ABSTRACT
Type III secretion systems (TTSS) are essential virulence determinants of many gram-negative bacteria and serve, upon physical contact with target cells, to translocate bacterial proteins directly across eukaryotic cell membranes. The
Shigella
TTSS is encoded by the
mxi/spa
loci located on its virulence plasmid. By electron microscopy secretons are visualized as tripartite with an external needle, a transmembrane domain, and a cytoplasmic bulb. In the present study, we generated a
Shigella spa32
mutant and studied its phenotype. The
spa32
gene shows low sequence homology to
Salmonella
TTSS1
invJ/spaN
and to flagellar
fliK
. The
spa32
mutant, like the wild-type strain, secreted the Ipas and IpgD, which are normally secreted via the TTSS, at low levels into the growth medium. However, unlike the wild-type strain, the
spa32
mutant could neither be induced to secrete the Ipas and IpgD instantaneously upon addition of Congo red nor penetrate HeLa cells in vitro. Additionally, the Spa32 protein is secreted in large amounts by the TTSS during exponential growth but not upon Congo red induction. Interestingly, electron microscopy analysis of the
spa32
mutant revealed that the needle of its secretons were up to 10 times longer than those of the wild type. In addition, in the absence of induction, the
spa32
mutant secreted normal levels of MxiI but a large excess of MxiH. Taken together, our data indicate that the
spa32
mutant presents a novel phenotype and that the primary defect of the mutant may be its inability to regulate or control secretion of MxiH.
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