Wegrzyn A, Wegrzyn G, Taylor K
Laboratory of Molecular Biology, Polish Academy of Sciences, Gdańsk.
Mol Gen Genet. 1995 May 20;247(4):501-8. doi: 10.1007/BF00293153.
We have shown previously that in amino acid-starved, relaxed (rel-) mutants of Escherichia coli replication of the lambda plasmid occurs via the lambda O-containing replication complex (RC) that was assembled prior to the onset of amino acid starvation and is inherited by one of the two daughter plasmid circles in each replication cycle. This replication is regulated neither by binding of the lambda O initiator to ori lambda, nor by the lambda Cro-mediated repression. Here we show that it is dependent on both RNA polymerase and DnaA functions, which is consistent with our recent finding that transcriptional activation of ori lambda is under the control of DnaA. In the system studied, DnaA-regulated transcriptional activation of ori lambda seems to be the only rate-limiting process. The lambda plasmid replication mediated by the inherited RC appeared to be independent of the functions of lambda P and DnaJ required in RC assembly In vitro experiments performed by others suggest that DnaJ first binds to the ori lambda-bound lambda O-lambda P-DnaB pre-primosome and subsequently lambda P complexed with DnaJ is preferentially recognized by DnaK-GrpE; chaperone-mediated rearrangement of this structure relieves DnaB helicase of lambda P inhibition. Recently we proposed that this process is directly coupled to the insertion of the pre-primosome between DNA strands transiently separated by transcription. This last-mentioned process may be required in lambda plasmid replication mediated by the inherited RC, which appeared in turn to be dependent on DnaK and GrpE functions.
我们之前已经表明，在氨基酸饥饿的大肠杆菌松弛（rel-）突变体中，λ质粒的复制通过含λO的复制复合体（RC）进行，该复合体在氨基酸饥饿开始之前组装而成，并在每个复制周期中由两个子代质粒环之一继承。这种复制既不受λO起始蛋白与ori λ的结合调控，也不受λCro介导的抑制作用调控。在此我们表明，它依赖于RNA聚合酶和DnaA的功能，这与我们最近发现ori λ的转录激活受DnaA控制是一致的。在所研究的系统中，DnaA调控的ori λ转录激活似乎是唯一的限速过程。由继承的RC介导的λ质粒复制似乎独立于RC组装中所需的λP和DnaJ的功能。其他人进行的体外实验表明，DnaJ首先与结合在ori λ上的λO-λP-DnaB前引发体结合，随后与DnaJ复合的λP优先被DnaK-GrpE识别；伴侣介导的这种结构重排解除了λP对DnaB解旋酶的抑制作用。最近我们提出，这个过程直接与前引发体插入由转录暂时分开的DNA链之间相偶联。在由继承的RC介导的λ质粒复制中可能需要最后提到的这个过程，而这反过来又似乎依赖于DnaK和GrpE的功能。
