Date Published: June 7, 2018
Publisher: Public Library of Science
Author(s): Mauro Tomassetti, Betiana S. Garavaglia, Cecilia V. Vranych, Natalia Gottig, Jorgelina Ottado, Hugo Gramajo, Lautaro Diacovich, Zonghua Wang.
Citrus canker is a disease caused by the phytopathogen Xanthomonas citri subsp. citri (Xcc), bacterium which is unable to survive out of the host for extended periods of time. Once established inside the plant, the pathogen must compete for resources and evade the defenses of the host cell. However, a number of aspects of Xcc metabolic and nutritional state, during the epiphytic stage and at different phases of infection, are poorly characterized. The 3-methylcrotonyl-CoA carboxylase complex (MCC) is an essential enzyme for the catabolism of the branched-chain amino acid leucine, which prevents the accumulation of toxic intermediaries, facilitates the generation of branched chain fatty acids and/or provides energy to the cell. The MCC complexes belong to a group of acyl-CoA carboxylases (ACCase) enzymes dependent of biotin. In this work, we have identified two ORFs (XAC0263 and XAC0264) encoding for the α and β subunits of an acyl-CoA carboxylase complex from Xanthomonas and demonstrated that this enzyme has MCC activity both in vitro and in vivo. We also found that this MCC complex is conserved in a group of pathogenic gram negative bacteria. The generation and analysis of an Xcc mutant strain deficient in MCC showed less canker lesions in the interaction with the host plant, suggesting that the expression of these proteins is necessary for Xcc fitness during infection.
In most of the organisms, including bacteria, archaea, fungi, algae, plants, and animals, the enzymatic complexes of biotin-dependent carboxylases catalyze fundamental metabolic reactions. These reactions are involved in the metabolism of fatty acids, carbohydrates, and amino acids, as well as in polyketide biosynthesis, urea utilization [1–4]. Biotin-dependent carboxylases contain three different components: the biotin carboxylase (BC), the biotin carboxyl carrier protein (BCCP) and the carboxyltransferase (CT). These components catalyze two separate hemi-reactions [5,6] (Fig 1A).
Phytopathogenic bacteria causing major plant diseases are frequently studied in terms of the role of bacterial protein secretion systems, bacterial effector proteins, pathogen associated molecular patterns (PAMPs), and pathogenicity factors in the triggering or overcoming of host defenses. However, infections caused by bacterial phytopathogens involve multiple adaptation processes, such as specific adherence of bacteria to host cells and tissues, and also adaptation of the bacterial metabolism to the nutrients availability and physical conditions existent in host tissues. In this work, we characterized a MCC complex from Xcc at the biochemical and genetic levels, and found that this enzyme, which may be involved in leucine catabolism, is expressed during infection and is necessary for survival into the citrus host tissue.