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Transposon Mutagenesis in Chlamydia trachomatis Identifies CT339 as a ComEC Homolog Important for DNA Uptake and Lateral Gene Transfer
LaBrie, Scott D. Dimond, Zoe E. Harrison, Kelly S. Baid, Srishti Wickstrum, Jason Schuland, Robert J. Hefty, P. Scott
LaBrie, Scott D.
Dimond, Zoe E.
Harrison, Kelly S.
Baid, Srishti
Wickstrum, Jason
Schuland, Robert J.
Hefty, P. Scott
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Abstract
Transposon mutagenesis is a widely applied and powerful genetic tool for
the discovery of genes associated with selected phenotypes. Chlamydia trachomatis is a
clinically significant, obligate intracellular bacterium for which many conventional genetic tools and capabilities have been developed only recently. This report describes the
successful development and application of a Himar transposon mutagenesis system for
generating single-insertion mutant clones of C. trachomatis. This system was used to
generate a pool of 105 transposon mutant clones that included insertions in genes encoding flavin adenine dinucleotide (FAD)-dependent monooxygenase (C. trachomatis 148
[ct148]), deubiquitinase (ct868), and competence-associated (ct339) proteins. A subset of
Tn mutant clones was evaluated for growth differences under cell culture conditions, revealing that most phenocopied the parental strain; however, some
strains displayed subtle and yet significant differences in infectious progeny production and inclusion sizes. Bacterial burden studies in mice also supported the
idea that a FAD-dependent monooxygenase (ct148) and a deubiquitinase (ct868)
were important for these infections. The ct339 gene encodes a hypothetical protein with limited sequence similarity to the DNA-uptake protein ComEC. A transposon insertion in ct339 rendered the mutant incapable of DNA acquisition during recombination experiments. This observation, along with in situ structural
analysis, supports the idea that this protein is playing a role in the fundamental
process of lateral gene transfer similar to that of ComEC. In all, the development
of the Himar transposon system for Chlamydia provides an effective genetic tool
for further discovery of genes that are important for basic biology and pathogenesis aspects.
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Date
2019-08-06
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American Society for Microbiology
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Keywords
Chlamydia trachomatis, genetic competence, mutagenesis, transposons
Citation
LaBrie SD, Dimond ZE, Harrison KS,
Baid S, Wickstrum J, Suchland RJ, Hefty PS.
2019. Transposon mutagenesis in Chlamydia
trachomatis identifies CT339 as a ComEC
homolog important for DNA uptake and lateral
gene transfer. mBio 10:e01343-19. https://doi
.org/10.1128/mBio.01343-19.