Kukolj G, DuBow M S
Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada.
J Biol Chem. 1992 Sep 5;267(25):17827-35.
The lytic-lysogenic switch in transposable, Mu-like bacteriophage D108 is governed by two divergent and slightly overlapping transcription units originating from the Pe and Pc promoters. DNase I footprinting and in vivo mutational analysis suggest that lysogeny is maintained by c-repressor occupancy of the O2 operator, which precludes RNA polymerase from binding to Pe. Lytic development is controlled by the Ner repressor, which binds to a site symmetrically situated between the converging promoters and, in the absence of other factors, prevents RNA polymerase from binding to either Pc or Pe. DNase I protection and potassium permanganate hypersensitivity in the presence of integration host factor (IHF), which binds and alters the DNA structure upstream of Pe, revealed that RNA polymerase was able to bind Pe irrespective of the Ner.DNA-bound complex, and partially unwind the Pe "-10 region." Ner repression of Pe transcription in vitro was significantly more effective in the absence of IHF. Using a cloned D108 early region-lacZ fusion in IHF-deficient and -proficient backgrounds, we also demonstrate this host factor's affect on ner-repressed Pe in vivo, and generate a system for isolating mutants in the regulatory genes and sites controlling this genetic switch. D108 lytic growth is proposed to occur through IHF-mediated activation of the phage Ner-repressed early operon.
可转座的类Mu噬菌体D108中的裂解-溶原开关由源自Pe和Pc启动子的两个反向且略有重叠的转录单元控制。DNA酶I足迹分析和体内突变分析表明，溶原状态通过O2操纵子上c-阻遏物的占据得以维持，这使得RNA聚合酶无法结合到Pe上。裂解发育由Ner阻遏物控制，它结合在两个汇聚启动子之间对称位置的一个位点上，在没有其他因子的情况下，阻止RNA聚合酶结合到Pc或Pe上。在存在整合宿主因子（IHF）的情况下进行的DNA酶I保护和高锰酸钾超敏性分析显示，IHF结合并改变Pe上游的DNA结构，无论Ner-DNA结合复合物如何，RNA聚合酶都能够结合到Pe上，并部分解开Pe的“-10区域”。在没有IHF的情况下，Ner在体外对Pe转录的抑制作用明显更强。利用在缺乏和具有IHF背景下的克隆D108早期区域-lacZ融合体，我们还证明了这种宿主因子在体内对受Ner抑制的Pe的影响，并建立了一个用于分离调控基因和控制此遗传开关位点突变体的系统。有人提出，D108的裂解生长是通过IHF介导的对噬菌体Ner抑制的早期操纵子的激活而发生的。
