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Identification and characterization of a direct activator of a gene transfer agent.

ABSTRACT: Gene transfer agents (GTAs) are thought to be ancient bacteriophages that have been co-opted into serving their host and can now transfer any gene between bacteria. Production of GTAs is controlled by several global regulators through unclear mechanisms. In Rhodobacter capsulatus, gene rcc01865 encodes a putative regulatory protein that is essential for GTA production. Here, I show that rcc01865 (hereafter gafA) encodes a transcriptional regulator that binds to the GTA promoter to initiate production of structural and DNA packaging components. Expression of gafA is in turn controlled by the pleiotropic regulator protein CtrA and the quorum-sensing regulator GtaR. GafA and CtrA work together to promote GTA maturation and eventual release through cell lysis. Identification of GafA as a direct GTA regulator allows the first integrated regulatory model to be proposed and paves the way for discovery of GTAs in other species that possess gafA homologues.

SUBMITTER: Fogg PCM

PROVIDER: S-EPMC6363796 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Identification and characterization of a direct activator of a gene transfer agent.

Fogg Paul C M PCM

Nature communications 20190205 1

Gene transfer agents (GTAs) are thought to be ancient bacteriophages that have been co-opted into serving their host and can now transfer any gene between bacteria. Production of GTAs is controlled by several global regulators through unclear mechanisms. In Rhodobacter capsulatus, gene rcc01865 encodes a putative regulatory protein that is essential for GTA production. Here, I show that rcc01865 (hereafter gafA) encodes a transcriptional regulator that binds to the GTA promoter to initiate produ ...[more]

PMID: 30723210

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Project description:VSH-1 is a mitomycin C-inducible prophage of the anaerobic spirochete Brachyspira hyodysenteriae. Purified VSH-1 virions are noninfectious, contain random 7.5-kb fragments of the bacterial genome, and mediate generalized transduction of B. hyodysenteriae cells. In order to identify and sequence genes of this novel gene transfer agent (GTA), proteins associated either with VSH-1 capsids or with tails were purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The N-terminal amino acid sequences of 11 proteins were determined. Degenerate PCR primers were designed from the amino acid sequences and used to amplify several VSH-1 genes from B. hyodysenteriae strain B204 DNA. A lambda clone library of B. hyodysenteriae B204 DNA was subsequently screened by Southern hybridization methods and used to identify and sequence overlapping DNA inserts containing additional VSH-1 genes. VSH-1 genes spanned 16.3 kb of the B. hyodysenteriae chromosome and were flanked by bacterial genes. VSH-1 identified genes and unidentified, intervening open reading frames were consecutively organized in head (seven genes), tail (seven genes), and lysis (four genes) clusters in the same transcriptional direction. Putative lysis genes encoding endolysin (Lys) and holin proteins were identified from sequence and structural similarities of their translated protein products with GenBank bacteriophage proteins. Recombinant Lys protein hydrolyzed peptidoglycan purified from B. hyodysenteriae cells. The identified VSH-1 genes exceed the DNA capacity of VSH-1 virions and do not encode traditional bacteriophage early functions involved in DNA replication. These genome properties explain the noninfectious nature of VSH-1 virions and further confirm its resemblance to known prophage-like, GTAs of other bacterial species, such as the GTA from Rhodobacter capsulatus. The identification of VSH-1 genes will enable analysis of the regulation of this GTA and should facilitate investigations of VSH-1-like prophages from other Brachyspira species.

Dataset Information

Identification and characterization of a direct activator of a gene transfer agent.

Publications

Identification and characterization of a direct activator of a gene transfer agent.

Similar Datasets

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