European Molecular Biology Laboratory, Postfach 10.2209, 6900 Heidelberg, FRG.
EMBO J. 1986 Dec 1;5(12):3391-9. doi: 10.1002/j.1460-2075.1986.tb04655.x.
Derivatives of the yeast 2mu plasmid with the cis-acting locus STB (also called REP3) are stably maintained if two plasmid-encoded proteins are present in trans. There are conflicting reports of both the extent of STB and its possible involvement in plasmid partition or copy number control. We have resolved the controversy by constructing 2micro derivatives with a conditional STB function, and showing that when STB is inactivated plasmids become concentrated in a small fraction of the population although the total number of plasmids remains unaltered. Moreover we show that STB consists of two functionally distinct domains which we call STB-proximal and STB-distal relative to the origin of replication. Although STB-proximal is sufficient for proper partitioning, this function is severely disrupted by active transcription from neighbouring sequences. STB-distal is important to protect STB-proximal and ORI from such transcription, and can be effeciently replaced by a 94-bp terminator fragment in an orientation-dependent manner. We find that STB-distal contains an additional element which depresses transcription from upstream promoters. We also describe the phenomenon of replicaton inhibition which we believe can exlain the anomalous instability of some yeast plasmids.
酵母 2μ 质粒的衍生物，如果存在两个质粒编码的蛋白质在顺式作用，就可以稳定地维持。关于 STB（也称为 REP3）的作用范围及其在质粒分配或拷贝数控制中的可能作用存在相互矛盾的报道。我们通过构建具有条件 STB 功能的 2μ 衍生物，解决了这一争议，并表明当 STB 失活时，质粒集中在种群的一小部分中，尽管质粒总数保持不变。此外，我们还表明，STB 由两个功能不同的结构域组成，相对于复制起点，我们称之为 STB 近端和 STB 远端。虽然 STB 近端足以进行适当的分配，但来自邻近序列的活跃转录会严重破坏这种功能。STB 远端对于保护 STB 近端和 ORI 免受这种转录至关重要，并且可以通过以依赖取向的方式有效地被一个 94 碱基对的终止子片段替换。我们发现 STB 远端包含一个额外的元件，它抑制了来自上游启动子的转录。我们还描述了复制抑制现象，我们认为这种现象可以解释某些酵母质粒的异常不稳定性。
