Lecture 13

Question 1

What is a virus?

Answer 1

An infectious agent that must grow or reproduce inside a host cell

Considered obligate intracellular parasites

Question 2

What are viruses composed of?

Answer 2

Nuclei acid, genetic material and protein coat (some also contain lipids in their coats)

Question 3

What is a bacteriophage?

Answer 3

Viruses that infect bacterial cells

Question 4

Features of phage lambda

Answer 4

• infects E.coli
• siphoviridae
• double stranded DNA genome (dsDNA)
• long flexible tail, icosahedral head

Question 5

Two phage lambda pathways

Answer 5

• lytic pathway
• lysogenic pathway

Question 6

Lambda and its two pathways

Answer 6

Yap
- phage finds an appropriate cell and uses its tail fibres to recognise receptors on cell surface
- Pham lambda is a TEMPERATE phage which means it can choose between two pathways
- lytic cell entrains a phage taking over the cell, making more of itself and causing cell lysis
- lysogenic it forms a stable relationship with e.coli and inject its DNA into the chromosome - forming a PROPHAGE - everything it reproduces the page is inherited vertically - phage can predict conditions and go into prophage induction (cell lysis) or lysogenic growth

Question 7

Lytic lifecycle definition

Answer 7

The phage lifecycle that results in lysis of the bacterial cell upon release of progeny phage

Question 8

Lysogenic life cycle

Answer 8

The phage lifecycle that results in stable carriage of the phage (prophage) within the host cell (lysogenic)

Question 9

Virulent phage definition

Answer 9

A phage that is only able to undergo replication via the lytic cycle

Question 10

Temperate phage definition

Answer 10

A phage that can replicate via either the lytic or lysogenic cyclones (e.g phage lambda)

Question 11

Lysogenic definition

Answer 11

A host cell that is harbouring a prophage during lysogeny

Question 12

Prophage definition

Answer 12

The latent form of a temperate phage that remains within the lysogen (e.g integrated into a host chromosome)

Question 13

Phage lambda lifecycle

Answer 13

Question 14

Step one in the formation of a bacteriophage

Answer 14

DNA injection and circularisation

• genome is linear in phage head
• following injection, the lambda genome circularises (staggered ends that are complementary )
• Aided by cohesive end sites (cos sites)

Question 15

Phage lambda genome in head vs in host

Answer 15

• linear in phage
• circularised in host

Question 16

Phage genome

Answer 16

Question 17

Lytic cycle: process of the phage assemble and release

Answer 17

1. Head and tail proteins are synthesised
2. DNA is packaged into heads
3. Tails are added
4. The is lysed and releasing new phage

Question 18

Lysogenic cycle: process of phage integration and maintainence

Answer 18

1. Lambda integrates
2. Prophage is stably maintained
3. Prophage passed to daughter cells

Question 19

In lysogeny how does integration of lambda DNA into host chromosome occur ?

Answer 19

• integration occurs between two sites
  = attP (attachment on phage)
  = attB (attachment on bacteria)
• there is a site-specific recombination requiring enzyme int (integrate)
• lambda is then linearised in host chromosome

Question 20

3 main questions to decide if lytic or lysogenic pathway

Answer 20

Question 21

The cascade of gene expression upon infection

Answer 21

Question 22

Very early gene expression events

Answer 22

• following injection, host RNA polymerase transcribes from two promoters
• PL (produces N). (Anti terminator)
• PR (produces Cro) (cro encodes the Cro protein. Expressed early in infection and the major player in establishing lytic growth)

N encodes an anti-terminator protein that enables transcription past 2 terminators resulting in early gene expression

Protein products for both possible pathways are produced early - but hasn’t committed yet

Question 23

Principle of anti-termination

Answer 23

Question 24

What are the three DNA-binding proteins of lambda

Answer 24

• Cro
• Cl
• CII

Question 25

What is Cro

Answer 25

A DNA-binding protein that represses transcription - Cro promotes the lytic cycle

Question 26

What is CII

Answer 26

A DNA-bidning protein that can activate or repress transcription - CI activates its own expression - represses genes requires for lytic cycle - maintains lysogeny

Question 27

What does CII do and Cro do

Answer 27

CII promotes lysogenic pathway Cro enables lytic pathway

Question 28

What protein is the deciding protein in lytic vs lysogenic - lots of protease

Answer 28

CII - host protease degrade CII - healthy cells produce high levels of protease - in actively growing cells, CII gets degraded - Cro protein wins the battle and the lytic cycle proceeds

Question 29

Status of CII and Cro in the late lytic cycle

Answer 29

CII is degraded Cro predominates - Cro represses expression of CI, all early genes then itself and the replication genes - Q is an anti-terminator that enables expression of late lytic genes (head,tail and lysis genes)

Question 30

What is the deciding protein - lytic vs lysogenic - less protease

Answer 30

- starved cells produce less protease - CII remains intact and stable - CII wins the battle over Cro and activates lysogenic pathway - CII wins the battle over Cro and activates lysogenic pathway

Question 31

Late lysogenic - status of CII

Answer 31

Not degraded CII is an activator protein that turns on: 1. Int resulting in integration of lambda into host chromosome 2. CI repressor, which activates own expression and binds to OL and OR to repress all other phage genes

Question 32

Process of lysogeny maintenance

Answer 32

- maintained as a prophage by CI - CI is a repressor of all genes but an activator itself - keeps phage genome ‘silent’ in bacterial chromosome until induction So CII activates int and Cl, which cause integration of phage and maintenance of lysogeny, respectively

Question 33

Summary of the decision.

Answer 33

Question 34

Answer 34

Maybe make into flash card