Journal: Infection and Immunity
Article Title: A Chemical Genetics Screen Reveals Influence of p38 Mitogen-Activated Protein Kinase and Autophagy on Phagosome Development and Intracellular Replication of Brucella neotomae in Macrophages
doi: 10.1128/IAI.00044-19
Figure Lengend Snippet: Chemical genetics screen identifies pathways that inhibit intracellular growth of B. neotomae. (A) Inhibitory effects of selected small molecules on intracellular replication in J774A.1 host cells at 48 h postinfection. Both B. neotomae (Bn) luminescence and CFU were measured in parallel. Data shown are means ± SD from at least two independent experiments, 3 replicates for luminescence data and 2 replicates for CFU data. Concentrations of the following tested compounds were in general above the IC50 (Table 1) and substantially below the CC50: arcyriaflavin A (3.6 μM), BX912 (3.6 μM), IKK16 (3.2 μM), I3M (3.6 μM), Ki20227 (2.0 μM), CYT387 (1.4 μM), Rhodblock6 (6.9 μM), SP600125 (10.2 μM), esomeprazole (10.2 μM), NS8593 (4.7 μM), and pinacidil (10.2 μM). (B) Inhibitory effects of selected small molecules on intracellular replication in primary murine bone marrow-derived macrophages (BMDM) at 48 h postinfection based on luminescence signals. Data shown are means ± SD from 4 replicates. Saponin (0.2%) and 0.3% DMSO in the absence of infection were used as positive and negative cytotoxicity controls, respectively. Significant suppression of luminescence was observed for all inhibitors in comparison with the DMSO control (P < 0.0001 by ANOVA and Dunnett’s post hoc multiple-comparison tests). (C) Effect of kinase target siRNA on intracellular growth of B. neotomae in J774A.1 host cells as assessed by luminescence. Data points represent means ± SD from at least three independent experiments. Significant suppression of luminescence (*, P < 0.001) was observed for siRNA knockdowns compared to the NTsi control. (D) Effect of siRNA knockdowns on mRNA expression assayed using same experimental protocol as the one described for panel C. Data points represent means ± SD from three independent samples and were normalized to the β-actin level within each sample and then to expression in untreated controls.
Article Snippet: IKK16 (IKK inhibitor VII) , APExBIO , B1586.
Techniques: Derivative Assay, Infection, Comparison, Control, Expressing
