Cis-acting, orientation-dependent, positive control system activates pheromone-inducible conjugation functions at distances greater than 10 kilobases upstream from its target in Enterococcus faecalis.
Cis-acting, orientation-dependent, positive control system activates pheromone-inducible conjugation functions at distances greater than 10 kilobases upstream from its target in Enterococcus faecalis.
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顺式作用、方向依赖性、正控制系统在粪肠球菌中距目标上游 10 kb 以上的距离处激活信息素诱导的缀合功能。
DOI：
10.1073/pnas.89.19.9020
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发表时间：
1992
影响因子：
11.1
通讯作者：
Dunny,GM
中科院分区：
文献类型：
--
作者：
Chung,JW;Dunny,GM
PrGB基因编码表面蛋白Asc10，Asc10介导细胞聚集，导致信息素诱导的四环素耐药质粒pCF10在粪肠球菌中的高频接合转移。通过Northern杂交和引物延伸对信使RNA的分析表明，PrGB的转录是信息素诱导的单顺反子。先前的转座子突变和对pCF10一个12kb碱基区的测序分析表明，包括prgR和prgs在内的几个基因需要激活pRGB的表达。这些调控基因与PRGB之间的距离(3-4kb)表明，这种激活可能在反式转录中起作用。为了验证这一点，构建并克隆了一个与pRGB融合的无启动子的LacZ基因，其中没有部分或全部调控基因。在一个拷贝数较高的质粒中克隆了该调节区的几个限制性片段，并在粪肠球菌中进行了大量的互补研究。在这些实验中没有观察到反式作用的互补。然而，当调节区和靶基因被克隆在同一质粒的不同位置，相隔12kb时，可以观察到PRGB的激活。有趣的是，只有当这些区域以与野生型pCF10上相同的相对方向克隆时，这种激活才会发生。这些结果表明，一个或多个调控分子可能结合到上游顺式作用位点，并沿着DNA跟踪到激活PRGB转录的靶部位。
The prgB gene encodes the surface protein, Asc10, which mediates cell aggregation, resulting in high-frequency conjugative transfer of the pheromone-inducible tetracycline-resistance plasmid pCF10 in Enterococcus faecalis. Messenger RNA analysis by Northern blot hybridization and primer extension indicates that prgB transcription is pheromone-inducible and monocistronic. Previous transposon mutagenesis and sequencing analysis of a 12-kilobase (kb) region of pCF10 indicated that several genes including prgR and prgS are required to activate expression of prgB. The distance (3-4 kb) between these regulatory genes and prgB suggested that the activation might function in trans. To test this, a promoterless lacZ gene fusion to prgB was constructed and cloned without some or all of the regulatory genes. Several restriction fragments of the regulatory region were cloned in a higher copy-number plasmid, and numerous complementation studies were carried out in E. faecalis. Complementation in trans was not observed in any of these experiments. However, when the regulatory region and target genes were cloned in different sites of the same plasmid, separated by as much as 12 kb, activation of prgB was observed. Interestingly, this activation occurred only when the regions were cloned in the same relative orientation in which they exist on wild-type pCF10. These results suggest that one or more regulatory molecules may bind to an upstream cis-acting site and track along the DNA to reach a target site to activate prgB transcription.
影响因子：
4.3
作者：
H. Sambrook
通讯作者：
H. Sambrook