Identification and characterization of a chromosomal virulence gene, vacJ, required for intercellular spreading of Shigella flexneri.
Intercellular spreading of shigellae is a prerequisite for shigellosis, although the molecular mechanisms underlying the phenomenon are still largely obscure. To elucidate some of these mechanisms, we performed random Tn10 insertion mutagenesis in Shigella flexneri YSH6000T and found a chromosomal locus in the NotI-J segment responsible for bacterial spreading. The locus affected in the mutant, designated vacJ, was neither involved in the invasion of epithelial cells nor in intracellular movement, but was required for intercellular spread. The vacJ mutant was capable of forming bacterium-containing membranous protrusions within the infected cell, but had diminished ability to move from the protrusions into the cytoplasm of the adjacent epithelial cells. Cloning and sequencing of the vacJ region indicated that the vacJ gene encoded a 28.0 kDa protein possessing a signal peptide at the N-terminus, which contained the motif characteristic of lipoproteins. The analysis of the vacJ product indicated that VacJ was exposed on the bacterial surface. The vacJ gene was distributed among shigellae and enteroinvasive Escherichia coli, and the constructed vacJ mutants failed to spread intercellularly, indicating that vacJ is a chromosomal gene essential for the pathogenicity of shigellae.
Suzuki
T
T
Department of Bacteriology, University of Tokyo, Japan.
Murai
T
T
Fukuda
I
I
Tobe
T
T
Yoshikawa
M
M
Sasakawa
C
C
D000595:Amino Acid Sequence; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D001426:Bacterial Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D002874:Chromosome Mapping; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004251:DNA Transposable Elements / Q000235:genetics; D004269:DNA, Bacterial / Q000302:isolation & purification; D004848:Epithelium; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016254:Mutagenesis, Insertional; D017422:Sequence Analysis, DNA; D012763:Shigella flexneri / Q000235:genetics* / Q000472:pathogenicity*; D014774:Virulence
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; C086588:VacJ protein, Shigella flexneri