Case study of bacterial pathogenesis

Question 1

Listeria monoctogenes

Answer 1

Can be ingested by contaminated food. All listeria spp are motile at low temperatures, facultative anaerobes nonsporulating. Only L.monocytogenes is pathogenic. Temperature-dependent expression of virulence factors: RNA thermometer. RNA blocks the ribsome, but at a certain temperature the RNA melts, allowing the ribosome to bind. Actin is involved in dispersing the bacteria and some sort of movement (comet tails)

Question 2

Actin

Answer 2

Globular actin is a soluble monomer that polymerises to form actin filaments. Filaments exhibit polarity. polymerisation is favoured at one end

Question 3

using mutants to identify bacterial factors involved in tail formation

Answer 3

two antibiotic resistant markers, one within transposon and one on plasmid. Transform bacteria in media culture containing both resistant genes. culture. Transposition of resistance cassette onto chromosomes. select on media containing only resistance on transposon, pool of individual deletion mutants. Take mtants and infect cells with each mutant. Identify the mutants the do not form tails. identify genes in question

Question 4

ActA

Answer 4

wildtype ActA restores tails. ActA is a surface protein, contains a signal sequence in its N-terminal region that directs secretion. if signal sequence is deleteted there is no tail formation. ActA bind F-actin, multiple regions important for interaction between F-actin and ActA in the N-terminal domain. ActA is a surface protein anchored to the membrane by this C-terminal region. if this region is deleted there is no tail formation.
ActA stimulates rapid actin assembly only in the presence of the Arp2/3 complex

Question 5

Listeria ActA mutants

Answer 5

Hopping- discontinuous tail, theres a problem with polymerisation
Dancers- tail forms on ‘long side’, there’s a problem with location

Question 6

Listeria ActA

Answer 6

discoverly revealed a family of cellular ptoetins called ‘nucleation (polymerisation) promoting factors’. They share amino acid similarity to ActA but are found in cells. Involved in stimulating cellular Arp2/3 dependent actin assembly. Like actA they bind and activate Arp2/3 complex. Important in key cellular signalling mechanisms relevant to cell movement and receptor signalling