title: A VapBC toxin-antitoxin module is a posttranscriptional regulator of metabolic flux in mycobacteria.
abstract: The largest family of toxin-antitoxin (TA) modules are encoded by the vapBC operons, but their roles in bacterial physiology remain enigmatic. Microarray analysis in Mycobacterium smegmatis overexpressing VapC/VapBC revealed a high percentage of downregulated genes with annotated roles in carbon transport and metabolism, suggesting that VapC was targeting specific metabolic mRNA transcripts. To validate this hypothesis, purified VapC was used to identify the RNA cleavage site in vitro. VapC had RNase activity that was sequence specific, cleaving single-stranded RNA substrates at AUAU and AUAA in vitro and in vivo (viz., MSMEG_2121 to MSMEG_2124). A bioinformatic analysis of these regions suggested that an RNA hairpin 3' of the AUA(U/A) motif is also required for efficient cleavage. VapC-mediated regulation in vivo was demonstrated by showing that MSMEG_2124 (dhaF) and MSMEG_2121 (dhaM) were upregulated in a ΔvapBC mutant growing on glycerol. The ΔvapBC mutant had a specific rate of glycerol consumption that was 2.4-fold higher than that of the wild type during exponential growth. This increased rate of glycerol consumption was not used for generating bacterial biomass, suggesting that metabolism by the ΔvapBC mutant was uncoupled from growth. These data suggest a model in which VapC regulates the rate of glycerol utilization to match the anabolic demands of the cell, allowing for fine-tuning of the catabolic rate at a posttranscriptional level.
authors.affiliation: Department of Biological Sciences, University of Waikato, Hamilton, New Zealand.
authors.forename: Joanna L
authors.initials: JL
authors.lastname: McKenzie
authors.forename: Jennifer
authors.initials: J
authors.lastname: Robson
authors.forename: Michael
authors.initials: M
authors.lastname: Berney
authors.forename: Tony C
authors.initials: TC
authors.lastname: Smith
authors.forename: Alaine
authors.initials: A
authors.lastname: Ruthe
authors.forename: Paul P
authors.initials: PP
authors.lastname: Gardner
authors.forename: Vickery L
authors.initials: VL
authors.lastname: Arcus
authors.forename: Gregory M
authors.initials: GM
authors.lastname: Cook
mesh_terms: D000992:Antitoxins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001427:Bacterial Toxins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000502:physiology; D002244:Carbon / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D004734:Energy Metabolism / Q000502:physiology*; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D020868:Gene Silencing; D005990:Glycerol / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D020102:Mycobacterium smegmatis / Q000235:genetics / Q000378:metabolism*; D040081:Protein Array Analysis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism
chemical_list: D000992:Antitoxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D004268:DNA-Binding Proteins; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; C077517:VapB protein, Bacteria; D002244:Carbon; D005990:Glycerol