ANTAGONISTIC CONTROLS REGULATE COPY NUMBER OF THE YEAST 2-MU PLASMID
ANTAGONISTIC CONTROLS REGULATE COPY NUMBER OF THE YEAST 2-MU PLASMID
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DOI：
10.1002/j.1460-2075.1987.tb02768.x
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发表时间：
1987-12-01
期刊：
影响因子：
11.4
通讯作者：
CESARENI, G
中科院分区：
文献类型：
--
作者：
MURRAY, JAH;SCARPA, M;CESARENI, G
内源2.mu。酵母质粒编码位点特异性重组酶 FLP，可导致质粒拷贝数扩增。使用过表达2.mu的菌株。染色体构建体中的质粒蛋白破坏基因产物的正常平衡，我们在此表明拷贝数是通过调节 FLP 的转录水平来控制的。 FLP 的表达通过质粒编码的 REP1 和 REP2 蛋白的联合作用受到 100 倍范围内的负调节，这些蛋白也在质粒分割中发挥作用。我们还表明，第四个质粒开放阅读框 D 的产物通过缓解 REP1 和 REP2 引起的抑制来增加 FLP 表达。这是该基因功能的首次演示，我们称之为 RAF。 RAF 的转录也受到 REP1 和 REP2 共同作用的抑制，但完全抑制所需的水平比抑制 FLP 所需的水平更高。因此，拷贝数受到 REP1-REP2 浓度的负调节，既可以通过 FLP 的直接抑制，也可以通过控制正向元件（抗阻遏物 RAF）间接调节。我们认为这些拮抗调节机制放大了拷贝数小幅下降产生的信号。
The endogenous 2.mu. plasmid of yeast encodes a site-specific recombinase FLP that can cause an amplification of plasmid copy number. Using strains overexpressing 2.mu. plasmid proteins from chromosomal constructs to disrupt the normal balance of gene products, we show here that copy number is controlled by regulating the transcript level of FLP. Expression of FLP is negatively regulated over a 100-fold range by the joint action of the plasmid-encoded REP1 and REP2 proteins, which also have a role in plasmid partition. We also show that the product of the fourth plasmid open reading frame D increases FLP expression by relieving the repression caused by REP1 and REP2. This is the first demonstration of a function for this gene, which we call RAF. The transcription of RAF is also repressed by REP1 and REP2 acting together, but requires a higher level for complete inhibition than that required to repress FLP. Copy number is therefore negatively regulated by REP1-REP2 concentration both by direct repression of FLP and indirectly, by control of the positive element, the anti-repressor RAF. We propose that these antagonistic regulatory mechanisms amplify the signal produced by a small fall in copy number.