Dataset Information

Toxin YafQ Reduces Escherichia coli Growth at Low Temperatures.

ABSTRACT: Toxin/antitoxin (TA) systems reduce metabolism under stress; for example, toxin YafQ of the YafQ/DinJ Escherichia coli TA system reduces growth by cleaving transcripts with in-frame 5'-AAA-G/A-3' sites, and antitoxin DinJ is a global regulator that represses its locus as well as controls levels of the stationary sigma factor RpoS. Here we investigated the influence on cell growth at various temperatures and found that deletion of the antitoxin gene, dinJ, resulted in both reduced metabolism and slower growth at 18°C but not at 37°C. The reduction in growth could be complemented by producing DinJ from a plasmid. Using a transposon screen to reverse the effect of the absence of DinJ, two mutations were found that inactivated the toxin YafQ; hence, the toxin caused the slower growth only at low temperatures rather than DinJ acting as a global regulator. Corroborating this result, a clean deletion of yafQ in the ?dinJ ?KmR strain restored both metabolism and growth at 18°C. In addition, production of YafQ was more toxic at 18°C compared to 37°C. Furthermore, by overproducing all the E. coli proteins, the global transcription repressor Mlc was found that counteracts YafQ toxicity only at 18°C. Therefore, YafQ is more effective at reducing metabolism at low temperatures, and Mlc is its putative target.

SUBMITTER: Zhao Y

PROVIDER: S-EPMC4996492 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Toxin YafQ Reduces Escherichia coli Growth at Low Temperatures.

Zhao Yueju Y McAnulty Michael J MJ Wood Thomas K TK

PloS one 20160824 8

Toxin/antitoxin (TA) systems reduce metabolism under stress; for example, toxin YafQ of the YafQ/DinJ Escherichia coli TA system reduces growth by cleaving transcripts with in-frame 5'-AAA-G/A-3' sites, and antitoxin DinJ is a global regulator that represses its locus as well as controls levels of the stationary sigma factor RpoS. Here we investigated the influence on cell growth at various temperatures and found that deletion of the antitoxin gene, dinJ, resulted in both reduced metabolism and ...[more]

PMID: 27557125

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Project description:We have shown the quorum-sensing signals acylhomoserine lactones (AHLs), autoinducer-2 (AI-2), and indole influence the biofilm formation of Escherichia coli. Here, we investigate how the environment, i.e., temperature, affects indole and AI-2 signaling in E. coli. We show in biofilms that indole addition leads to more extensive differential gene expression at 30ï°C (186 genes) than at 37ï°C (59 genes), that indole reduces biofilm formation (without affecting growth) more significantly at 25ï°C and 30ï°C than at 37ï°C, and that the effect is associated with the quorum-sensing protein SdiA. The addition of indole at 30ï°C compared to 37ï°C most significantly repressed genes involved in uridine monophosphate (UMP) biosynthesis (carAB, pyrLBI, pyrC, pyrD, pyrF, and upp) and uracil transport (uraA). These uracil-related genes are also repressed at 30ï°C by SdiA, which confirms SdiA is involved in indole signaling. Also, compared to 37ï°C, indole more significantly decreased flagella-related qseB, flhD, and fliA promoter activity, enhanced antibiotic resistance, and inhibited cell division at 30ï°C. In contrast to indole and SdiA, the addition of (S)-4,5-dihydroxy-2,3-pentanedione (the AI-2 precursor) leads to more extensive differential gene expression at 37ï°C (63 genes) than at 30ï°C (11 genes), and, rather than repressing UMP synthesis genes, AI-2 induces them at 37ï°C (but not at 30ï°C). Also, the addition of AI-2 induces the transcription of virulence genes in enterohemorrhagic E. coli O157:H7 at 37oC but not at 30oC. Hence, cell signals cause diverse responses at different temperatures, and indole- and AI-2-based signaling are intertwined. Experiment Overall Design: We performed seven sets of microarray experiments in LB medium (Table 2): (i) biofilm cells of the BW25113 wild-type strain grown for 7 h at 30Â°C vs. 37Â°C to determine the effect of temperature on E. coli biofilm formation (initial absorbance of 0.05), (ii) biofilm cells of the tnaA mutant grown for 7 h with 1 mM indole vs. no indole at 30Â°C to determine the effect of temperature on indole signaling in E. coli (initial absorbance of 0.05), (iii) biofilm cells of the tnaA mutant grown for 7 h with 1 mM indole vs. no indole at 37Â°C to determine the effect of temperature on indole signaling in E. coli, (iv) suspension cells of the wild-type strain vs. the sdiA mutant at an absorbance of 4.0 at 600 nm at 30Â°C (since production of indole takes place primarily in the stationary phase) to discern the genes that SdiA controls (initial absorbance of 0.05), (v) biofilm cells of the sdiA mutant grown for 7 h with 1 mM indole vs. no indole at 30Â°C to determine gene expression in response to indole in the absence of SdiA, (vi) suspension cells of the luxS mutant with 100 micro M AI-2 vs. no AI-2 at 30Â°C for 3 h to determine the effect of temperature on AI-2 signaling in E. coli (initial absorbance of 0.5), and (vii) suspension cells of the luxS mutant with 100 micro M AI-2 vs. no AI-2 at 37Â°C for 3 h to determine the effect of temperature on AI-2 signaling in E. coli. Ten g of glass wool (Corning Glass Works, Corning, N.Y.) was used to form large amounts of biofilms (Ren et al., 2004a) in 250 mL LB in 1 L Erlenmeyer shake flasks. RNA was isolated from the suspension and biofilm cells as described previously (Ren et al., 2004b).

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Toxin YafQ Reduces Escherichia coli Growth at Low Temperatures.

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Toxin YafQ Reduces Escherichia coli Growth at Low Temperatures.

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