FIG 1.

The pks island-positive nonhemolytic mutant E. coli 536-HDM requires exogenous UvrY to induce a cytopathic effect in HeLa cells. (A) HeLa cells were either not infected or infected with the indicated E. coli strains to a multiplicity of infection (MOI) of 200. After 4 hours of infection, HeLa cells were washed to remove bacteria and further cultivated. At 72 h postinfection, cells were washed and Giemsa stained. Scale bars, 50 μm. The uvrY plasmid pCA9505 (50) was used to complement the uvrY deletion in E. coli strain 536-HDM. As a control, E. coli strain 536-HDM was transformed with the uvrY-negative variant of plasmid pCA9505, namely, pCA9505-MluI (90). (B) N-Myristoyl-d-asparagine (C14-asparagine) quantification (mean values ± SEM) in bacterial cultures of E. coli strains M1/5, 536, 536-HDM, 536-HDM (pCA9505), and 536-HDM (pCA9505-MluI) by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The data presented in the graph were obtained from three biological replicates. ****, P < 0.0001; ***, P < 0.001; **, P < 0.01; 1-way ANOVA with Bonferroni correction. (C) DNA cross-link formation of plasmid DNA exposed to strains M1/5, 536, 536-HDM, 536-HDM (pCA9505), and 536-HDM (pCA9505-MluI) was visualized after migration under alkaline denaturing conditions. M, DNA size marker (1 kb plus DNA ladder, Invitrogen). This image is representative of three independent experiments.