Molecular and physiological characterisation of Arabidopsis thaliana mutants with altered fermentative metabolism

In plants hypoxia, i.e. the scarce availability of oxygen in tissues, is a common condition. It can be caused by external stressful events, such as floods, or be present physiologically, in tissues in which the diffusion of oxygen is limited, such as in meristems and fruits. This also happens during the in vitro regeneration of plants, when cell proliferation leads to the formation of structures called calli, inside which oxygen is scarce. When cells do not have access to enough oxygen, they cannot use the electron transport chain to regenerate oxidized cofactors; therefore, they use alternative metabolic pathways such as lactic fermentation, alcoholic fermentation, and non-circular citric acid pathways to allow glycolysis to continue. In this work, we analyzed plants of the species Arabidopsis thaliana mutant for genes fundamental for these metabolic pathways (individually or in higher-order combinations of mutations), namely lactate dehydrogenase (LDH), the two hypoxia-inducible isoforms of pyruvate decarboxylase (PDC1/2) and alanine aminotransferases (ALTAA1/2). We subjected the plants to treatments in anoxic and hypoxic environments and then evaluated their growth and survival and post-translational modifications of proteins (lactylation and acetylation). We then analyzed the in vitro regeneration of these mutants, evaluating the time required for the formation of sprouts. In addition, we analyzed differences in gene expression between mutant and wild-type plants at different times of regeneration using the RNA-seq technique. The work revealed for the first time the differential contribution of pyruvate pathways in hypoxic response and callus regeneration, while also shedding new light on the role of fermentations in de novo organogenesis.