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摘要
One of the best understood systems in genetic regulatory biology is the so-called "genetic switch" that determines the choice the phage-encoded CI repressor binds cooperatively to tripartite operators, O-L and O-R, in a defined pattern, thus blocking the transcription at two lytic promoters, P-L and P-R, and auto-regulating the promoter, P-RM, which directs CI synthesis by the prophage. Fine-tuning of the maintenance of lysogeny is facilitated by interactions between CI dimers bound to O-R and O-L through the formation of a loop by the intervening DNA segment. By using a purified in vitro transcription system, we have genetically dissected the roles of individual operator sites in the formation of the DNA loop and thus have gained several new and unexpected insights into the system. First, although both O-R and O-L are tripartite, the presence of only a single active CI binding site in one of the two operators is sufficient for DNA loop formation. Second, in PL, unlike in P-R, the promoter distal operator site, O(L)3, is sufficient to directly repress PL. Third, DNA looping mediated by the formation of CI octamers arising through the interaction of pairs of dimers bound to adjacent operator sites in OR and OL does not require OR and OL to be aligned "in register", that is, CI bound to "out-of-register" sub-operators, for example, O(L)1 similar to O(1)2 and O(R)2 similar to O(R)3, can also mediate loop formation. Finally, based on an examination of the mechanism of activation of PRM when only O(R)1 or O(R)2 are wild type, we hypothesize that RNA polymerase bound at P-R interferes with DNA loop formation. Thus, the formation of DNA loops involves potential interactions between proteins bound at numerous cis-acting sites, which therefore very subtly contribute to the regulation of the "switch".
- 出版日期2016-11-6
- 单位NIH
