Highlight:Trehalose is a double-edged sword for both partners in the citrus–Xanthomonas interaction, as it is necessary for bacterial survival but also triggers citrus defence responses.

BackgroundSeveral bacterial plant pathogens colonize their hosts through the secretion of effector proteins by a Type III protein secretion system (T3SS). The role of T3SS in bacterial pathogenesis is well established but whether this system is involved in multicellular processes, such as bacterial biofilm formation has not been elucidated. Here, the phytopathogen Xanthomonas citri subsp. citri (X. citri) was used as a model to gain further insights about the role of the T3SS in biofilm formation.ResultsThe capacity of biofilm formation of different X. citri T3SS mutants was compared to the wild type strain and it was observed that this secretion system was necessary for this process. Moreover, the T3SS mutants adhered proficiently to leaf surfaces but were impaired in leaf-associated growth. A proteomic study of biofilm cells showed that the lack of the T3SS causes changes in the expression of proteins involved in metabolic processes, energy generation, exopolysaccharide (EPS) production and bacterial motility as well as outer membrane proteins. Furthermore, EPS production and bacterial motility were also altered in the T3SS mutants.ConclusionsOur results indicate a novel role for T3SS in X. citri in the modulation of biofilm formation. Since this process increases X. citri virulence, this study reveals new functions of T3SS in pathogenesis.

Cerulenin is a fungal toxin that inhibits both eukaryotic and prokaryotic ketoacyl‐acyl carrier protein synthases or condensing enzymes. It has been used experimentally to treat cancer and obesity, and is a potent inhibitor of bacterial growth. Understanding the molecular mechanisms of resistance to cerulenin and similar compounds is thus highly relevant for human health. We have previously described a Bacillus subtilis cerulenin‐resistant strain, expressing a point‐mutated condensing enzyme FabF (FabF[I108F]) (i.e. FabF with isoleucine 108 substituted by phenylalanine). We now report the crystal structures of wild‐type FabF from B. subtilis, both alone and in complex with cerulenin, as well as of the FabF[I108F] mutant protein. The three‐dimensional structure of FabF[I108F] constitutes the first atomic model of a condensing enzyme that remains active in the presence of the inhibitor. Soaking the mycotoxin into preformed wild‐type FabF crystals allowed for noncovalent binding into its specific pocket within the FabF core. Interestingly, only co‐crystallization experiments allowed us to trap the covalent complex. Our structure shows that the covalent bond between Cys163 and cerulenin, in contrast to that previously proposed, implicates carbon C3 of the inhibitor. The similarities between Escherichia coli and B. subtilis FabF structures did not explain the reported inability of ecFabF[I108F] (i.e. FabF from Escherichia coli with isoleucine 108 substituted by phenylalanine) to elongate medium and long‐chain acyl‐ACPs. We now demonstrate that the E. coli modified enzyme efficiently catalyzes the synthesis of medium and long‐chain ketoacyl‐ACPs. We also characterized another cerulenin‐insensitive form of FabF, conferring a different phenotype in B. subtilis. The structural, biochemical and physiological data presented, shed light on the mechanisms of FabF catalysis and resistance to cerulenin. Database Crystallographic data (including atomic coordinates and structure factors) have been deposited in the Protein Data Bank under accession codes http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4LS5, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4LS6, http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4LS7 and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4LS8.