Lecture 3: Innate Immunity and Trypanosome Lytic Factor (TLF) Flashcards
What are the two morphological T. brucei subspecies found in Africa that cause distinct disease patterns in humans?
T. brucei gambiense (chronic infection)
T. brucei rhondesiense (acute infection)
What is Nagana and what causes this disease?
Cattle disease caused by T. b. bruci, T. congolense and T. vivax
Which trypanosome species causes the disease surra?
T. evansi
Which trypanosome parasites are humans susceptible and resistant to?
Susceptible to infection by T. b. gambiense and T. b rhodesiense
Resistant to infection by T. b. brucei, T. congolense, T. vivax and T. evansi
Why are humans resistant to infection by T. b. brucei, T. congolense, T. vivax and T. evansi?
These trypanosomes are susepctible to lysis by Trypanosome lytic factors (TLFs)
True or false: TLFs are innate immune effectors?
True (they are not complement proteins or antibodies though)
Why are humans susceptible to infection by T. b. gambiense and T. b rhodesiense?
They are resistant to killing by TLFs (have developed mechanisms that counteract the lytic effect of TLF)
What is needed for the lytic activity of TLFs?
uptake by endocytosis and trafficking to the lysosomal compartment within the trypanosome
what effect does normal human serum (NHS) have on T. b. brucei?
cell swelling and lysis
which trypanosoma brucei subspecies can be killed by serum from humans and some primates?
Trypanosoma brucei brucei
(not rhodesience or gambiense)
How was it identified that the TLF was a high density lipoprotein?
serum from patients with Tangier disease (characterised by deficiency in HDL) had no trypanocidal activity due to absence of TLFs
What are the two distinct lipoprotein complexes present in human blood that exhibit trypanosome lytic activity?
TLF1 and TLF2
What are the components of TLF1?
- Apo-lipoprotein A-1 (APOA1)
- Haptoglobin-related protein (Hpr)
- Apo-lipoprotein L-1 (APOL1)
- Trace amounts of apo-lipoprotein A-II and paraoxonase
What are the components of TLF2?
- Apo-lipoprotein A-1 (APOA1)
- Haptoglobin-related protein (Hpr)
- Apo-lipoprotein L-1 (APOL1)
- IgM
What is APOL1?
a key component of HDLs and is a cofactor of lecithin cholesterol acyltransferase (which recruits free cholesterol to the lipid core of the HDL particles and transport cholesterol to the liver for secretion or re-use)
Which human TLF is described as a lipid-poor protein complex?
TLF2
What is the role of the haptoglobin-related protein in the TLFs?
binds Hb but is not removed from blood circulation via receptor-mediated process in liver (not recognised by receptors) and plays an important role in promoting TLF1 uptake by binding of Hpr-Hb to T. b. brucei TbHpHbR receptor
Which TLF component is necessary for the lysis of the parasites (the lytic component)?
APOL1
What was the result of TbHpHbR gene knockout parasites?
more resistant to lysis - hence the receptor is required for efficient lysis by mediating the uptake of TLF
What are two unique characteristics of APOL1?
has N-terminal signal sequence meaning it is secreted from cells
it is exclusively associated with HDL particles int he circulation
What are the three main domains of the APOL1 protein?
- N-terminal pore-forming domain (contains10 helices homologous to bacterial colicins that forms pores in the membrane)
- Membrane-addressing domain (inserts into membrane)
- C-terminal SRA-interacting domain
Describe the mechanism of APOL1 induced lysis of T. b. brucei
- endocytosis of APOL1 contaning HDL particles by TbHpHbR
- low endosomal pH promotes conformational change in APOL1 that exposes the pore forming domain and allows membrane insertion
- this leads to influx of chloride ions and osmotic imbalance that cause lysosome swelling and cell death
How may the fact that T. b. brucei is deficient in haem biosynthesis result in TLF arming mechanism?
parasite requires haem from Hb so causes erythrocyte damage to release haem into the plasma which is then taken up into the parasite by binding to Hpr and endocytosis via TbHpHbR receptor
What is the role of TbKIFC1?
has cholesterol-trafficking activity that contributes to high plasma membrane fluidity
what is the effect of KIFC1 knockdown in cells on Ab-VSG clearance and TLF2 internalisation?
- depletion of KIFC1 = cell lysis occurred at a slower rate that WT cells
–> Ab-VSG complexes moved slower to flagellar pocket
–> clearance of Ab-VSG complexes is likely a driving force for TLF2 internalisation and trypanosome killing (TLF2 bound IgMs bind to VSG and are internalised at the flagellar pocket to introduce APOL1 into the parasite for killing)
What is SRA?
Serum resistance Associated gene expressed by T. b. rhodesiense that is a truncated VSG protein lacking surface-exposed loops and is not expressed on the surface of parasite
What is the relationship between the location of TLF and SRA in parasite?
TLF and SRA were shown to co-localise in endosomes and lysosomes
how is it believes that SRA expression in T. b. rhodesiense confers resistance to lysis by APOL1?
TLF internalised through flagellar pocket
- SRA is able to block the inserting of APOL1 into the endosomal membrane (blocking swelling of parasite and subsequent death)
What was the result of SRA overexpression in the TLF1 sensitive T. b. brucei parasite?
reduction in lysis compared to the parasites without the SRA protein
Where was the T. b. rhodesiense SRA gene found in the parasite genome?
It is an expression-site associated gene (ESAG) found in the expression site of a truncated VSG
What three mechanisms does T. b. gambiense use to evade lysis by TLF?
mutation in the HpHb receptor results in inactivation and reduced uptake of APOL1
increase degradation of APOL1 due to lower pH in early endosome and earlier endosomal protease activity = faster degradation of APOL1
Prevention of APOL1 membrane insertion - TgsGP (T. b. gambeinse specific glycoprotein) a shortened VSG that is GPI anchored in lysosomes causing stiffening and increasing curvature of membrane to prevent APOL1 insertion
What was found in the African genome population that conferred protection against T. b. rhodesiense infection?
Two genetic polymorphisms in the human APOL1 gene:
- G1 allele (2 amino acid substitutions)
- G2 allele (2 amino acid deletion at C-terminus)
How was it identified that APOL1 deficiency is linked to T. evansi infection?
T. evansi does not normally cause infection in humans since sensitive to TLF
but patient infected with T. evansi was found to have a frameshift mutation in APOL1 gene that meant they were not producing functional APOL1 and were therefore susceptible to T. evansi infection
True or false: T. b. gambiense is resistant to TLF by expression of SRA?
False: T. gambeinse does not express SRA (unlike T. rhodesiense) but has 3 independent complementary mechanisms of TLF lysis evasion
