Figure 21.1.

Autophagy increases M. tuberculosis phagosome maturation and eliminates mycobacteria in macrophages. A. Autophagy reduces mycobacterial survival. RAW264.7 macrophages were infected with BCG for 1 h and incubated with or without starvation medium in the presence or absence of rapamycin, 3-methyladenine (3MA), or wortmannin (Wm) for 2 h. Cells were lysed to determine mycobacterial viability. B. and C. Beclin 1 is important for autophagic killing of mycobacteria. RAW264.7 cells were transfected with siRNAs against Beclin 1or scramble control. Protein knockdown was allowed to proceed for 48 h. Mycobacterial viability was determined as in A. Immunoblot analysis was performed to validate Beclin 1knockdown level using Actin as a loading control. D. and E. Autophagic induction enhances acidification and acquisition of a lysosomal protease by the mycobacterial phagosome. RAW264.7 cells were transfected with siRNAs against proteins of interest. Cells were infected with BCG and stained with LysoTracker Red or antibodies against cathepsin D. Quantitative analysis of percent colocalization was performed. Confocal images of cells transfected with siRNAs scramble control subjected to starvation treatment are shown as examples. Data, means ± SEM from three different experiments, **p ∷: 0.01, ^{p 2' 0.05. Fig. 21.1A is modified from Gutierrez et al. (2004). Figs. 21.1B and 21.C are modified from Delgado et al. (2008).