Playing with Pyr: alternate sources of mitochondrial pyruvate fuel plant respiration

One of the most distinctive features of plant metabolism is a high degree of flexibility at the pathway level. Consider the textbook example of glucose respiration by glycolysis, the TCA cycle, and the mitochondrial electron transport chain. In animals, if metabolic flow at any point through this standard pathway is disrupted, the organism will face major, often lethal, problems. By contrast, if you disrupt a particular enzyme along this pathway in plants, most of the time there will be no obvious effect. Plants still require the ability to respire glucose; they just usually have a viable metabolic detour. Metabolic flexibility likely represents a series of adaptations enabling plants to better survive diverse environmental stresses.

The pattern of enhanced metabolic complexity holds true in the case of plant transporters (Lee and Millar, 2016). The obvious intracellular transporter needed for glucose respiration is a mitochondrial pyruvate transporter to import cytosolic pyruvate produced by glycolysis and fuel the mitochondrial TCA cycle. In mice, loss of the mitochondrial pyruvate carrier (MPC) is embryo lethal (Vanderperre et al., 2016). By contrast, as Le et al. (2021) demonstrate in this issue, disruption of Arabidopsis MPC1 leads to a complete loss of the MPC protein complex, but no obvious growth phenotype. To be sure, previous research has detected an enhanced cadmium sensitive phenotype for Atmpc1 (He et al., 2019); but under most circumstances, MPC deficient plant mitochondria apparently obtain sufficient pyruvate by another route (or two). To identify and qualify the contribution of these routes, Le et al. combined detailed metabolic labeling with genetic and pharmaceutical inhibition of target enzymes.

When isolated mitochondria are fed with external ¹³C-labelled pyruvate, mpc1 mitochondria display negligible rates of pyruvate import. This means that another mitochondrial pyruvate transporter does not compensate for the loss of MPC; it must be something else. In plants, mitochondrial NAD malic enzyme (NAD-ME) interconverts malate and pyruvate and thus represents an internal source for mitochondrial pyruvate. When fed with external ¹³C-labelled malate, mpc1 mitochondria display increased production of ¹³C-labelled pyruvate. This means that mpc1 mitochondria display increased metabolic flux through NAD-ME, which may allow sufficient mitochondrial pyruvate production from imported malate.

When the authors produced a triple mutant me1.me2.mpc1, which lacked both NAD-ME and MPC1 activity, they observed a moderately retarded growth phenotype compared with me1.me2 or mpc1. Isolated me1.me2.mpc1 mitochondria could neither uptake external pyruvate nor produce pyruvate internally from malate. This result was surprising because if the me1.me2.mpc1 mitochondria cannot acquire pyruvate, the plants should display a more severe growth phenotype owing to respiratory malfunction. Could there be a third way for plant mitochondria to obtain pyruvate in vivo?

Diurnal metabolomic analysis revealed that me1.me2.mpc1 leaves produced few metabolite shifts compared to WT, which centered on increased alanine and 2-oxoglutarate levels. Both of these metabolites are products of a single reaction whereby pyruvate receives the nitrogen (amino) group of glutamate to become alanine. This reversible pyruvate:glutamate transamination is catalyzed by the enzyme alanine aminotransferase, which exists in both the mitochondria and cytosol and could allow alanine import to be a third source of mitochondrial pyruvate. Indeed, the authors found the growth of me1.me2.mpc1 seedlings were hypersensitive to cycloserine, an inhibitor of alanine aminotransferase (see Figure). Ultimately, it required the sequential disruption of three independent pyruvate transport mechanisms to observe the effect of mitochondrial pyruvate starvation in plants. Further metabolic flux studies will be required to understand how each pathway contributes to mitochondrial pyruvate supply under varying developmental and environmental circumstances.

Figure.

Genetic and pharmaceutical disruptions reveal three pathways of mitochondria pyruvate supply. mpc1 and especially me1.me.2.mpc1 seedlings show conditional growth phenotypes upon the inhibition of alanine aminotransferase by cycloserine treatment. Adapted fromLe et al. (2021) Figure 6.