Actin Re-Organization Induced by Chlamydia trachomatis Serovar D - Evidence for a Critical Role of the Effector Protein CT166 Targeting Rac
Figure 9
C. trachomatis D induced cytopathic effects are diminished by over-expression of a constitutive active Rac1 mutant, or by CNF1.
HeLa cells were transiently transfected with the pEGFP-C1 vector encoding for human Rac1-G12V, or with the empty control vector. After 24 h the cells were either mock infected or infected with C. trachomatis D (MOI 100). Depicted is the rhodamine-phalloidin staining of the actin cytoskeleton and the fluorescence of GFP-fusion protein expressing cells at 2 h post infection (A). GFP-positive cells (cell diameter <30 µm) were analyzed for cell rounding (* indicates significant differences comparing mock-infected with C. trachomatis D-infected control cells, p<0.05; ** indicates significant differences comparing infected control cells with infected Rac1-G12V expressing cells, p<0.005) (B). The average cell diameter of GFP-positive cells was determined (* indicates significant differences comparing mock-infected with C. trachomatis D-infected control cells, p<0.05; ** indicates significant differences comparing infected control cells with infected Rac1-G12V expressing cells, p<0.005) (C). HeLa cells were treated with 0.1 µg/ml CNF1 from E. coli for 6 h, or with buffer. Subsequently, cells were either mock infected or infected with C. trachomatis D (MOI 100). At 2 h post infection the actin cytoskeleton was stained with rhodamine-phalloidin (not shown) and rounded cells (cell diameter <30 µm) were quantified and given as ratio in percent (** indicates significant differences as compared to the corresponding mock-infected cells, p<0.05; * indicates significant differences comparing infected control cells with infected CNF1-treated cells, p<0.05) (D). The average cell diameter was determined (*/** indicates significant differences as compared to the corresponding mock-infected cells, p<0.05/p<0.005; ** indicates significant differences comparing infected control cells with infected CNF1-treated cells, p<0.05) (E). Depicted is the mean ± SD of three independent experiments.