The role of mitochondrial malate metabolism in plant energy homeostasis in <em>Arabidopsis thaliana</em>

Abstract

Malate plays a central role in plant physiology and metabolism. In mitochondria, it functions in the tricarboxylic acid cycle as a provider of carbon skeletons and is therefore essential for cellular energy metabolism. In addition, malate acts as an osmolyte, a regulator of pH, and a major component of root exudates. Beyond these functions, malate serves as an important carrier of carbon and reducing equivalents between chloroplasts, the cytosol, and mitochondria, making it critical for cellular homeostasis. Despite its importance, the specific contributions of malate-converting systems in distinct subcellular compartments remain poorly understood.<br /> In the first chapter of this work, we investigated the consequences of impaired mitochondrial malate-conversion capacity in <em>Arabidopsis thaliana</em>. To this end, we generated triple mutants (<em>mdh1xme1xme2</em>) lacking the major mitochondrial malate dehydrogenase (MDH1) and both subunits of NAD-dependent malic enzyme (ME1 and ME2). Under different photoperiods and light regimes, these mutants displayed a conditional phenotype. In particular, under short-day and low-light conditions, they exhibited reduced growth, impaired photosynthetic performance, a more reduced cytosolic redox state, and altered chloroplast ultrastructure compared with wild type. These findings demonstrate that mitochondrial malate metabolism is essential for coordinating respiratory energy supply, redox balance, and photosynthesis under energetic-limited conditions.<br /> In the second chapter, because ME is predominantly active as heterodimer, we asked whether ME1 might remain functional in the absence of ME2 and MDH1 in plant mitochondria. To address this, we analyzed T-DNA insertion mutants affecting only MDH1 and ME2. Although ME1 was undetectable in <em>mdh1xme2</em>, suggesting a functional loss similar to that observed in the triple mutant, the double mutants showed more severe defects in growth and photosynthesis under short-day, low-light conditions than either the wild type and the triple mutant. Whole-genome analysis revealed a major genomic rearrangement associated with the <em>MDH1</em> T-DNA insertion. This structural variant, which was absent from the triple mutant, altered gene expression and likely intensified the observed phenotype. These results underscore the importance of verifying genomic integrity in T-DNA insertion lines when interpreting genotype-phenotype relationships.<br /> In the final chapter, we examined organelle communication by analyzing the cytosolic redox status of mutants lacking mitochondrial MDH1 or MDH2. Disruption of the malate shuttles in mitochondria resulted in a more reduced cytosolic environment underscoring the importance of mitochondrial malate metabolism in the regulation of cellular redox balance.<br /> Overall, this thesis advances our understanding of mitochondrial malate metabolism and its central role in plant homeostasis, particularly in the coordination of growth, energy metabolism, and redox balance under changing environmental conditions.