Figure 4. Modulation of HspB translocation by HcpG and HcpC during H. pylori infection.

(a) ELISA performed to determine the specificity of HcpC-HspB interaction, in vitro. (b–e) FACS analyses of HspB translocation in single- and double hcp gene mutants and in the parent HpG27MA WT strain (b) 3, (c) 6, (d) 12, and (e) 24 h after infection of serum-starved confluent AGS cell cultures. Overlapping histograms of mean geometric intensity from individual infections were generated using the WINMDI software program (version 2.9; Joseph Trotter, Scripps Research Institute, LA Jolla, CA). x-axis, α-HspB fluorescence; y-axis, number of events measured for each sample (10⁴ cells/sample). A shift of fluorescence to the left indicates reduced intensity. The FACS parameters are listed in supplementary methods available in File S1. Inset, bar graphs showing the geometric mean intensity of α-HspB fluorescence at each time point after infection with mutant and WT strains, respectively. The strains are color-coded as follows: Black, WT; blue, ΔhcpC; red, ΔhcpG; and green, ΔhcpG,ΔhcpC. The results show the averages from three experiments (± SD). (f). Model of observed functional interactions of HcpC, HcpG, and HspB translocation depending on the temporal context of the infection.