Defects in flagellin glycosylation affect the virulence of Pseudomonas syringae pv. tabaci 6605.
Flagellar motility and its glycosylation are indispensable for the virulence of Pseudomonas syringae pv. tabaci 6605. Six serine residues of the flagellin protein at positions 143, 164, 176, 183, 193 and 201 are glycosylated, and the glycan structure at 201 was determined to consist of a trisaccharide of two L-rhamnosyl residues and a modified 4-amino-4,6-dideoxyglucosyl (viosamine) residue. To investigate the glycan structures attached to the other serine residues and to identify the glycans important for virulence, Ser/Ala-substituted mutants were generated. Six mutant strains that each retained a single glycosylated serine residue were generated by replacing five of the six serine residues with alanine residues. MALDI-TOF mass analysis of flagellin proteins revealed that the major component of each glycan was a trisaccharide basically similar to that at position 201, but with heterogeneity in glycoform distribution. Swarming motility and amounts of acylhomoserine lactones (AHLs) as quorum-sensing signal molecules were significantly reduced, especially in the S143-5S/A, S164-5S/A and S201-5S/A mutants, whereas tolerance to antibiotics was increased in these three mutants. All the mutants showed lower ability to cause disease on host tobacco plants. These results supported our previous finding that glycosylation of the most externally located sites on the surface of the flagellin molecule, such as S176 and S183, is required for virulence in P. syringae pv. tabaci 6605. Furthermore, it is speculated that flagellum-dependent motility might be correlated with quorum sensing and antibiotic resistance.
Taguchi
Fumiko
F
Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka 1-1-1, Okayama 700-8530 Japan.
Yamamoto
Masanobu
M
National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan.
Ohnishi-Kameyama
Mayumi
M
National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan.
Iwaki
Masako
M
Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka 1-1-1, Okayama 700-8530 Japan.
Yoshida
Mitsuru
M
National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan.
Ishii
Tadashi
T
Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687, Japan.
Konishi
Tomoyuki
T
Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki 305-8687, Japan.
Ichinose
Yuki
Y
Graduate School of Natural Science and Technology, Okayama University, Tsushima-naka 1-1-1, Okayama 700-8530 Japan.
D054742:Acyl-Butyrolactones / Q000378:metabolism; D019943:Amino Acid Substitution; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D024881:Drug Resistance, Bacterial; D005408:Flagellin / Q000737:chemistry*; D006031:Glycosylation; D008826:Microbial Sensitivity Tests; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011135:Polysaccharides, Bacterial / Q000096:biosynthesis; D044224:Pseudomonas syringae / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D053038:Quorum Sensing; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014026:Nicotiana / Q000382:microbiology; D014774:Virulence
D054742:Acyl-Butyrolactones; D000900:Anti-Bacterial Agents; D011135:Polysaccharides, Bacterial; D005408:Flagellin