Institut für Molekulare Biowissenschaften, Karl-Franzens-Universität Graz, Humboldtstrasse 50, A-8010 Graz, Austria.
Microbiology (Reading). 2013 Dec;159(Pt 12):2481-2491. doi: 10.1099/mic.0.071738-0. Epub 2013 Oct 1.
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 PY, 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 PY 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 PY promoter of plasmid R1. We show that TraJ binds to PY DNA in vivo and that H-NS acts as a silencer of the PY 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.
F 型接合质粒的 DNA 转移（tra）基因的表达对于功能性 IV 型分泌机制的组装以及随后质粒 DNA 从供体转移到受体细胞是必需的。tra 基因的转录依赖于质粒编码的 TraJ 蛋白对单个启动子 PY 的激活。我们在这里确定了来自各种 F 样质粒亚群的 TraJ 蛋白的质粒特异性，发现只有其同源激活子和沙门氏菌质粒 pSLT 的 TraJ 才能实现 R1 质粒的接合和 PY 启动子的激活，而 F 或 R100 TraJ 蛋白则不能。此外，我们还对质粒 R1 的 PY 启动子进行了表征。我们表明 TraJ 在体内与 PY DNA 结合，而 H-NS 作为 PY 启动子的沉默子。在天然质粒环境中，H-NS 沉默转移基因的表达和水平质粒 DNA 转移。与 F 质粒的发现相反，缺乏 H-NS 并没有消除 ArcA 和 TraJ 对达到完全 tra 基因表达和 DNA 转移活性的要求。我们提出，除了被动去沉默活性外，ArcA 和 TraJ 都在协同刺激 tra 操纵子转录和随后的 DNA 转移中发挥直接作用。
