FIG 3.

Entry assay identifies new components required for postentry processes during Brucella infection. (A) Representative images from the entry assay showing nuclei (DAPI) of HeLa cells and intracellular Brucella abortus (GFP) for control condition (mock) and cells treated with siRNAs against CDC42 or VPS35. HeLa cells were infected with B. abortus expressing GFP under a tetracycline-inducible system for 8 h (see Materials and Methods). Bars = 100 μm. (B) Scatter plot in double logarithmic scale showing infection scores measured for the entry assay (8 hpi) versus endpoint assay (48 hpi), normalized to the respective mock data set (Table S3). For the entry assay, cells containing single bacteria were considered infected, and the final readout is proportional to the median number of bacteria per infected cells. For the endpoint assay, only cells containing replicating bacteria were considered infected (Fig. 1 and Materials and Methods). Each data point corresponds to the average of all siRNAs or esiRNAs targeted against the gene of interest (n = 3). The straight fit (oblique line, r² = 0.763) indicates a globally high correlation between both assays. The blue region shows all points within 1 standard deviation (SD) of the fitted data. The genes falling out of this range are marked in red. For ease of visualization, only the averaged values over all RNAi products targeting a given gene are displayed.