The Mxi-Spa type III secretory pathway of Shigella flexneri requires an outer membrane lipoprotein, MxiM, for invasin translocation

Invasion of epithelial cells by Shigella flexneri is mediated by a set of t ranslocated bacterial invasins, the Ipa proteins, and its dedicated type II I secretion system, called Mxi-Spa. We show here that mxiM, part of the mri -spa locus in the S. flexneri virulence plasmid, encodes an indispensable t ype III secretion apparatus component, required for both Ipa translocation and tissue culture cell invasion. We demonstrated that mature MxiM, first i dentified as a putative lipoprotein, is lipidated in vivo. Consistent with features of known lipoproteins, MxiM (i) can be labeled with [H-3]palmitate and [2-H-3]glycerol, (ii) is associated with the cell envelope, (iii) is s ecreted independently of the type III pathway, and (iv) requires an intact lipoprotein modification and processing site for full activity. The lipidat ed form of MxiM was detected primarily in the outer membrane, where it esta blishes a peripheral association with the inner leaflet. Through analysis o f subcellular Ipa distribution in a mxiM null mutant background, MxiM was f ound to be required for the assembly and/or function of outer, but not inne r, membrane regions of Mxi-Spa. This function probably requires interaction s with other Mxi-Spa subunits within the periplasmic space. We discuss impl ications of these findings with respect to the function of MxiM and the str ucture of Mxi-Spa as a whole.