Oxaloacetic acid mediates ADP-dependent inhibition of mitochondrial complex II-driven respiration
Brian D Fink1, Fan Bai1, Liping Yu2,3
1From the Department of Internal Medicine/Endocrinology and Metabolism.
Abstract:
We recently reported a previously unrecognized mitochondrial respiratory phenomenon. When [ADP] was held constant ("clamped") at sequentially increasing concentrations in succinate-energized muscle mitochondria in the absence of rotenone (commonly used to block complex I), we observed a biphasic, increasing then decreasing, respiratory response. Here we investigated the mechanism. We confirmed decades-old reports that oxaloacetate (OAA) inhibits succinate dehydrogenase (SDH). We then used an NMR method to assess OAA concentrations (known as difficult to measure by MS) as well as those of malate, fumarate, and citrate in isolated succinate-respiring mitochondria. When these mitochondria were incubated at varying clamped ADP concentrations, respiration increased at low [ADP] as expected given the concurrent reduction in membrane potential. With further increments in [ADP], respiration decreased associated with accumulation of OAA. Moreover, a low pyruvate concentration, that alone was not enough to drive respiration, was sufficient to metabolize OAA to citrate and completely reverse the loss of succinate-supported respiration at high [ADP]. Further, chemical or genetic inhibition of pyruvate uptake prevented OAA clearance and preserved respiration. In addition, we measured the effects of incremental [ADP] on NADH, superoxide, and H2O2 (a marker of reverse electron transport from complex II to I). In summary, our findings, taken together, support a mechanism (detailed within) wherein succinate-energized respiration as a function of increasing [ADP] is initially increased by [ADP]-dependent effects on membrane potential but subsequently decreased at higher [ADP] by inhibition of succinate dehydrogenase by OAA. The physiologic relevance is discussed.
Insights
Mitochondrial respiration initially increases with higher adenosine diphosphate (ADP) levels but then decreases due to oxaloacetate (OAA) inhibiting succinate dehydrogenase (SDH). Pyruvate metabolism resolves this OAA-induced inhibition.
Area of Science:
- Mitochondrial physiology
- Bioenergetics
- Biochemistry
Background:
- Mitochondrial respiration is crucial for cellular energy production.
- Adenosine diphosphate (ADP) levels influence respiratory rates.
- Succinate dehydrogenase (SDH) is a key enzyme in the citric acid cycle and electron transport chain.
Purpose of the Study:
- To elucidate the mechanism behind a previously observed biphasic mitochondrial respiratory response.
- To investigate the role of oxaloacetate (OAA) in regulating succinate-energized respiration.
- To understand the impact of varying adenosine diphosphate (ADP) concentrations on mitochondrial function.
Main Methods:
- Utilized clamped adenosine diphosphate (ADP) concentrations in isolated succinate-respiring mitochondria.
- Employed Nuclear Magnetic Resonance (NMR) spectroscopy to quantify oxaloacetate (OAA) and related metabolites.
- Assessed the effects of pyruvate and inhibitors on mitochondrial respiration, NADH, superoxide, and hydrogen peroxide (H2O2).
Main Results:
- Respiration initially increased with rising ADP due to membrane potential changes.
- Higher ADP concentrations led to decreased respiration, correlated with oxaloacetate (OAA) accumulation.
- Pyruvate addition metabolized OAA, reversing the ADP-induced respiratory inhibition; inhibiting pyruvate uptake prevented OAA clearance.
Conclusions:
- Succinate-energized respiration is biphasic: initially enhanced by ADP-dependent membrane potential shifts, then inhibited at higher ADP by OAA-induced succinate dehydrogenase (SDH) blockade.
- Oxaloacetate (OAA) accumulation is a key factor in the decline of respiration at elevated adenosine diphosphate (ADP) levels.
- Pyruvate metabolism plays a critical role in clearing OAA and maintaining mitochondrial respiratory function under varying substrate conditions.
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