Journal: PLoS Pathogens
Article Title: PknG senses amino acid availability to control metabolism and virulence of Mycobacterium tuberculosis
Figure Lengend Snippet: garA is required for growth of M . tuberculosis in vitro , survival in macrophages, and virulence in mice. ( A ) M . tuberculosis lacking garA was unable to grow on standard 7H10 medium unless supplemented with asparagine. Plasmid-borne garA restored the defect, but variants of garA with mutations at threonine 21 in the phosphorylation motif (ETTS) gave only partial complementation. Strains were grown in Middlebrook 7H9 plus 30 mM asparagine then washed and diluted in standard 7H9 and spotted onto standard 7H10 with or without 10 mM asparagine. Photographs are representative of at least 3 independent experiments. ( B ) M . tuberculosis lacking garA (red squares) had a defect in growth and survival in differentiated THP-1 cells compared to parental M . tuberculosis H37Rv (black circles). Re-introduction of GarA (black triangles) or variants of GarA lacking a single phosphorylation site (grey crosses and squares) restored growth but variant GarA lacking both phosphorylation sites (green triangles) did not. Data points show the mean and standard deviation from four replicates and are representative of two independent experiments. ( C ) M . tuberculosis lacking garA was avirulent in mice as it was eliminated from the lungs. BALB/C mice were infected intranasally with 10 5 bacilli and bacterial burden was measured on days 1, 7, 21 and 28. Data points show the bacterial burden in individual animals. The bacterial burden of mice infected with Δ garA Mt (red squares) was significantly lower than those infected with M . tuberculosis H37Rv (black circles), or complemented Δ garA Mt (black triangles) at all time points from day 7 (p
Article Snippet: Bacterial strains, media, and culture M . tuberculosis H37Rv and M . smegmatis mc2 155 were routinely cultured on Middlebrook 7H10 agar (Oxoid) with 10% ADN (0.5% bovine serum albumin, 0.2% dextrose, 0.085% NaCl) and Middlebrook 7H9 medium (Oxoid) with 10% ADN and 0.05% Tween 80.
Techniques: In Vitro, Mouse Assay, Plasmid Preparation, Variant Assay, Standard Deviation, Infection