Random transposon mutagenesis identifies genes essential for transformation in Methanococcus maripaludis

Dallas R. Fonseca, Madison B. Loppnow, Leslie A. Day, Elisa L. Kelsey, Mohd Farid Abdul Halim, Kyle C. Costa

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

Natural transformation, the process whereby a cell acquires DNA directly from the environment, is an important driver of evolution in microbial populations, yet the mechanism of DNA uptake is only characterized in bacteria. To expand our understanding of natural transformation in archaea, we undertook a genetic approach to identify a catalog of genes necessary for transformation in Methanococcus maripaludis. Using an optimized method to generate random transposon mutants, we screened 6144 mutant strains for defects in natural transformation and identified 25 transformation-associated candidate genes. Among these are genes encoding components of the type IV-like pilus, transcription/translation associated genes, genes encoding putative membrane bound transport proteins, and genes of unknown function. Interestingly, similar genes were identified regardless of whether replicating or integrating plasmids were provided as a substrate for transformation. Using allelic replacement mutagenesis, we confirmed that several genes identified in these screens are essential for transformation. Finally, we identified a homolog of a membrane bound substrate transporter in Methanoculleus thermophilus and verified its importance for transformation using allelic replacement mutagenesis, suggesting a conserved mechanism for DNA transfer in multiple archaea. These data represent an initial characterization of the genes important for transformation which will inform efforts to understand gene flow in natural populations. Additionally, knowledge of the genes necessary for natural transformation may assist in identifying signatures of transformation machinery in archaeal genomes and aid the establishment of new model genetic systems for studying archaea.

Original languageEnglish (US)
Pages (from-to)537-548
Number of pages12
JournalMolecular Genetics and Genomics
Volume298
Issue number3
DOIs
StatePublished - May 2023

Bibliographical note

Funding Information:
We thank Michael Rother for providing strains and constructs for the purification of transposases and generation of transposon mutants. This study was supported by the National Science Foundation under grant number MCB-2148165.

Publisher Copyright:
© 2023, The Author(s).

Keywords

  • Archaea
  • Competence
  • Genetics
  • Natural transformation
  • Transposon mutagenesis

PubMed: MeSH publication types

  • Journal Article

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