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Enterobacteria phage λ (lambda phage) is a temperate bacteriophage that infects Escherichia coli. Once the phage is inside its host, it may integrate itself into the host's DNA. In this state, λ is called a prophage and stays resident within the host's genome, without apparent harm. Thus the prophage is duplicated with every cell division of the host. This is referred to as the "lysogenic pathway". The DNA of the prophage that is expressed in that state codes for proteins that look out for signs of stress in the host cell. Stress can be from starvation, poisons (like antibiotics), or other factors that can damage or destroy the host. At that point, the prophage reactivates, excises itself from the DNA of the host cell and enters its lytic cycle. The reactivated phage takes apart the host's DNA and produces large amounts of its own mRNA, so as to produce prodigious amounts of phage units. When all resources of the host are depleted from building new phages, the host cell is lysed (the cell wall is broken down), and the new phages are released. The integration of phage λ takes place at a special attachment site in the bacterial genome, called attλ. The sequence of the att site is called attB and consists of the parts B-O-B', whereas the complementary sequence in the circular phage genome is called attP and consists of the parts P-O-P'. The integration itself is a sequential exchange (see genetic recombination) via a Holliday junction and requires both the phage protein Int and the bacterial protein IHF (integration host factor). Both Int and IHF bind to attP and form an intasome, a DNA-protein-complex designed for site-specific recombination of the phage and host DNA. The original BOB' sequence is changed by the integration to B-O-P'-phage DNA-P-O-B'. The phage DNA is now part of the host's genome. Repressor The repressor found in the phage lambda is a notable example of the level of control possible over gene expression by a very simple system. It forms a 'binary switch' with two genes under mutually exclusive expression. In the following paragraphs, we will write genes in italics and their associated proteins in roman. For instance, cI refers to the gene, while cI is the resulting protein encoded by that gene. The lambda repressor gene system consists of (from left to right on the chromosome): The lambda repressor is a dimer also known as the cI protein. It regulates the transcription of the cI protein and the Cro protein. The life cycle of lambda phages is controlled by cI and Cro proteins. The lambda phage will remain in the lysogenic state if cI proteins predominate, but will be transformed into the lytic cycle if cro proteins predominate. The cI dimer may bind to any of three operators, OR1, OR2, and OR3, in the order OR1 > OR2 > OR3. Binding of a cI dimer to OR1 enhances binding of a second cI dimer to OR2, an effect called cooperativity. Thus, OR1 and OR2 are almost always simultaneously occupied by cI. However, this does not increase the affinity between cI and OR3, which will be occupied only when the cI concentration is high. Lifecycle Lytic Lifestyle [xis and int regulation of insertion and excision] Lysenogenic (or Lysogenic) Lifestyle Induction [Control of phage genome excision in induction] Protein Function Overview cro; Transcription inhibitor, binds OR3, OR2 and OR1 (affinity OR3 > OR2 > OR1, ie. preferentially binds OR3). At low concentrations blocks the RM promoter (preventing cI production). At high concentrations downregulates its own production through OR2 and OR1 binding. cI; Transcription inhibitor, binds OR1, OR2 and OR3 (affinity OR1 > OR2 > OR3, ie. prefferentially binds OR1). At low concentrations blocks the R promoter (preventing cro production). At high concentrations downregulates its own production through OR2 and OR3 binding. Also inhibits transcription from the L promoter. Succeptable to cleavage by RecA cII; Transcription activator, binds cIII. Activates transcription from the antiq, RE and I promoters. Low stability due to succeptability to cellular proteases (especially in healthy cells and cells undergoing the SOS response), slightly stabilised by binding to cIII. cIII; cII binding protein, protects cII from degradation by cellular proteases. N; RNA binding protein and RNA polymerase cofactor, binds RNA (at Nut sites) and transfers onto any oncoming RNApol. Alters the recognition of termination sites, so normal RNA polymerase termination signals are ignored and RNA synthesis continues into distal phage genes. Q; DNA binding protein and RNApol cofactor, binds DNA (at Qut sites) and transfers onto any oncoming RNApol. Alters the recognition of termination sequences, so normal ones are ignored and special Q termination sequences are effective instead. xis; excisionase and integrase regulator, manages excision and insertion of phage genome into the host's genome. int; integrase, manages insertion of phage genome into the host's genome. In Conditions of low int concentration there is no effect. If xis is low in concentration and int high the n this leads to the insertion of the phage genome. If xis and int have high (and approximately equal) concentrations this leads to the excision of phage genomes from the host's genome. A, B, C, D, E, F, Z, U, V, G, T, H, M, L, K, I, J Shown on diagram as head and tail, A-F code for phage head genes, Z-J code for phage tail genes. The order shown here is as found on the genome, reading in a clockwise direction; structural proteins, self assemble with the phage genome into daughter phage particles. S, R Shown on diagram as lysis. The order shown here is as found on the genome, reading in a clockwise direction; lysis promoters, cause the host cell to undergo lysis at high enough concentrations. OP Shown on diagram as O replication P; DNA replication promoter, promotes the specific replication of only the phage genome. sib not a protein, but a vital conserved DNA sequence; Forms a stable hairpin loop structure in transcribed mRNA. Attracts degradation of mRNA by RNAaseIII. attp not a protein, but a vital conserved DNA sequence; point of action of int and xis in insertion and excision of the phage genome into the host's genome. Corresponding attb found in the host's genome at the point of insertion. | |||||||
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