Regulation in a Lysogen

In a lysogenic bacterium, the genes of the phage lytic cycle must be turned off, or the cell would die. Turnoff is caused by a phage-coded repressor (usually a protein, but in some cases, as with phage P4, an antisense RNA). The repressor prevents transcriptional initiation from key phage promoters. This turns off almost all phage transcription. Repressor acts directly on only a few promoters (in l, there are two). These are the promoters from which lytic development is initiated. Phage genes expressed late in lytic development are turned off indirectly, by the lack of early gene products needed to activate their expression. The skeletal control system in l (where the repressor is coded by the cl gene) is shown in figure 7-7. Repressor not only turns off transcription from the lytic promoters pL and pR but also

Figure 7-7 Outline of gene control in a l lysogen. Repressor (product of gene cl) binds to operator sites (oL, oR, also known as OL and OR, respectively), turning off transcription from pL (also known as PL) and pR (also known as PR) and turning on cI transcription from pM. Repressor blocks transcription of late genes indirectly, by preventing transcription from gene Q, whose product turns on late gene transcription of gene N, whose product antiterminates early gene transcription.

Figure 7-7 Outline of gene control in a l lysogen. Repressor (product of gene cl) binds to operator sites (oL, oR, also known as OL and OR, respectively), turning off transcription from pL (also known as PL) and pR (also known as PR) and turning on cI transcription from pM. Repressor blocks transcription of late genes indirectly, by preventing transcription from gene Q, whose product turns on late gene transcription of gene N, whose product antiterminates early gene transcription.

promotes its own transcription. This is accomplished by specific binding to a cluster of three repressor-binding sites at each of the two operators. Late genes determining cell lysis and virion components are not expressed in the lysogen because their transcription is turned on only when the Q protein antiterminates the pR0 transcript, and the Q gene is transcribed from pR.

The lysogenic state is quite stable. About once in 104 cell divisions, lytic development is spontaneously activated, and the cell lyses and liberates phage. In many phages (including l), lytic development can be induced experimentally. The most convenient method of induction is to heat a strain that harbors a prophage with a ts mutation in the cl gene, so that the repressor is thermosensitive and denatures at high temperature. A more natural method is to treat the cell with various agents (such as ultraviolet light) that activate the bacterial SOS system.

The SOS system is a global regulatory system responsive to DNA damage. The basic circuitry is shown in figure 7-8. When DNA is damaged by irradiation or other means, single-strand oligonucleotides are formed as breakdown products. These activate a coprotease activity of the RecA protein (which also mediates homology-dependent recombination). This coprotease activity promotes cleavage of a master control protein, LexA. RecA is considered a copro-tease rather than a protease because the LexA protein can undergo spontaneous cleavage (autodigestion) under nonphysiological conditions, such as extreme pH, in the absence of RecA. LexA represses, directly or indirectly, a large number of genes whose products facilitate repair and recovery from DNA damage. So when LexA is cleaved in response to DNA damage, these proteins are synthesized. Many phage repressors (including 1's) mimic LexA in their susceptibility to coproteolysis by activated RecA. When repressor is thus inactivated, lytic development ensues in almost every cell of a treated culture. Spontaneous phage production is RecA-dependent and therefore presumed to result from rare sporadic DNA damage, such as may occur when a replication fork occasionally stalls.

excises.

Figure 7-8 The bacterial SOS system and its relation to induction of lytic development. DNA damage causes the disintegration of some DNA to smaller degradation products, of which single-stranded DNA is especially effective. Single-stranded DNA activates the RecA protein to a form RecA* whose coprotease activity accelerates the spontaneous cleavage of the LexA protein into two inactive polypeptides. The normal function of LexA is to repress (either directly or indirectly) the transcription of damage-inducible (din) genes of the host, many of which function in DNA repair. The repressor proteins of many phages (including l) mimic LexA, and their cleavage is likewise stimulated, inducing lytic development.

Figure 7-8 The bacterial SOS system and its relation to induction of lytic development. DNA damage causes the disintegration of some DNA to smaller degradation products, of which single-stranded DNA is especially effective. Single-stranded DNA activates the RecA protein to a form RecA* whose coprotease activity accelerates the spontaneous cleavage of the LexA protein into two inactive polypeptides. The normal function of LexA is to repress (either directly or indirectly) the transcription of damage-inducible (din) genes of the host, many of which function in DNA repair. The repressor proteins of many phages (including l) mimic LexA, and their cleavage is likewise stimulated, inducing lytic development.

The repressor concentration is delicately poised. It is sufficient to control phage gene expression, but too much repressor impedes induction. Excess repressor synthesis is avoided because, although repressor stimulates transcription from pM at moderate concentrations, high repressor concentrations repress transcription from pM.

If a lysogenic cell is externally infected by another phage of the carried type, the repressor present in the cell binds to the operator sites on the entering phage and prevents it from initiating lytic development. The lysogenic cell is said to be immune. Repressor (and hence immunity) are specific. A cell lysogenic for l can be lytically infected by unrelated phages such as Mu-1, and vice versa. Among the natural relatives of l, there are a variety of repressor types; thus l and 434, which insert at the same chromosomal site, make different repressors, and each phage makes plaques on lysogens of the other.

The circuitry has variations, even among close relatives of l. Phage P22, for example, encodes, in addition to a repres-sor, an antirepressor protein that can bind to repressor and neutralize its activity. Antirepressor synthesis is turned off in lysogenic cells by a "maintenance" protein, Mnt, which represses transcription of the antirepressor gene. Phages such as P1 that lysogenize by plasmid formation also control most of their genes by repression. Their problem is inherently more complex than that of l or Mu-1, because the lysogen must express not only the repressor gene but also those genes needed for plasmid replication and partitioning.

Although there are many examples of latency among eukaryotic viruses, there are few close analogues of lyso-geny. Retroviruses, for example, insert their DNA and can replicate as part of a host chromosome, but they carry out a productive cycle of infection while inserted. When retroviruses are carried permanently in an inactive state, it is generally because either the virus has mutated to a defective form or the particular host cell type is unable to support a complete infectious cycle. The closest thing to lysogeny occurs with some of the nonintegrated viruses, such as papilloma and members of the Herpes family. In both these cases there seems to be a programmed potentiality either to be carried as a plasmid (analogous to P1) or to initiate a productive infection that culminates in virus liberation and cell death. In some of the herpesviruses this occurs in nondividing nerve cells, where the plasmid form need not replicate to persist; however, Epstein-Barr virus can establish a carried state in replicating lymphocytes.

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