LAST MINUTE Flashcards
How do NK cells perform receptor mediated cytotoxicity?
Fas/FasL - target cell/NK cell
– results in caspase 8/10 activation leading to caspase cascade –> degradation of cellular proteins, DNA, and structural components, leading to apoptotic cell death
TRAIL/TRAIL R - v similar to Fas/FasL
– these seem more specialised to targeting cancer cells (express R-1, R-2)
– TRAIL-R decoys (eg R-3, R-4) exist on healthy cells, preventing apoptosis
TNF
– can bind TNFR1 and 2 to induce apoptosis
What are some examples of activatory receptors on NK cells?
CD16: a low affinity receptor for FC region of IgG - binds to opsonising antibodies to allow for ADCC
NKG2D: homodimeric CLR, recognises stress signals
NKp46: - recognises viral hemagglutinins
- and ecto-calreticulin, an ER stress protein
What are some examples of inhibitory receptors on NK cells?
NKG2A/CD94: contains an ITIM in its cytoplasmic tail. recognises HLA-E, a non classical MHC I
inhibitory KIRs: recognise HLA-A, B, C (classical MHC class I molecules). contain ITIMs in cytoplasmic tails
Siglec-7: recognises Sialic acid-containing glycans found on the surface of healthy cells. also plays a significant role in inhibiting IgE-mediated mast cell activation
What are the functions of the different NK cell subsets, and what markers are present on each?
CD56 bright: rare in blood but abundant in certain tissue. can rapidly produce immunomodulatory cytokines
(CD56, CCR7, CSF2)
CD56 dim: dominant in peripheral blood. cytotoxic effector role by releasing cytolytic granules
(CD16, CD56, CX3CR1)
tissue-resident NK (trNK): heterogenous population found across various tissues. cytokine and KIR profile differ from conventional NK cells
(CCR7, CD69, EOMES, ICAM1)
adaptive NK: long lived. can rapidly proliferate and produce cytokines upon viral reexposure (memory). also plays role in malaria defence. newest subset
(B3GAT1 (CD57), KLRC2)
How do NK cells perform granule-mediated cytotoxicity?
through formation of an immune synapse w target cell via integrin binding (LFA-1 on NK cells, and ICAM-1 on target cell)
lytic granules transported along microtubules and fuse w PM to be released into synaptic cleft, a tight space where granules can be released into target cell
– this prevents release into environment which would be detrimental
How do granzymes kill target cells?
caspase activation
– granzyme B cleavage and activation of caspase 3 = DNA fragmentation
mitochondrial dysfunction
– granzyme B cleavage of Bid into tBid (truncated = active), induces Bax and Bak activation and oligomerisation = formation of holes in mitochondria OM
– granzyme A degrades Complex I subunit in electron transport chain (ETC) causing ROS generation
caspase independent apoptosis
– granzyme A cleaves SET complex, proteins that bind and protect DNA = ssDNA breaks via nuclease
How does perforin form holes in the PM?
in the presence of a lipid bilayer, C2 domain binds Ca2+ to dock into membrane and then oligomerise to create channel = 14-20 nm in diameter
alternate pathway proposes that perforin can be endocytosed and form pores from within vesicle to release granzyme
granzymes = 5nm
How are NK cells unaffected by the perforn/granzyme system?
SB9: granzyme B inhibitor
– in cytoplasm, granzyme B substrate = conformational changes and deactivation
Lamp-1 (CD107a): degranulation marker
– inhibits binding of perforin to NK PM
Cathespin B: perforin cleavage
– cleaves perforin, prevent insertion PM. expressed on PM during degranulation
– knockouts do not show increased susceptibility to cytotoxicity = redundancy, ood thing
How do NK cells perform receptor mediated cytotoxicity?
Fas/FasL - target cell/NK cell
– results in caspase 8/10 activation leading to caspase cascade –> degradation of cellular proteins, DNA, and structural components, leading to apoptotic cell death
TRAIL/TRAIL R - v similar to Fas/FasL
– these seem more specialised to targeting cancer cells (express R-1, R-2)
– TRAIL-R decoys (eg R-3, R-4) exist on healthy cells, preventing apoptosis
TNF
– can bind TNFR1 and 2 to induce apoptosis
What are the TLR adaptor proteins?
- MyD88
- TRIF
- TIRAP/MAL
- TRAM
- TRAF3
TLR signalling largely divided into MyD88-dependent and TRIF-dependent pathways
MyD88: utilised by all TLRs
= activates NK-kB and MAPKs for induction of inflammatory cytokine genes
TRIF is recruited to TLR3 and 4
= promotion of alternative pathway that leads to activation of IRF3, NK-kB, and MAPKs for induction of type I IFK and inflammatory cytokine genes
Give some detail about TLR 1/2 and 2/6.
1/2 detect triacyl lipopeptides
2/6 detects diacyl lipopeptides
= highly expressed cell wall components in gram+ bacteria
can also detect alarmins -> endogenous ligands eg heat shock proteins
signal through MyD88 (TIRAP/Mal facilitates connection to receptor)
downstream signalling triggers SEAP promoter = secretion of alkaline phosphatase (ALP) or induce production of pro-inf cytokines eg TNF a, IL, IFN
– ALP dephosphorylates peptidoglycan, previnting binding to receptors
Give some detail about TLR4.
detects LPS
- uses MD-2, an accessory protein, to assist in detection and dimerisation
extracellular LBP binds LPS monomer -> delivery to soluble or membrane CD14 -> transfer LPS to TLR4/MD-2 complex = homodimerisation -> dimerisation of TIR -> binding of MyD88
this activates TF NF-kB and MAPK -> transcription of pro-inf cytokines
endocytosis of LPS-TLR4/MD-2 complex leads to TRIF/TRAM-dependent pathway -> induction of IRF3 and IFNs
Give some detail about TLR7 and 8?
detect GU-rich short ssRNA in endosomes/lysosomes
- signal through MyD88
- nuclear translocation of AP-1, NF-kB, and IRFs
- phosphorylation of IRFs promote induction of interferon stimulated response element (ISRE), w expression of IFN
== production of pro-inf cytokines
What is the cGAS-STING pathway?
cyclic GMP-AMP synthase (cGAS) is a cytosolic receptor that detect cytosolic dsDNA, from bacteria, viruses, or damaged cell
- detection of dsRNA by cGAS to produce cGAMP from ATP and GTP
- cGAMP binds to STING dimer present on ER membrane and activates its signalling
- STING activates the kinase TBK1 to phosphorylate IRF3 which enters nucleus and induces expression of type 1 interferon genes
What is AIM2?
AIM2 recognizes dsDNA in the cytoplasm from viral or bacterial infection or mitochondrial DNA released from damaged cells
AIM2 binds to dsDNA via its HIN200 domain
The PYD of AIM2 interacts with the PYD of ASC (adaptor), leading to oligomerization of ASC
ASC recruits and activates pro-caspase-1 by forming a large multiprotein complex.
Active caspase-1 cleaves pro-IL-1β and pro-IL-18 into their mature, secreted forms
Caspase-1 also induces pyroptosis, a form of inflammatory cell death
