Protein S-glutathionylation and sex dimorphic effects on hydrogen peroxide production by dihydroorotate dehydrogenase in liver mitochondria

Dihydroorotate dehydrogenase (DHODH) oxidizes dihydroorotate to orotate for pyrimidine biosynthesis, donating electrons to the ubiquinone (UQ) pool of mitochondria. DHODH has a measurable rate for hydrogen peroxide (H2O2) production and thus contributes to cellular changes in redox tone. Protein S-glutathionylation serves as a negative feedback loop for the inhibition of H(2)O(2 )by several alpha-keto acid dehydrogenases and respiratory complexes in mitochondria, as well as ROS sources in liver cytoplasm. Here, we report this redox signaling mechanism also inhibits H(2)O(2 )production by DHODH in liver mitochondria isolated from male and female C57BL6N mice. We discovered that low amounts of the glutathionylation catalyst, disulfiram (50-500 nM), almost abolished H(2)O(2 )production by DHODH in mitochondria from male mice. Similar results were collected with diamide, however, higher doses (1000-5000 mu M) were required to elicit this effect. Disulfiram and diamide also significantly suppressed H(2)O(2 )production by DHODH in female liver mitochondria. However, liver mitochondria from female mice were more resistant to disulfiram or diamide-mediated inhibition of H(2)O(2 )genesis when compared to samples from males. Analysis of the impact of disulfiram and diamide on DHODH activity revealed that both compounds inhibited the dehydrogenase directly, however the effect was less in female mice. Additionally, disulfiram and diamide impeded the use of dihydroorotate fueled oxidative phosphorylation in mitochondria from males and females, although samples collected from female rodents displayed more resistance to this inhibition. Taken together, our findings demonstrate H(2)O(2 )production by DHODH can be inhibited by glutathionylation and sex can impact this redox modification.

Automated summary

Dihydroorotate dehydrogenase (DHODH) is involved in pyrimidine biosynthesis and can be inhibited by S-glutathionylation, which is a negative feedback loop for hydrogen peroxide (H2O2) production. This redox modification can be influenced by sex differences in liver mitochondria.