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Steps for Shigella Gatekeeper Protein MxiC Function in Hierarchical Type III Secretion Regulation*

A. D. Roehrich,E. Bordignon,S. Mode,D. Shen,Xia Liu,Maria Pain,I. Murillo,Isabel Martinez-Argudo,Isabel Martinez-Argudo,R. Sessions,A. Blocker

Published 2016 in Journal of Biological Chemistry

ABSTRACT

Type III secretion systems are complex nanomachines used for injection of proteins from Gram-negative bacteria into eukaryotic cells. Although they are assembled when the environmental conditions are appropriate, they only start secreting upon contact with a host cell. Secretion is hierarchical. First, the pore-forming translocators are released. Second, effector proteins are injected. Hierarchy between these protein classes is mediated by a conserved gatekeeper protein, MxiC, in Shigella. As its molecular mechanism of action is still poorly understood, we used its structure to guide site-directed mutagenesis and to dissect its function. We identified mutants predominantly affecting all known features of MxiC regulation as follows: secretion of translocators, MxiC and/or effectors. Using molecular genetics, we then mapped at which point in the regulatory cascade the mutants were affected. Analysis of some of these mutants led us to a set of electron paramagnetic resonance experiments that provide evidence that MxiC interacts directly with IpaD. We suggest how this interaction regulates a switch in its conformation that is key to its functions.

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