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Lecture 4

The document summarizes the replication mechanisms of lambda phage, specifically comparing the regulatory mechanisms that determine whether the phage follows the lytic or lysogenic cycle. It explains that early transcription can establish the lytic cycle by producing the Cro and N proteins. The N protein allows transcription to continue through the middle and late stages to produce phage particles. Alternatively, the cI repressor protein establishes lysogeny by binding operator sequences and preventing transcription of the lytic genes. The ultimate fate depends on competition between Cro and cI for binding sites on the operator.

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MuhammadFakhriAimi
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63 views

Lecture 4

The document summarizes the replication mechanisms of lambda phage, specifically comparing the regulatory mechanisms that determine whether the phage follows the lytic or lysogenic cycle. It explains that early transcription can establish the lytic cycle by producing the Cro and N proteins. The N protein allows transcription to continue through the middle and late stages to produce phage particles. Alternatively, the cI repressor protein establishes lysogeny by binding operator sequences and preventing transcription of the lytic genes. The ultimate fate depends on competition between Cro and cI for binding sites on the operator.

Uploaded by

MuhammadFakhriAimi
Copyright
© Attribution Non-Commercial (BY-NC)
Available Formats
Download as PPT, PDF, TXT or read online on Scribd
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Learning Outcomes:

At the end of this lecture, students will be able to: Explain replication mechanism in lambda phage Differentiate the regulatory mechanism involved in the lytic and lysogeny cycle

Events leading to lytic cycle


transcription occurs in 3 stages ; early, middle, late. Early transcription establishes the lytic cycle After infection of a sensitive host (bacteria), early transcription proceeds from major promoters situated to the left (PL) and right (PR) of the repressor gene (cI). As host RNA polymerase transcribes the lambda genome, two proteins are produced: cro (for "control of repressor and other things") from PR N from PL If the lytic pathway is followed, transcription of the remainder of the viral genes occurs, and assembly of the virus particles will occur. The N protein functions in this process, ensuring that transcription does not terminate.

transcripts from PL and PR stop at termination site tL and tR1. switches from early to middle stage transcription by antitermination. The N gene product (antiterminator protein) switches the transcription process. The N gene product (expressed from PL) interacts with RNA polymerase and stops the action of host termination protein, allowing it to ignore the stop signal (tL & tR). This allows the PL and PR transcripts to extend into genes required for middle stage (ie. red, O, P genes-DNA synthesis) cro gene (for control of repression and other things) encodes an antirepressor protein- inhibits transcription of the repressor operon, including the cI gene the cro gene product accumulates sufficiently to turn off the cI gene (coding for the lambda-repressor) and firmly establish a lytic cycle

There is also a second antitermination system involved in the lytic cycle Transcription of the right operon terminates again, somewhat downstream from a gene designated Q. In cells that enter a lytic cycle, the Q gene product accumulates and functions as another antiterminator, allowing the late operon (actually an extension of the right operon) to transcribe genes needed for completion of the lytic cycle, including those for formation of the phage head and tail.

The Q product anti terminates the short PR transcript and so extending the transcript into the late genes (S, R) and crosses into the Cos region releasing many mature phages. N and Q are genes responsible for positive regulation, which are essential for phage growth and plaque formation. *However, an N- phage is able to produce a small plaque if the termination site tR2 is removed by a deletion termed nin (N independent) N- nin

N
att

N
cI cro

N
cII O P Q SR AJ

int

gam red xis cIII N

Pint

tL1

PL

PRM PR tR1 PRE

tR2 PR t6S tR3


6S RNA

N protein
CIII Recombination proteins

Cro
CII Q protein

Replication proteins

Repressor protein, cI
The path to lysogeny occurs differently, involving a cI gene product. cI gene product is a lambda repressor protein PRE (a repressor establishment promoter) expresses the repressor gene, cI, to establish lysogeny. Its function is to bind to operator sequences and prevent transcription of both the left and right operons. Expression of cI will induce the phage genome to integrate into the host genome. When integrated, only the cI will be produced, it tends to be self sustaining so as to maintain the lysogenic state. PRM (a repressor maintenance promoter) expresses the repressor gene, cI, to maintain lysogeny. Transcription of cI is enhanced by high levels of cII protein The cI gene product controls the transcription process. (In a lysogen, the repression results in immunity of host to superinfection)

Lambda decision: Lysogeny or lytic phase?


The virus adopts the lytic or lysogenic path early following infection of the host bacterium. The fate of the viral genetic material is governed by a competition between the cro and cI proteins. Both can bind to the same operator region. The region has three binding zones--cro and cI occupy these zones in reverse order. The protein, which is able to occupy the preferred regions of the operator first, stimulates its further synthesis and blocks synthesis of the other protein.

Lytic genes are repressed by cI. cI is repressed by cro. cII (a transcription activator)-increases rate of initiation at PRE induces transcription of cI. cII levels are controlled by presence of nutrients. The cII protein is constantly degraded by proteases present in the cell. If levels of cII remain below a critical level, transcription from PR and PL continues and the phage undergoes a productive replication cycle which culminates in lysis of the cell and the release of phage particles. If the concentration of cII protein builds up, transcription of cI is enhanced phage goes through lysogeny cycle Physiological stress and particularly ultraviolet irradiation of cells results in the induction of a host cell protein, RecA. This protein, whose normal function is to induce the expression of cellular genes which permit the cell to adapt to and survive in altered environmental conditions, cleaves the cI repressor protein. Phage goes through lytic cycle

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