Functional characterization of the type III secretion ATPase SsaN encoded by Salmonella pathogenicity island 2.
A type III secretion system (T3SS) is utilized by a large number of gram-negative bacteria to deliver effectors directly into the cytosol of eukaryotic host cells. One essential component of a T3SS is an ATPase that catalyzes the unfolding of proteins, which is followed by the translocation of effectors through an injectisome. Here we demonstrate a functional role of the ATPase SsaN, a component of Salmonella pathogenicity island 2 T3SS (T3SS-2) in Salmonella enterica serovar Typhimurium. SsaN hydrolyzed ATP in vitro and was essential for T3SS function and Salmonella virulence in vivo. Protein-protein interaction analyses revealed that SsaN interacted with SsaK and SsaQ to form the C ring complex. SsaN and its complex co-localized to the membrane fraction under T3SS-2 inducing conditions. In addition, SsaN bound to Salmonella pathogenicity island 2 (SPI-2) specific chaperones, including SsaE, SseA, SscA, and SscB that facilitated translocator/effector secretion. Using an in vitro chaperone release assay, we demonstrated that SsaN dissociated a chaperone-effector complex, SsaE and SseB, in an ATP-dependent manner. Effector release was dependent on a conserved arginine residue at position 192 of SsaN, and this was essential for its enzymatic activity. These results strongly suggest that the T3SS-2-associated ATPase SsaN contributes to T3SS-2 effector translocation efficiency.
Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.
Yukie
Y
Yoshida
Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.
Tsuyoshi
T
Miki
Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.
Sayaka
S
Ono
Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.
Takeshi
T
Haneda
Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.
Masahiro
M
Ito
Department of Microbiology, School of Pharmacy, Kitasato University, Tokyo, Japan.
Nobuhiko
N
Okada
D000251:Adenosine Triphosphatases / Q000235:genetics* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D058947:Bacterial Secretion Systems / Q000235:genetics*; D005260:Female; D015964:Gene Expression Regulation, Bacterial; D044404:Genomic Islands; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps; D012480:Salmonella Infections / Q000382:microbiology; D012486:Salmonella typhimurium / Q000201:enzymology / Q000235:genetics* / Q000472:pathogenicity*; D016415:Sequence Alignment; D014774:Virulence
D001426:Bacterial Proteins; D058947:Bacterial Secretion Systems; D018832:Molecular Chaperones; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases