK4 has a helical domain, ABC1 domain, and a kinase-like domain and is similar to yeast Abc1/Coq8 which is required for coenzyme Q10 (CoQ10) biosynthesis. ADCK4 localizes to mitochondrial matrix and associates with the inner membranes. Interestingly, patients with ADCK4 mutations exhibit reduced cellular CoQ10 contents. However, the role of ADCK4 at the molecular level is unclear. To address this, we knocked out ADCK4 in cultured podocytes and HK-2, a proximal tubule cell line using CRISPR/Cas9. ADCK4 knockout did not affect cell viability in both cell lines. The basal levels of CoQ10 levels of podocytes were three-fold higher compared to those of HK-2 cells. The levels of CoQ10 were severely decreased in ADCK4 knockout podocytes compared to control podocytes, whereas CoQ10 contents in HK-2 cells were not different. As CoQ10 is required for electron transfer from complex I and II to complex III of mitochondrial respiratory chain, we measured complex II+III activities. Complex II+III activities were defective only in podocytes, but not in HK-2 cells. In addition, transmission electron microscopy showed that ADCK4 knockout resulted in disintegrated mitochondria and loss of cristae formation in cultured podocytes, but not in HK-2 cells, suggesting that ADCK4 is indispensable for maintaining mitochondrial function in podocytes. In conclusion, our results showed that podocytes are more vulnerable to loss of ADCK4 than HK-2 cells and this may explain why individuals with ADCK4 mutations mostly manifest only SRNS.