The unipolar Shigella surface protein IcsA is targeted directly to the bacterial old pole: IcsP cleavage of IcsA occurs over the entire bacterial surface

Shigella flexneri is an intracellular pathogen that is able to move within the cytoplasm of infected cells by the continual assembly of actin onto one pole of the bacterium. IcsA, an outer membrane protein, is localized to th e old pole of the bacterium and is both necessary and sufficient for actin assembly. IcsA is slowly cleaved from the bacterial surface by the protease IcsP (SopA). Absence of IcsP leads to an alteration in the distribution of surface IcsA, such that the polar cap is maintained and some IcsA is distr ibuted along the lateral walls of the bacillus, The mechanism of unipolar l ocalization of IcsA and the role of IcsP in its unipolar localization are i ncompletely understood. Here, we demonstrate that cleavage of IcsA occurs e xclusively in the outer membrane and that IcsP is localized to the outer me mbrane. In addition, we show that IcsA at the old pole is susceptible to cl eavage by IcsP and that native IcsP is active at the pole. Taken together, these data indicate that IcsP cleaves IcsA over the entire bacterial surfac e, Finally, we show that, immediately after induction from a tightly regula ted promoter, IcsA is expressed exclusively at the old pole in both the ics P(-) icsA(-) and the icsA- background. These data demonstrate that unipolar localization of IcsA results from its direct targeting to the pole, follow ed by its diffusion laterally in the outer membrane.