GENE-REGULATION AT THE RIGHT OPERATOR (OR) OF BACTERIOPHAGE-LAMBDA .2. OR1, OR2, AND OR3 - THEIR ROLES IN MEDIATING THE EFFECTS OF REPRESSOR AND CRO
GENE-REGULATION AT THE RIGHT OPERATOR (OR) OF BACTERIOPHAGE-LAMBDA .2. OR1, OR2, AND OR3 - THEIR ROLES IN MEDIATING THE EFFECTS OF REPRESSOR AND CRO
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DOI：
10.1016/0022-2836(80)90303-4
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发表时间：
1980-01-01
影响因子：
5.6
通讯作者：
PTASHNE, M
中科院分区：
文献类型：
--
作者：
MEYER, BJ;MAURER, R;PTASHNE, M
噬菌体的正确操作者，OR含有3个位点，每个位点被2个调节蛋白识别。这些蛋白质是。抑制子、cI基因的产物和cro基因的产物。这些蛋白质与OR的相互作用导致了对来自相邻启动子PR和PRM的转录的多种调节作用。每个操作网站在介导这些影响所发挥的作用进行了研究。体内PR和PRM的调节与λ的关系分析阻遏物或CRO蛋白浓度。利用体外重组构建的两种类型的杂合操纵子。一种类型，携带在噬菌体载体上，携带与噬菌体融合的lacZ基因。PR或... PRM.在细胞[大肠杆菌]中，这些噬菌体之一溶原性，β-的水平半乳糖苷酶是PR或PRM活性的量度。在另一种类型的杂合操纵子中，携带在质粒上，λ cI或cro基因与lac启动子融合并合成λ。调节蛋白由lac阻遏物控制。携带这些质粒之一的细胞合成增加量的.阻遏物或cro响应于IPTG [异丙基硫代-β- D-吡喃半乳糖苷]。因此，在一个典型的实验中，合成β-在携带一种噬菌体载体（作为原噬菌体）加上一种质粒的细胞中测量半乳糖苷酶作为IPTG浓度的函数。通过研究突变体的行为，揭示了OR位点在调节PR和PRM中的作用，一些新分离的突变体在OR中具有序列变化。除了一个突变之外，所有突变都位于OR 1、OR 2或OR 3中，并且每个突变都降低了相应位点对调节蛋白的亲和力。另一个是prmup-1，位于OR 2和OR 3之间。然而野生型PRM仅在阻遏物存在下有效地起作用，而prmup-1允许在阻遏物不存在下以高频率在PRM处发生起始。OR 1或OR 2（或两者）被任一蛋白质占据抑制PR。OR 3被任一蛋白质占据抑制PRM。仅与OR 2结合的阻遏物刺激PRM。在溶原细胞中，PR被抑制而PRM被刺激，因为阻遏物主要与OR 1和OR 2结合，但很少与OR 3结合。相反，随着cro蛋白的浓度在体内积累，首先PRM在OR 3被占据时关闭，然后PR在OR 1和OR 2被占据时关闭。这些事实解释了这两种调节蛋白的不同生理作用。提出了一个分子模型来解释这些调节作用的机制，其中考虑到规则描述合作结合的阻遏位点OR。
The right operator of bacteriophage .lambda., OR, contains 3 sites, each of which is recognized by 2 regulatory proteins. These proteins are the .lambda. repressor, the product of the cI gene, and the cro gene product. The interactions of these proteins with OR results in a variety of regulatory effects on transcription from the adjacent promoters PR and PRM. The roles played by each operator site in mediating these effects were studied. The regulation of PR and PRM in vivo as a function of .lambda. repressor or cro protein concentration was analyzed. Two types of hybrid operons constructed by recombination in vitro were utilized. One type, carried on a phage vector, bears the lacZ gene fused either to the .lambda. PR or the .lambda. PRM. In cells [Escherichia coli] lysogenic for one of these phages, the level of .beta.-galactosidase is a measure of the activity of PR or PRM. In the other type of hybrid operon, carried on a plasmid, a .lambda.cI or cro gene is fused to the lac promoter and synthesis of the .lambda. regulatory protein is controlled by the lac repressor. Cells bearing one of these plasmids synthesize increased amounts of .lambda. repressor or cro in response to increased amounts of IPTG [isopropylthio-.beta.-D-galactopyranoside]. In a typical experiment, therefore, the synthesis of .beta.-galactosidase, as a function of IPTG concentration, was measured in a cell bearing one of the phage vectors (as prophage) plus one of the plasmids. The roles of the OR sites in regulating PR and PRM were revealed by studying the behavior of mutants, some newly isolated, bearing sequence changes in OR. All but one mutation are located in OR1, OR2 or OR3, and each decreases the affinity of the corresponding site for the regulatory proteins. The other, prmup-1, is located between OR2 and OR3. Whereas wild-type PRM functions efficiently only in the presence of repressor, prmup-1 allows initiation to occur at PRM at high frequency in the absence of repressor. Occupation of OR1 or OR2 (or both) by either protein represses PR. Occupation of OR3 by either protein represses PRM. Repressor bound only to OR2 stimulates PRM. In a lysogen, PR is repressed and PRM is stimulated because repressor is bound predominantly to OR1 and OR2 but rarely to OR3. In contrast, as the concentration of cro protein accumulates in vivo, first PRM is turned off as OR3 is occupied and then PR is turned off as OR1 and OR2 are occupied. These facts explain the contrasting physiological effect of these 2 regulatory proteins. A molecular model is presented to explain the mechanism of each of these regulatory effects which takes into account the rules describing cooperative binding of repressor to sites in OR.