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. 2017 Jul 25;8(4):e01047-17. doi: 10.1128/mBio.01047-17

FIG 8 .

FIG 8 

Expression of the human ppGpp hydrolase enzyme Mesh1 prevents ppGpp accumulation in yeast and allows normal growth. (a) Production of ppGpp following DOX induction in strain BY4741-112-135 (135; ppGpp synthetase) was prevented in a strain coexpressing MESH1 (BY4741-112-142; 142). Cultures were induced with DOX (5 μg/ml) and incubated for 6 h prior to analysis of intracellular nucleotide composition by LC-UV. Nucleotide abundances, normalized to picomoles per milligram (dry weight) of cells, are the averages of triplicate cultures (± the standard deviation [SD]). Strains BY4741-112-134 (134; empty vector) and BY4741-112-139 (139; Mesh1) are shown as controls. (b) Serial dilution plate growth assay of the strains shown in panel a when grown on YNB with 0.5% glucose, with (5 DOX) or without (0 DOX) 5 μg/ml DOX. Growth of strain 135 (ppGpp synthetase) was inhibited, while growth of 142 (ppGpp synthetase plus Mesh1 ppGpp hydrolase) was not. (c) DOX-induced ppGpp synthesis lengthens both doubling time and lag time in microtiter plate cultures of strain 135 (ppGpp synthetase), but coexpression of MESH1 restored these growth characteristics to levels observed in the control strains. The data are averages (± SD) of four replicates grown in YNB with 0.5% glucose, with (+) or without (−) 5 μg/ml DOX.