Lecture 10 - Bacteriophage Lambda λ

What family does Bacteriophage λ belong to?

Siphoviridae (Greek: siphon = tube)

What host does Bacteriophage λ infect?

E. coli

Is Bacteriophage λ enveloped or non-enveloped?

Non-enveloped (naked)

What type of genome does λ phage have?

Linear double-stranded DNA (dsDNA), monopartite, 48 kb

Describe the physical structure of Bacteriophage λ.

Icosahedral capsid (63 nm) with a long, flexible, non-contractile tail (135 nm) and a terminal fiber.

What kind of sequence allows circularization of the genome?

cos (cohesive end) sequences

What enzyme seals the circularized genome?

Host DNA ligase

What happens when bacteriophage λ injects dsDNA into e.coli cell.

The genome circularizes by cos sequences which are adhesive ends

What does it mean when we say Bacteriophage λ is temperate phage?

It can integrate its genetic material into the host bacterial chromosome, becoming a prophage, allowing it to replicate alongside the host without immediately causing cell death (lysogeny), but can also switch to a lytic cycle and replicate rapidly, leading to cell lysis

What protein mediates attachment to the host?

J protein (trimer at tail tip)

What receptor does λ use on E. coli?

LamB (normal role is transport of maltose-inducible into cell as a porin)

What triggers genome injection into the host?

J protein interaction with LamB causes rearrangement of terminal fiber which allows genome injection

What does PL stand for and what genes does it primarily express?

Left Promoter and N (antiterminator), CIII,

What does PR stand for and what genes does it primarily express?

Right Promoter and cro, CII, O, and P

What does PRM stand for and what genes does it primarily express?

Promoter for Repressor Maintenance and CI

What does PRE stand for and what genes does it primarily express?

Promoter for Repressor Establishment and cro

What does PI stand for and what genes does it primarily express?

Promoter for Integrase and xis, Int.

Which mRNAs are first made after circularization?

N mRNA from PL and cro mRNA from PR

What is the function of the N protein?

Antiterminator that promotes early gene expression

What sequences does N protein bind to?

NUT (N utilization) sequences

What proteins are involved in replication initiation?

O and P proteins

What does the O protein do?

Binds to the replication origin (ori) and melts AT-rich region

What does the P protein do?

Recruits E. coli helicase (DnaB) and host DNA polymerase to copy the phage DNA

What replication mechanism is used early on?

θ (theta) replication which is bidirectional and produces progeny circles

What replication mechanism is used during lytic growth?

Rolling circle replication which creates multimeric dsDNA concatemers

What is the role of the Q protein?

Antiterminator that enables late gene expression (transcription pf lysis, head, tail proteins)

What are the two lysis genes

R, transglycosidase and S, holin

What is the function of the R gene?

Transglycosidase that degrades the bacterial peptidoglycan

What is the function of the S gene?

Holin that forms pores in the membrane for R to access peptidoglycan

When does the λ enters lysogeny

When there is low nutrients and cell is metabolically sluggish/stationary phase

What happens under low nutrient conditions?

Less HfLA so CII is not degreaded. CIII stabilizes CII through protection → lysogeny favored

What happens under high nutrient conditions?

More HfLA so CII is degraded. cell favours lysis

What do CII and CIII proteins do?

CII promotes CI and integrase (Int) expression; CIII stabilizes CII

What does CI and Int do?

Int is the integrase enzyme necessary for λ DNA integration into the host chromosome and CI is the repressor protein

What is the prophage?

Integrated λ DNA in host genome

What is the only phage gene expressed during lysogeny?

CI repressor

How does CI repress lytic genes?

Binds operators near PR and blocks cro transcription

What else does CI provide besides repression?

Superinfection exclusion from other phages

How does CI maintain lysogeny?

CI dimers help form the CI repression loop which pulls the operators together in the middle 

What triggers the switch from lysogeny to lysis?

Host DNA damage (e.g., UV light) triggering the SOS response

What happens to CI during SOS response?

CI is cleaved, relieving repression

What does Cro do?

Binds OR operators to shut down PRM and CI expression and moves to lytic cycle

What genes mediate excision of prophage DNA?

Xis (excision), Int(intgrase), and IHF (integration host factor)