DNA packaging bias and differential expression of gene transfer agent genes within a population during production and release of the Rhodobacter capsulatus gene transfer agent, RcGTA.
Rhodobacter capsulatus produces a gene transfer agent (GTA) called RcGTA. RcGTA is a phage-like particle that packages R. capsulatus DNA and transfers it to other R. capsulatus cells. We quantified the relative frequency of packaging for each gene in the genome by hybridization of DNA from RcGTA particles to an R. capsulatus microarray. All genes were found within the RcGTA particles. However, the genes encoding the RcGTA particle were under-packaged compared with other regions. Gene transfer bioassays confirmed that the transfer of genes within the RcGTA structural cluster is reduced relative to those of other genes. Single-cell expression analysis, by flow cytometry analysis of cells containing RcGTA-reporter gene fusion constructs, demonstrated that RcGTA gene expression is not uniform within a culture. This phenomenon was accentuated when the constructs were placed in a strain lacking a putative lysis gene involved in RcGTA release; a small subpopulation was found to be responsible for ∼ 95% of RcGTA activity. We propose a mechanism whereby high levels of RcGTA gene transcription in the most active RcGTA-producing cells cause a reduction in their packaging frequency. This subpopulation's role in producing and releasing the RcGTA particles explains the lack of observed cell lysis in cultures.
Department of Biology, Memorial University of Newfoundland, 232 Elizabeth Ave., St John's, NL, A1B 3X9, Canada.
Alexander P
AP
Hynes
Ryan G
RG
Mercer
David E
DE
Watton
Colleen B
CB
Buckley
Andrew S
AS
Lang
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics; D042003:DNA Packaging; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D022761:Gene Transfer, Horizontal; D017930:Genes, Reporter; D008969:Molecular Sequence Data; D016354:Rhodobacter capsulatus / Q000235:genetics* / Q000378:metabolism*
D001426:Bacterial Proteins; D004269:DNA, Bacterial