Direct and quantitative analysis of altered metabolic flux distributions and cellular ATP production pathway in fumarate hydratase-diminished cells.

ABSTRACT: Fumarate hydratase (FH) is an enzyme in the tricarboxylic acid (TCA) cycle, biallelic loss-of-function mutations of which are associated with hereditary leiomyomatosis and renal cell cancer. However, how FH defect modulates intracellular metabolic fluxes in human cells has remained unclear. This study aimed to reveal metabolic flux alterations induced by reduced FH activity. We applied ¹³C metabolic flux analysis (¹³C-MFA) to an established cell line with diminished FH activity (FH^dim) and parental HEK293 cells. FH^dim cells showed reduced pyruvate import flux into mitochondria and subsequent TCA cycle fluxes. Interestingly, the diminished FH activity decreased FH flux only by about 20%, suggesting a very low need for FH to maintain the oxidative TCA cycle. Cellular ATP production from the TCA cycle was dominantly suppressed compared with that from glycolysis in FH^dim cells. Consistently, FH^dim cells exhibited higher glucose dependence for ATP production and higher resistance to an ATP synthase inhibitor. In summary, using FH^dim cells we demonstrated that FH defect led to suppressed pyruvate import into mitochondria, followed by downregulated TCA cycle activity and altered ATP production pathway balance from the TCA cycle to glycolysis. We confirmed that ¹³C-MFA can provide direct and quantitative information on metabolic alterations induced by FH defect.

SUBMITTER: Noguchi S 

PROVIDER: S-EPMC7400513 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature