Activation of transcription from Lambda lytic promoters, PL and PR by Cro: direct role in the epigenetic switch. As stated before, the Lambda Cro protein facilitates the phage's lytic growth by an epigenetic switch. It has been shown that Cro does so by turning off CI synthesis by repressing the PRM promoter by binding to OR3 after phage infection or prophage induction. Thus in the absence of CI the lytic promoters would be derepressed to help lytic growth. Cro binds to OR3 with a C1/2 of 100 nM. However, it takes a long time after the PRM turn-off by Cro to decrease the CI level by SOS cleavage to the level that would cause PR and PL derepression for facilitating phage lytic growth. The existing CI has to be diluted out by cell division for a full switch to lytic mode of the phage even after CI synthesis has been turned off by Cro. But repression of PL and PR happens sooner. To explain the paradox, we proposed a direct role of Cro in helping PL and PR transcription even in the presence of CI. Our idea has been corroborated by the following results. We studied the regulation of PR, PL and PRM in the presence of both CI and Cro in vivo and in vitro. Although these experiments are still in progress but it is clear that under the conditions in which CI at 120 nM concentration represses PR and PL 90%, the presence of even a very low concentration of Cro (75 nM) causes significant derepression of PL and PR establishing that the 'genetic switch' by Cro is more direct rather than the indirect model proposed earlier. It was assumed that the actions of Cro and CI during induction are independent of each other. We found Cro increases PL and PR in the presence of 120 nM CI. In vitro, the lytic promoters became active compared with CI-only control when a low concentration of Cro was simultaneously made available, suggesting that Cro alleviates the repression of PL and PR by CI. This is also true in vivo. When Cro protein was made in trans from a plasmid to a repressed prophage, a lacZ reporter gene linked to PR derepressed b-galactosidase synthesis as judged by red color on MacConkey-lactose agar plates. No red color was observed from the equivalent strain harbors the plasmid without cro. These paradigm-shifting results contradict the classical theory of the genetic switch in Lambda prophage induction, suggesting that Cro primarily causes prophage induction by inactivating the action of CI repressor to derepress the lytic promoters PR and PL long before it turns off CI synthesis. The details would be discussed in the presentation.
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