9. A second repressor is needed for lytic infection
11.9 A second repressor is needed for lytic infection |
We started this chapter by saying that lambda has the alternatives of entering lysogeny or starting a lytic infection. Lysogeny is initiated by establishing an autogenous maintenance circuit that inhibits the entire lytic cascade through applying pressure at two points. The program for establishing lysogeny proceeds through some of the same events that we described earlier in terms of the lytic cascade (expression of delayed early genes via expression of N is needed). We now face a problem. How does the phage enter the lytic cycle?
What we have left out of this account so far is the role of gene cro, which codes for another repressor. Cro is responsible for preventing the synthesis of the repressor protein; this action shuts off the possibility of establishing lysogeny. cro mutants usually establish lysogeny rather than entering the lytic pathway, because they lack the ability to switch events away from the expression of repressor.
Cro forms a small dimer (the subunit is 9 kD) that acts within the immunity region. It has two effects:
- It prevents the synthesis of repressor via the maintenance circuit; that is, it prevents transcription via PRM.
- It also inhibits the expression of early genes from both PL and PR.
This means that, when a phage enters the lytic pathway, Cro has responsibility both for preventing the synthesis of repressor and (subsequently) for turning down the expression of the early genes.
Cro achieves its function by binding to the same operators as (cI) repressor protein. Cro includes a region with the same general structure as the repressor; a helix-2 is offset at an angle from recognition helix-3. (The remainder of the structure is different, demonstrating that the helix-turn-helix motif can operate within various contexts.) Like repressor, Cro binds symmetrically at the operators.
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Figure 11.19 Two proteins that use the two-helix arrangement to contact DNA recognize lambda operators with affinities determined by the amino acid sequence of helix-3. |
The sequences of Cro and repressor in the helix-turn-helix region are related, explaining their ability to contact the same DNA sequences (see Figure 11.19). Cro makes similar contacts to those made by repressor, but binds to only one face of DNA; it lacks the N-terminal arms by which repressor reaches around to the other side.
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Figure 11.27 The lytic cascade requires Cro protein, which directly prevents repressor maintenance via PRM, as well as turning off delayed early gene expression, indirectly preventing repressor establishment |
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Figure 11.26 A cascade is needed to establish lysogeny, but then this circuit is switched off and replaced by the autogenous repressor-maintenance circuit. |
How can two proteins have the same sites of action, yet have such opposite effects? The answer lies in the different affinities that each protein has for the individual binding sites within the operators. (Also Cro has no activating region.) Let us just consider OR, where more is known, and where Cro exerts both its effects. The series of events is illustrated in Figure 11.27. (Note that the first two stages are identical to those of the lysogenic circuit shown in Figure 11.26.)
The affinity of Cro for OR3 is greater than its affinity for OR2 or OR1. So it binds first to OR3. This inhibits RNA polymerase from binding to PRM. So Cro’s first action is to prevent the maintenance circuit for lysogeny from coming into play.
Then Cro binds to OR2 or OR1. Its affinity for these sites is similar, and there is no cooperative effect. Its presence at either site is sufficient to prevent RNA polymerase from using PR. This in turn stops the production of the early functions (including Cro itself). Because CII is unstable, any use of PRE is brought to a halt. So the two actions of Cro together block all production of repressor.
So far as the lytic cycle is concerned, Cro turns down (although it does not completely eliminate) the expression of the early genes. Its incomplete effect is explained by its affinity for OR1 and OR2, which is about eight times lower than that of repressor. This effect of Cro does not occur until the early genes have become more or less superfluous, because pQ is present; by this time, the phage has started late gene expression, and is concentrating on the production of progeny phage particles.
