StarD7 Protein Deficiency Adversely Affects the Phosphatidylcholine Composition, Respiratory Activity, and Cristae Structure of Mitochondria.

Phosphatidylcholine (PC) is a major phospholipid of mitochondria, comprising 40-50% of both the outer and the inner membranes. However, PC must be imported from its production organelles because mitochondria lack the enzymes essential for PC biosynthesis. In a previous study, we found that StarD7 mediates the intracellular transfer of PC to mitochondria. Therefore, in this study, we analyzed the contribution of StarD7 to the maintenance of mitochondrial phospholipid content and function using siRNA-mediated knockdown and knock-out (KO) of the StarD7 gene in HEPA-1 cells. Real time analysis of respiratory activity demonstrated that the oxygen consumption rate and activity of mitochondrial complexes were impaired in StarD7-KD cells. To confirm these results, we established StarD7-KO HEPA-1 cells by double nicking using CRISPR/Cas9n. As expected, StarD7-KD and -KO cells showed a significant reduction in mitochondrial PC content. The ATP level and growth rate of KO cells were notably lower compared with wild-type cells when cultured in glucose-free galactose-containing medium to force cells to rely on mitochondrial ATP production. In KO cells, the level of the MTCO1 protein, a primary subunit of complex IV, was reduced without a concomitant decrease in its mRNA, but the level was restored when StarD7-I was overexpressed. StarD7-KO cells showed impaired formation of the mitochondrial supercomplexes and exhibited a disorganized cristae structure, with no changes in optic atrophy 1 protein. These findings indicate that StarD7 plays important roles in maintaining the proper composition of mitochondrial phospholipids as well as mitochondrial function and morphogenesis.

PON3 knockout mice are susceptible to obesity, gallstone formation, and atherosclerosis.

We report the engineering and characterization of paraoxonase-3 knockout mice (Pon3KO). The mice were generally healthy but exhibited quantitative alterations in bile acid metabolism and a 37% increased body weight compared to the wild-type mice on a high fat diet. PON3 was enriched in the mitochondria-associated membrane fraction of hepatocytes. PON3 deficiency resulted in impaired mitochondrial respiration, increased mitochondrial superoxide levels, and increased hepatic expression of inflammatory genes. PON3 deficiency did not influence atherosclerosis development on an apolipoprotein E null hyperlipidemic background, but it did lead to a significant 60% increase in atherosclerotic lesion size in Pon3KO mice on the C57BL/6J background when fed a cholate-cholesterol diet. On the diet, the Pon3KO had significantly increased plasma intermediate-density lipoprotein/LDL cholesterol and bile acid levels. They also exhibited significantly elevated levels of hepatotoxicity markers in circulation, a 58% increase in gallstone weight, a 40% increase in hepatic cholesterol level, and increased mortality. Furthermore, Pon3KO mice exhibited decreased hepatic bile acid synthesis and decreased bile acid levels in the small intestine compared with wild-type mice. Our study suggests a role for PON3 in the metabolism of lipid and bile acid as well as protection against atherosclerosis, gallstone disease, and obesity.