Agrobacterium tumefaciens is a facultative phytopathogen and causative agent of crown gall disease, capable of attaching to surfaces via a polarly localized unipolar polysaccharide (UPP) adhesin. The UPP is composed of two distinct forms, one characterized by N-acetylglucosamine residues (UPPGlcN) and the other by N-acetylgalactosamine residues (UPPGalN). Two genes with presumptive roles in dTDP-L-rhamnose biosynthesis, rfbA and rfbD, were identified in a transposon screen to be involved specifically in UPPGalN biosynthesis. Examination of independent rfbA and rfbD mutations validates the transposon mutant phenotypes, with UPPGalN specific defects observed as well as general attachment defects and UPP-mediated cellular aggregation across all rfb mutants. Additionally, despite retaining flagella, mutations in the rfb genes impart flagellar motility defects. Suppressor mutations that rescue the non-motile {Delta}rfbD phenotype disrupt the remaining rfb genes (rfbA, rfbB, and rfbC), the phosphoglucomutase exoC and a putative glycosyl transferase ATU-RS21610. Single deletion mutants in the dTDP-L-rhamnose pathway also have hallmarks of compromised outer membrane integrity, such as increased sensitivity to high-molecular weight antibiotics and increased expression of target genes for the widely conserved ChvG-ChvI two component system outer membrane stress response. Thus, defects in the dTDP-L-rhamnose pathway cause multiple cell surface deficiencies, resulting in outer membrane stress, impaired flagellar motility, defective UPP production, and irregular adhesion.
Reynolds, I. P., Westin, I., Wood, C., Fuqua, C.
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