Identification of a VapBC toxin–antitoxin system in a thermophilic bacterium Thermus thermophilus HB27

Extremophiles volume 21, pages153–161(2017)Cite this article

Abstract

There are 12 putative toxin–antitoxin (TA) loci in the Thermus thermophilus HB27 genome, including four VapBC and three HicBA families. Expression of these seven putative toxin genes in Escherichia coli demonstrated that one putative VapC toxin TTC0125 and two putative HicA toxins, TTC1395 and TTC1705, inhibited cell growth, and co-expression with cognate antitoxin genes rescued growth, indicating that these genes function as TA loci. In vitro analysis with the purified TTC0125 and total RNA/mRNA from E. coli and T. thermophilus showed that TTC0125 has RNase activity to rRNA and mRNA; this activity was inhibited by the addition of the purified TTC0126. Translation inhibition assays showed that TTC0125 inhibited protein synthesis by degrading mRNA but not by inactivating ribosomes. Amino acid substitutions of 14 predicted catalytic and conserved residues in VapC toxins to Ala or Asp in TTC0125 indicated that nine residues are important for its in vivo toxin activity and in vitro RNase activity. These data demonstrate that TTC0125-TTC0126 functions as a VapBC TA module and causes growth inhibition by degrading free RNA. This is the first study to identify the function of TA systems in T. thermophilus.

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Abbreviations

Dox:

Doxycycline

DTT:

Dithiothreitol

IPTG:

Isopropyl-β-d-thiogalactopyranoside

PAGE:

Polyacrylamide gel electrophoresis

PBS:

Phosphate-buffered saline

PMSF:

Phenylmethanesulfonylfluoride

RNase:

Ribonuclease

TA:

Toxin–antitoxin

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Author information

Affiliations

  1. Faculty of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8572, Japan

    Yuqi Fan, Takayuki Hoshino & Akira Nakamura

Authors
  1. Yuqi Fan
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  2. Takayuki Hoshino
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  3. Akira Nakamura
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Corresponding author

Correspondence to Akira Nakamura.

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Communicated by S. Albers.

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Fan, Y., Hoshino, T. & Nakamura, A. Identification of a VapBC toxin–antitoxin system in a thermophilic bacterium Thermus thermophilus HB27. Extremophiles 21, 153–161 (2017). https://doi.org/10.1007/s00792-016-0891-1

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Keywords

  • Thermus thermophilus
  • Toxin–antitoxin system
  • VapBC
  • Ribonuclease