trypanosome antigenic variation

Question 1

african trypanosome lifecycle

Answer 1

• tse tse fly bites infected reservoir host
• uptake of trypomastigote
• → procyclic trypomastigote
• → epimastigote
• → metacyclic trypomastigote in salivary glands
• injected into mammal upon biting
• long slender trypomastigote
  
  • replicating bloodstream form

Question 2

quorum sensing in trypanosomes

Answer 2

• to assess population levels
• prevents host death
• high population levels:
  
  • long slender trypomastigote forms short stumpu trypomastigote
  • replicating to non-replicating
  • can differentiate into procyclic form
• common in parasites that have coevolved with host

Question 3

immune evasion

Answer 3

• extracellular parasite - immune evasion is key
• involves antigenic coat switching
• different coats required in the fly and the mammal

Question 4

infection cycles

Answer 4

• periodic fluctuation in number of t. brucei in patient’s blood
• patient produces antibodies to eliminate parasite
• a few parasites switch coats so they survive and numbers increase again
• parasite relies on ability to change coat more quickly than the host can mount an immune response

Question 5

VSG coats

Answer 5

• variant surface glycoprotein
• analogous to giardia VSPs
• single variant of VSG densely packed on surface of bloodstream trypomastigotes at any one time
• shields shorter invariant surface molecules and receptors from host

Question 6

VSG components

Answer 6

• variable N-terminus
• conserved C-terminus
• GPI anchor attaching to plasma membrane
  
  • essentially lipid within a lipid
  • mobile, fluid, dense

Question 7

transferrin receptor

Answer 7

• example of a hidden receptor
• steals iron from the host for the parasite’s use
• receptor hidden by VSGs
• transferrin binds so parasite can steal it
• also occurs with hexose transporters

Question 8

VSG amino acid sequence

Answer 8

• highly variable sequence for evasion
• tertiary structure very similar and highly conserved
  
  • selection pressure for diversiy and conservation of shape
  • as coat is switched old and new VSGs mix
  • must be able to sit close together so need similar shape

Question 9

flagellar pocket

Answer 9

• hides bulk of invariant receptors
• inaccessible to antibodies
• site of endocytosis in t. brucei
  
  • endocytic vesicles involved in nutrient uptake bud off from here
• very high rates of endocytosis in t. brucei
  
  • entire VSG coat internalised every 12 minutes

Question 10

VSG internalisation

Answer 10

• recycles VSGs (doesn’t destroy)
• coat cleaning mechanism
• antibodies or C3b fragments attached to VSGs are stripped off
• VSG returned to surface
• in low titre antibody environmentsthis is enough to avoid elimination
• will be overwhelmed at high titres

Question 11

hydrodynamics and coat cleaning

Answer 11

• hydrodynamic forces help remove antibodies
• attached antibody may act as a sail on the aprasite surface
• Ab-VSG complex swept more quickly to the flagellar pocket than unbound VSGs

Question 12

VSG expression sites

Answer 12

• where the single active VSG is transcribed
• telomeric polycistronic transcription unit
• transcribed as one but contains other genes
• VSG gene always at telomeres
• contains polymorphic ESAGs as well as VSG
• very high transcription rates
• 15-20 per cell
  
  • highly variable structures

Question 13

ESAGs

Answer 13

• expression site associated genes
• mainly receptor molecules
• most functions unknown
• may play a role in host adaptation

Question 14

bloodstream vs metacyclic VSGs

Answer 14

• different VSGs better suited to either stage
• their VSG expression sites fulfil different needs

Question 15

bloodstream VSG expression sites

Answer 15

• ~20 sites
• large polycistronic unit
• 300-400 VSGs can be accessed
• efficient gene conversion of 70bp arrays
• maximal diversity

Question 16

metacyclic VSG expression sites

Answer 16

• ~25 sites
• monocistronic
• inefficient/no gene conversion
  
  • no/few 70bp repeats
• maximal diversity at population level

Question 17

silent VSGs

Answer 17

• 1000s of silent VSG genes at any time
• internal chromosome arrangement in arrays at subtelomeres
• ~1500 VSG genes and pseudogenes in T. brucei
• ~10% of total genome
• silent VSG cassettes can be switched in to expression sites
  
  • different mechanisms depending on stage of infection

Question 18

duplicate gene conversion of silent VSG cassettes

Answer 18

• occurs due to flanking homology allowing for VSG gene switching
• copy of a silent VSG gene can be moved over to active expression site
• throw out copy already there
• uses homologous recombination with 70bp homologous regions upstream and conserved region downstream
• important during chronic infection

Question 19

mechanisms of VSG gene switching

Answer 19

• duplicative gene conversion
• segmental gene conversion
• telomere exchange
• ES switching

Question 20

segmental gene conversion

Answer 20

• similar to duplicative
• parts of other genes are moved in creating a novel, chimeric VSG gene

Question 21

telomere exchange

Answer 21

• exchange of VSG genes at two telomere ends
• limited access to VSG gene pool

Question 22

ES switching

Answer 22

• involves silencing transcription at one expression site and activating it at another
• only allows switching among the 15 or so expression sites
• more important during early infection when host has not been immunised to a range of VSGs
• access to small VSG gene pool

Question 23

T. brucei genome

Answer 23

• only 10% of VSG genes are in tact
  
  • others are pseudogenes
• mosaic VSGs from segmental gene conversion are composed of pseudogenes
• allows parasite to create new coats at late stages of infection
• too difficult for the parasite to maintain all these genes active

Question 24

coat switching hierarchy

Answer 24

• switching to certain VSGs is preferred
• unknown mechanism
• useful for there to be some level of homogeneity within a population to prevent exposing the host to too many variants