Silencing and activating type IV secretion genes of the F-like conjugative resistance plasmid R1

Abstract

Expression of DNA transfer (tra) genes of F-type conjugative plasmids is required for the assembly of a functional type IV secretion machinery and subsequent plasmid DNA transfer from donor to recipient cells. Transcription of tra genes depends on the activation of a single promoter, designated P Y , by the plasmid encoded TraJ protein. We here determine plasmid specificity of TraJ proteins from various subgroups of F-like plasmids and find that plasmid R1 conjugation and P Y promoter activation can be achieved only by its cognate activator and by TraJ of the Salmonella plasmid pSLT and not by F or R100 TraJ proteins. In addition, we characterize the P Y promoter of plasmid R1. We show that TraJ binds to P Y DNA in vivo and that H-NS acts as a silencer of the P Y promoter. In the natural plasmid context, H-NS silences transfer gene expression and horizontal plasmid DNA transfer. In contrast to what was found for the F plasmid, lack of H-NS did not abolish the requirement for ArcA and TraJ to reach full tra gene expression and DNA transfer activity. We propose that, besides a passive de-silencing activity, both ArcA and TraJ play a direct role in synergistically stimulating tra operon transcription and subsequent DNA transfer.

Social behavior and decision making in bacterial conjugation

“…This repression of foreign genes termed “xenogeneic silencing” was also found in plasmid F (Will and Frost, 2006) and the naturally repressed plasmid R1 where H-NS contributes to the low frequency of transfer competent cells in a population of cells carrying this CP (Wagner et al, 2013). …”

“…While FinP is a small regulatory RNA that is produced as a countertranscript to the translation initiation region of the TraJ mRNA, FinO is an RNA chaperone that is required for efficient suppression of TraJ expression (Arthur et al, 2003). In populations of donor cells—under optimal conditions promoting growth and cell division—only in few cells (1–10 out of 1000 potential donor cells) TraJ escapes this negative FinOP mediated control and promotes transcription of tra genes together with the host encoded transcriptional activator ArcA-P (Strohmaier et al, 1998; Frost and Koraimann, 2010; Wagner et al, 2013). Once initiated, a positive feedback-loop leads to a burst of tra gene expression which ensures the transformation into a transfer competent cell (Dempsey, 1989; Pölzleitner et al, 1997).…”

“…These findings are consistent with the observation in liquid media where tra gene expression is low in the presence of plasmid R1 compared to a de-repressed mutant. Furthermore, gene transfer in liquid media occurs at a low frequency indicating that only about 1% of donor cells have activated their tra genes and transformed into transfer competent cells (Wagner et al, 2013). …”

Crystal Structures of the F and pSLT Plasmid TraJ N-Terminal Regions Reveal Similar Homodimeric PAS Folds with Functional Interchangeability

“…4 To further test the allelic specificity of different TraJ proteins, we assessed the ability of TraJ from F, pSLT, and R100 to drive transcription from an F P Y - lacZ fusion reporter plasmid, pJLac101-P Y . 23 Each of the TraJ proteins was expressed from the pK184 expression plasmid such that each was produced at close to physiological levels.…”

“…3 TraJ is thought to function to relieve P Y from H-NS (histone-like nucleoid structuring protein) repression as well as to activate P Y in an independent mechanism together with a host factor, ArcA. 4−6 As a central regulator of bacterial conjugation, the intracellular level of TraJ is stringently regulated at multiple levels by different host and plasmid-encoded factors. CRP (catabolite repressor protein) and Lrp (leucine-responsive protein) regulate traJ transcription, 7−9 whereas Hfq (host factor for Qβ replicase) and the plasmid-encoded FinOP (fertility inhibition) system affect TraJ translation.…”