D-Lactate dehydrogenase links methylglyoxal degradation to the ETC through CYTc in A. thaliana. Plant Phys

WELCHEN ELINA; SCHMITZ J; FUCHS F; GARCÍA LUCILA ; WAGNER S; WIENSTROER J; SCHERTL P; BROUN H-P; SCHWARZLÄNDER M; GONZALEZ DANIEL H; MAURINO VERÓNICA

PLANT PHYSIOLOGY.

AMER SOC PLANT BIOLOGISTS

Lugar: Rockville; Año: 2016

0032-0889

Glycolysis generates methylglyoxal (MGO) as an unavoidable, cytotoxic by-product in plant cells. MGO scavenging is performed by the glyoxalase system, which produces D-lactate as an end product. D-lactate dehydrogenase is encoded by a single gene in A. thaliana (D-LDH; At5g06580). It catalyses in vitro the oxidation of D-lactate to pyruvate using FAD as cofactor; knowledge of its function in the context of the plant cell remains sketchy. Blue native polyacrylamide electrophoresis of mitochondrial extracts combined with in gel activity assays using different substrates and tandem mass spectrometry allowed us to definitely show that D-LDH acts specifically on D-lactate, it is active as a dimer and does not associate with respiratory supercomplexes of the inner mitochondrial membrane. The combined use of cytochrome c (CYTc) loss-of-function mutants and respiratory Complex III inhibitors showed that CYTc acts as the in vivo electron acceptor of D-LDH. CYTc loss-of-function mutants, as well as the D-LDH mutants, were more sensitive to D-lactate and MGO, indicating that they function in the same pathway. In addition, overexpression of D-LDH and CYTc increased tolerance to D-lactate and MGO. Together with fine-localization of D-LDH, the functional interaction with CYTc in vivo strongly suggests that D-lactate oxidation takes place in the mitochondrial intermembrane space delivering electrons to the respiratory chain through CYTc. These results provide a comprehensive picture of the organisation and function of D-LDH in the plant cell and exemplify how the plant mitochondrial respiratory chain can act as a multi-functional electron sink for reductant from cytosolic pathways.