Figure 10. Cooperative invasion at membrane ruffles.

(A) Scheme explaining our quantification strategy for cooperative invasion. (B) HeLa cells were incubated with S.Tm^SopE(pGFP) at the indicated m.o.i. for 9 min, followed by washing, fixation and staining of actin and extracellular bacteria (anti-LPS antibody; staining before permeabilization). Left panel: quantification of the fraction of cells carrying ruffles. Middle panel: quantification of the number of “inside” and “outside” S.Tm^SopE(pGFP) per individual ruffle. Right Panel: number of invaded S.Tm^SopE(pGFP) per cell, calculated by multiplying the number of intracellular bacteria per ruffle with the fraction of ruffling cells. The black line indicates the estimated number of invaded S.Tm^SopE(pGFP) assuming independent invasion events, extrapolated from experimental data of ruffles with only one associated bacterium at the lowest m.o.i.. Each data point summarizes 4×150 cells for the analysis of cellular ruffling and 4×25 cells for inside and outside bacteria from 2 independent experiments. Error bars: standard deviation.