The gene transfer agent of Rhodobacter capsulatus (RcGTA) is a bacteriophage-like genetic element with the sole known function of horizontal gene transfer. Homologues of RcGTA genes are present in many members of the alphaproteobacteria and may serve an important role in microbial evolution. Transcription of RcGTA genes is induced as cultures enter the stationary phase; however, little is known about cis-active sequences. In this work, we identify the promoter of the first gene in the RcGTA structural gene cluster. Additionally, gene transduction frequency depends on the growth medium, and the reason for this is not known. We report that millimolar concentrations of phosphate posttranslationally inhibit the lysis-dependent release of RcGTA from cells in both a complex medium and a defined medium. Furthermore, we found that cell lysis requires the genes rcc00555 and rcc00556, which were expressed and studied in Escherichia coli to determine their predicted functions as an endolysin and holin, respectively. Production of RcGTA is regulated by host systems, including a putative histidine kinase, CckA, and we found that CckA is required for maximal expression of rcc00555 and for maturation of RcGTA to yield gene transduction-functional particles.

Yeast iso-1-cytochrome c (Cc) mutants have been constructed with Phe, Tyr, Gly, Ser, Leu, and Ile at position 82, each with Thr substituted for Cys at position 102. Their long-range electron transfer with zinc-substituted cytochrome c peroxidase (ZnCcP) has been studied by two kinetic techniques. The charge-separated complex, [(ZnCcP)+,FeIICc] converts to [ZnCcP,FeIIICc] by a single, intracomplex electron transfer step that is not governed by "gating" through possible rapid dissociation of the complex or isomerization (for example, heme-ligand) by FeIICc subsequent to its formation from FeIIICc. In every variant with an aliphatic residue at position 82 of Cc, the rate of this electron transfer process is approximately 10(4) slower at approximately 0 degrees C than for the two variants with aromatic residues.

Gene transfer agents (GTAs) morphologically resemble small, double-stranded DNA (dsDNA) bacteriophages; however, their only known role is to package and transfer random pieces of the producing cell genome to recipient cells. The best understood GTA is that of Rhodobacter capsulatus, termed RcGTA. We discovered that homologues of three genes involved in natural transformation in other bacteria, comEC, comF, and comM, are essential for RcGTA-mediated gene acquisition. This paper gives genetic and biochemical evidence that RcGTA-borne DNA entry into cells requires the ComEC and ComF putative DNA transport proteins and genetic evidence that putative cytoplasmic ComM protein of unknown function is required for recipient capability. Furthermore, the master regulator of RcGTA production in <1% of a cell population, CtrA, which is also required for gene acquisition in recipient cells, is expressed in the vast majority of the population. Our results indicate that RcGTA-mediated gene transfer combines key aspects of two bacterial horizontal gene transfer mechanisms, where donor DNA is packaged in transducing phage-like particles and recipient cells take up DNA using natural transformation-related machinery. Both of these differentiated subsets of a culture population, donors and recipients, are dependent on the same response regulator, CtrA. IMPORTANCEHorizontal gene transfer (HGT) is a major driver of bacterial evolution and adaptation to environmental stresses. Traits such as antibiotic resistance or metabolic properties can be transferred between bacteria via HGT; thus, HGT can have a tremendous effect on the fitness of a bacterial population. The three classically described HGT mechanisms are conjugation, transformation, and phage-mediated transduction. More recently, the HGT factor GTA was described, where random pieces of producing cell genome are packaged into phage-like particles that deliver DNA to recipient cells. In this report, we show that transport of DNA borne by the R. capsulatus RcGTA into recipient cells requires key genes previously thought to be specific to natural transformation pathways. These findings indicate that RcGTA combines central aspects of phage-mediated transduction and natural transformation in an efficient, regulated mode of HGT.T he first evidence of prokaryotic genetic exchange was transformation, a term coined in 1928 by Griffith (1). Subsequently, conjugation was observed in 1946 (2), followed by transduction, which was discovered in 1952 (3). More recently, another prokaryotic mode of horizontal gene transfer, dependent on an extracellular particle called a gene transfer agent (GTA), was described (4), and GTAs subsequently have been discovered in diverse prokaryotes (5).GTAs varying in morphology have been reported, although most resemble small, tailed double-stranded DNA (dsDNA) bacteriophages. The general criteria that define a GTA are (i) the DNA packaged within the head is insufficient to carry the GTA structural genes; (ii) all GTA particles package only random parts o...

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Uncertainty surrounding the role of Dientamoeba fragilis in human disease could be due in part to the existence of pathogenic and nonpathogenic variants. Evidence for two genetically distinct forms was obtained using PCR-restriction fragment length polymorphism analysis of ribosomal genes. Future studies in humans will need to take D. fragilis diversity into account.