Bacteriophage Lambda Flashcards
What are the minimal functions of a bacteriophage?
- Protection of nucleic acid
- Delivery of nucleic acid
- Conversion of infected cell to produce phage
- Release of phage
Briefly describe the steps of the lytic cycle.
1) Phage attaches to bacterium, with the tail recognising receptors on the cell surface
2) DNA injected into bacterium (nucleic acid may have to be unpackaged if filamentous)
3) Enzymes for DNA synthesis are made and replication begins (early development)
4) Genomes, heads and tails are made, and the DNA is packaged into the heads and the tails attached (late development)
5) Lysis occurs; the cell is broken to release progeny phages. Some filamentous phage ‘bud off’ instead of causing cell lysis.
Briefly describe the steps of the lysogenic growth cycle.
1) Similarly to the lytic cycle, the phage attaches to the bacterium and injects its DNA
2) Phage DNA is integrated into the bacterial genome
This is a slower process than the lytic cycle but less risky.
What is a virulent phage?
Always enters the lytic cycle (can not undergo lysogeny), and so these strains have often incurred mutations as entering the lysogenic cycle can be beneficial for phage survival.
What is a temperate phage?
Has the ability to enter both the lytic and lysogenic cycles.
What is a lysogen?
A bacterial cell containing DNA with incorporated phage DNA in it.
What is a prophage?
Refers to the phage DNA within the bacterial chromosome.
What is immunity in terms of the lysogenic cycle?
Immunity refers to a lysogen now being immune to infection of phage particles from the same species (second infection wont lead to lytic cycle). This is as repressor proteins have been translated that are specific to the operon on that species of phage DNA.
What is induction?
When the lytic cycle is induced from the lysogenic. This occurs due to the cleaving of the dimeric repressor protein, or by changing the concentration of the CII protein.
Describe how conditions affect whether the lytic or lysogenic cycle is entered.
Conditions favoured by lysogenic: low nutrient levels and high MOI (multiplicity of infection) i.e. greater number of phage than bacterial cells. This is as some phage will not be able to find cells to infect and thus will be exposed to the environment/nucleases. Low nutrient levels may cause the bacterial cell to go into a dormant state so lytic cycle cannot happen as can’t take over cellular machinery.
Conditions favoured by lytic: low MOI i.e. more bacterial cells than phage particles, as phage progeny will have lots of cells to infect.
How can you distinguish if a phage is virulent or temperate experimentally?
By growing phage on a bacterial lawn. If the phage is virulent then the lawn will be clear as it made ‘holes’ in the cells. If the phage is temperate then there will be an area that is clear (due to initial lytic cycle), but when the lysogenic cycle is induced the cells will grow over this clear area to make it turbid.
Why is it important that the bacteriophage lambda DNA forms a circle when in the bacterial cell?
It is needed as the promoter for the late genes (structural components) is before the S region where the circle is joined, and the late genes are after this region. If it was not made into a circle then the head and tail of the phage would not be made (would not be expressed as promoter in wrong position). The circularisation allows the formation of the operon, but this is not needed when the phage is not in the cell.
Describe the lytic cycle cascade in bacteriophage lambda.
1) Host RNA polymerase initially controls the lytic cycle and transcribes the early genes. From the first promoter to the first termination site, a set of genes are transcribed.
2) These genes are translated to regulatory proteins, and in bacteriophage lambda this protein is an antitermination factor which allows the RNA pol to carry on past the termination site (stops RNA pol from recognising terminator sequence).
3) The RNA pol is then able to transcribe the delayed early expression genes, and again regulatory proteins are subsequently translated. Also structural genes begin to be transcribed.
4) Transcription of the late genes is then allowed, and phage components can then be built.
Groups of genes are expressed in an ordered manner.
In bacteriophage lambda, the regulatory protein produced during lytic development is the antitermination factor. What other regulatory proteins are there in different phages and how do they allow transcription of the delayed early and late genes?
- A new RNA polymerase may be produced (different from the initial host polymerase) that can recognise the 2nd promoter and transcribe this next section of the DNA sequence.
- Sigma factor may be produced, which changes the conformation of the host RNA polymerase so it is able to recognise the next phage promoter sequence.
What is pN?
The N protein, the result of the transcription of the N gene (part of the immediate early genes transcribed in lytic development). It is an anti-termination factor and so allows the delayed early genes to be transcribed, as it interacts with RNA pol and stops it from recognising the first terminator sequence. Permits transcription past N and cro (early genes).
Where is the cI gene located and what does it encode?
It lies between the left and right promoters (lambda has left and right transcription units). It encodes for the repressor, and has 2 promoters (Prm and Pre).
