The transjugation machinery of Thermus thermophilus: Identification of TdtA, an ATPase involved in DNA donation

PLoS Genet. 2017 Mar 10;13(3):e1006669. doi: 10.1371/journal.pgen.1006669. eCollection 2017 Mar.

Abstract

In addition to natural competence, some Thermus thermophilus strains show a high rate of DNA transfer via direct cell-to-cell contact. The process is bidirectional and follows a two-step model where the donor cell actively pushes out DNA and the recipient cell employs the natural competence system to take up the DNA, in a hybrid transformation-dependent conjugation process (transjugation). While the DNA uptake machinery is well known as in other bacterial species that undergo transformation, the pushing step of transjugation remains to be characterized. Here we have searched for hypothetical DNA translocases putatively involved in the pushing step of transjugation. Among candidates encoded by T. thermophilus HB27, the TdtA protein was found to be required for DNA pushing but not for DNA pulling during transjugation, without affecting other cellular processes. Purified TdtA shows ATPase activity and oligomerizes as hexamers with a central opening that can accommodate double-stranded DNA. The tdtA gene was found to belong to a mobile 14 kbp-long DNA element inserted within the 3' end of a tRNA gene, flanked by 47 bp direct repeats. The insertion also encoded a homolog of bacteriophage site-specific recombinases and actively self-excised from the chromosome at high frequency to form an apparently non-replicative circular form. The insertion also encoded a type II restriction endonuclease and a NurA-like nuclease, whose activities were required for efficient transjugation. All these data support that TdtA belongs to a new type of Integrative and Conjugative Element which promotes the generalized and efficient transfer of genetic traits that could facilitate its co-selection among bacterial populations.

MeSH terms

  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Computational Biology
  • DNA Restriction Enzymes / metabolism
  • DNA, Bacterial / genetics*
  • Escherichia coli / metabolism
  • Microscopy, Electron
  • Mutation
  • Phenotype
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Thermus thermophilus / genetics
  • Thermus thermophilus / metabolism*
  • Transformation, Bacterial

Substances

  • Bacterial Proteins
  • DNA, Bacterial
  • Recombinant Proteins
  • DNA Restriction Enzymes
  • Adenosine Triphosphatases

Grants and funding

This work was supported by grants from the Spanish Ministry of Economy and Competitiveness (BIO2013-44963-R and BIO2016-77031-R to JB and BFU2014-55475-R to JRC), the FP7-PEOPLE-2012-IAPP from the European Union (Grant number 324439 to JB), and the Comunidad Autónoma de Madrid (S2013/MIT-2807 to JRC). An institutional grant from Fundación Ramón Areces to CBMSO is also acknowledged. AB and IB hold FPI fellowships from the Spanish Ministry of Education. CPM is a PhD fellow in the La Caixa Foundation International Fellowship Program (La Caixa/CNB). Hiring of NGQ through project BIO2013-50068-EXP is also acknowledged. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.