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. Author manuscript; available in PMC: 2011 Jun 27.
Published in final edited form as: Mol Microbiol. 2010 Feb;75(4):924–941. doi: 10.1111/j.1365-2958.2009.07027.x

Fig. 3. Overexpression of PcrV or PcrV (Δ3–21).

Fig. 3

PcrV or the signal-sequenceless version PcrV(Δ3–21) were overexpressed in a wild-type strain background either in the presence or absence of calcium. The effect on the export of effectors was monitored using a lacZ reporter inserted in the exoS locus (A). Overexpression of PcrV or PcrV (Δ3–21) was confirmed by western blot on cell pellets of overexpressing strains (B, RpoA served as loading control). (C) Wild-type PcrV or a mutant lacking the export signal (Δ3–21) were expressed from a plasmid in a strain in which the chromosomal copy of pcrV had been deleted and in which expression of the type III secretion genes was uniformly up-regulated (ΔexsE). Where indicated, the strain also harbored a chromosomal copy of pcrG, which had been modified to express an internally VSV-G tagged version of the protein, PcrG(i3VG). Both input cell-lysates and output fractions after immunoprecipitation with the anti-VSV-G tag antibody were separated by SDS-PAGE and probed for the presence of PcrV or the VSV-G-tagged PcrG as indicated.