Abstract 1177 Inhibition of mitochondrial transaminase impairs complex II respiration at low inner membrane potential

We previously showed that the transaminase inhibitor, aminooxyacetic acid (AAc), reduced respiration energized by succinate + glutamate in mitochondria isolated from mouse hindlimb muscle, an effect that required a reduction in membrane potential (ΔΨ) with resultant accumulation of oxaloacetate and consequent inhibition of succinate dehydrogenase (SDH). To specifically assess the effect of the mitochondrial transaminase, glutamic-oxaloacetic transaminase (GOT2), we performed respiratory and metabolic studies in wild-type (WT) and inducible muscle GOT2 knock-out (mGOT2KO) mice as well as in WT and CRISPR- generated GOT2 KO C2C12 cells. GOT2 gene expression was substantially, but not completely, reduced for both KO mice and cells. O2 flux in succinate (10mM) + glutamate (0.5mM)-energized mitochondria of mGOT2KO mice was reduced versus WT, but only at lower membrane potential generated by added ADP. This effect of ADP was associated with oxaloacetate accumulation in both WT and KO mice. Similar effects on respiration energized by 10mM succinate + 0.5mM glutamate (but not pyruvate + malate) were seen in GOT2KO C2C12 cells versus WT at lower ΔΨ generated by 1.0μM FCCP. But, interestingly, ADP addition to C2C12 cells (both KO and WT) did not reduce ΔΨ as much as FCCP and did not reduce respiration. Moreover, although metabolite data suggested oxaloacetate inhibition of SDH may have had a role in reducing respiration in the presence of FCCP, we could not directly detect oxaloacetate. Intact cell respiration by GOT2KO C2C12 cells (versus WT) assessed by Seahorse respirometry was reduced in the presence of FCCP. In summary, GOT2 KO reduced mitochondrial respiration at low ΔΨ. Based on differential substrate effects this occurred largely at complex II. The research is funded by the NIH award, 1 R01 DK123043-01A1 (William Sivitz, PI) and by the Iowa Fraternal Order of the Eagles (William Sivitz, PI).