TY - JOUR
T1 - Vibrio choleraeMARTX toxin heterologous translocation of beta-lactamase and roles of individual effector domains on cytoskeleton dynamics
AU - Dolores, Jazel S.
AU - Agarwal, Shivani
AU - Egerer, Martina
AU - Satchell, Karla J.F.
N1 - Publisher Copyright:
© 2014 John Wiley & Sons Ltd.
PY - 2015/2/1
Y1 - 2015/2/1
N2 - Summary: The Vibrio choleraeMARTXVc toxin delivers three effector domains to eukaryotic cells. To study toxin delivery and function of individual domains, the rtxA gene was modified to encode toxin with an in-frame beta-lactamase (Bla) fusion. The hybrid RtxA::Bla toxin was Type I secreted from bacteria; and then Bla was translocated into eukaryotic cells and delivered by autoprocessing, demonstrating that the MARTXVc toxin is capable of heterologous protein transfer. Strains that produce hybrid RtxA::Bla toxins that carry one effector domain in addition to Bla were found to more efficiently translocate Bla. In cell biological assays, the actin cross-linking domain (ACD) and Rho-inactivation domain (RID) are found to cross-link actin and inactivate RhoA, respectively, when other effector domains are absent, with toxin autoprocessing required for high efficiency. The previously unstudied alpha-beta hydrolase domain (ABH) is shown here to activate CDC42, although the effect is ameliorated when RID is also present. Despite all effector domains acting on cytoskeleton assembly, the ACD was sufficient to rapidly inhibit macrophage phagocytosis. Both the ACD and RID independently disrupted polarized epithelial tight junction integrity. The sufficiency of ACD but strong selection for retention of RID and ABH suggests these two domains may primarily function by modulating cell signaling.
AB - Summary: The Vibrio choleraeMARTXVc toxin delivers three effector domains to eukaryotic cells. To study toxin delivery and function of individual domains, the rtxA gene was modified to encode toxin with an in-frame beta-lactamase (Bla) fusion. The hybrid RtxA::Bla toxin was Type I secreted from bacteria; and then Bla was translocated into eukaryotic cells and delivered by autoprocessing, demonstrating that the MARTXVc toxin is capable of heterologous protein transfer. Strains that produce hybrid RtxA::Bla toxins that carry one effector domain in addition to Bla were found to more efficiently translocate Bla. In cell biological assays, the actin cross-linking domain (ACD) and Rho-inactivation domain (RID) are found to cross-link actin and inactivate RhoA, respectively, when other effector domains are absent, with toxin autoprocessing required for high efficiency. The previously unstudied alpha-beta hydrolase domain (ABH) is shown here to activate CDC42, although the effect is ameliorated when RID is also present. Despite all effector domains acting on cytoskeleton assembly, the ACD was sufficient to rapidly inhibit macrophage phagocytosis. Both the ACD and RID independently disrupted polarized epithelial tight junction integrity. The sufficiency of ACD but strong selection for retention of RID and ABH suggests these two domains may primarily function by modulating cell signaling.
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U2 - 10.1111/mmi.12879
DO - 10.1111/mmi.12879
M3 - Article
C2 - 25427654
AN - SCOPUS:84922602285
SN - 0950-382X
VL - 95
SP - 590
EP - 604
JO - Molecular Microbiology
JF - Molecular Microbiology
IS - 4
ER -