class i pathway prediction of proteasomal cleavage and tap binidng
DESCRIPTION
Class I pathway Prediction of proteasomal cleavage and TAP binidng. Can Keşmir, TBB, Utrecht University, NL & CBS, BioCentrum, DTU. Outline. MHC class I epitopes Antigen processing Proteasome Specificity and Polymorphism Prediction methods TAP Binding motif Evolution Immune escape. - PowerPoint PPT PresentationTRANSCRIPT
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Class I pathwayPrediction of proteasomal cleavage and
TAP binidng
Can Keşmir,TBB, Utrecht University, NL &
CBS, BioCentrum, DTU
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Outline
• MHC class I epitopes– Antigen processing
• Proteasome– Specificity and Polymorphism– Prediction methods
• TAP– Binding motif
• Evolution• Immune escape
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Peptide generation in the class I pathway
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Proteasomal cleavage
• ~20% of all peptide bonds are cleaved• Average peptide length 8-9 amino acids• Not all peptide bonds are equally likely cleaved
• Cleavage more likely after hydrophobic than after hydrophilic amino acids
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Proteasome specificity
• Low polymorphism– Constitutive & Immuno-
proteasome
• Evolutionary conserved• Stochastic and low specificity
– Only 70-80% of the cleavage sites are reproduced in repeated experiments
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Proteasome evolution (1 unit)
Constitutive
Immuno
Human (Hs) - HumanDrosophila (Dm) - Fly
Bos Taurus (Bota) - CowOncorhynchus mykiss (Om) - Fish
…
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Immuno- and Constitutive proteasome specificity
...LVGPTPVNIIGRNMLTQL..
P1 P1’
Immuno Constitutive
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• NetChop– Neural network based method
• PaProc– Partially non-linear method (a neural
network without hidden neurons????)• SMM (stabilized matrix method)• FragPredict
– Based on a statistical analysis of cleavage-determining amino acid motifs present around the scissile bond (i.e. also weight matrix like)
Predicting proteasomal cleavage
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NetChop20S-3.0In vitro digest data from the constitutive proteasome
Toes et al., J.exp.med. 2001
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NetChop 3.0 Cterm (MHC ligands)
LDFVRFMGVMSSCNNPA LVQEKYLEYRQVPDSDP RTQDENPVVHFFKNIVT TPLIPLTIFVGENTGVP LVPVEPDKVEEATEGEN YMLDLQPETTDLYCYEQ PVESMETTMRSPVFTDN ISEYRHYCYSLYGTTLE AAVDAGMAMAGQSPVLR QPKKVKRRLFETRELTD LGEFYNQMMVKAGLNDD GYGGRASDYKSAHKGLK KTKDIVNGLRSVQTFAD LVGFLLLKYRAREPVTK SVDPKNYPKKKMEKRFV SSSSTPLLYPSLALPAP FLYGALLLAEGFYTTGA
• NetChop-3.0 C-term– Trained on class I
epitopes– Most epitopes are
generated by the immunoproteasome
– Predicts the processing specificity
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Prediction performance
SensTPAP
Spec TN
AN
CC TP TN FN FPPPAN APPN
TPFP
APAN
Aroc=0.5
Aroc=0.8
1 - spec
Sen
s
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Predicting proteasomal cleavage
-0.4-0.2
00.20.40.60.8
1
Per
form
ance
Sens Spec CC
0
0.5
1
Perf
orm
an
ce
CC PCC Aroc
CC 0.12 0.1 0.41 0.48
PCC 0.13 0.48 0.55
Aroc 0.56 0.82 0.85
FragPredict PAProCI Netchop20S NetChop20S-3.0
NetChop-3.0
NetChop20S--3.0
• Relative poor predictive performance–For MHC prediction CC~0.92 and AUC~0.95
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Proteasome specificity
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What does TAP do?
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TAP affinity prediction
• Transporter Associated with antigen Processing• Binds peptides 9-18 long• Binding determined mostly by N1-3 and C terminal amino acids
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TAP binding motif matrix
Peters et el., 2003. JI, 171: 1741.
A low matrix entry corresponds to an amino acid well suited for TAP binding
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Predicting TAP affinity
9 meric peptides >9 meric
Peters et el., 2003. JI, 171: 1741.
ILRGTSFVYV-0.11 + 0.09 - 0.42 - 0.3 = -0.74
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Proteasome, TAP and MHC co-evolution
• Antigen processing and presentation is highly ineffective• Only 1 in 200 peptides will bind a given MHC complex• If proteasome and TAP do not effectively produce MHC restricted peptides, antigen processing would be a severe bottleneck for antigen recognition
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Co-evolution of Proteasome, TAP and MHC
• CP-P1: Constitutive proteasome specificity at P1 position• TAP-9: TAP motif at P9 position• MHC-9: Average MHC motif at P9
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Co-evolution of Proteasome, TAP and MHC
• IP-P1: Immuno proteasome specificity at P1 position• CP-P1: Constitutive proteasome specificity at P1 position• TAP-9: TAP motif at P9 position• MHC-9: Average MHC motif at P9
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Co-evolution (continued)
Kesmir et al. Immunogenetics, 2003, 55:437
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What is going on at the N terminal?
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S T R K F L D G N E M T L . . .
Epitope identification
TAP precursor A2 Epitope FLDGNEMTL
FLDGNEMTL 2.0100 KFLDGNEMTL -2.5300
RKFLDGNEMTL -3.7400 TRKFLDGNEMTL -2.4400
0.0101 0.6483 0.9955 0.9984 0.4299 0.2261 0.0103 0.0265 0.0099 0.0099 0.9590 0.4670 0.9989
Proteasomal cleavage
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N terminal trimming
>50% need 2-3 amino acids N terminal trimming
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TAP and proteasome independent presentation
• CTL epitopes are presented at the cell surface on TAP deficient cell lines•Some CTL epitopes have very poor TAP binding affinity• Dominant CTL epitopes can have very poor C terminal cleavage signal • Many CTL epitope have strong internal cleavage sites
• Other important players in the class I pathway
– Signal peptides– Sec61– Diffusion– Proteases
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Immune escape
• Pathogens evolve under strong selection pressure to avoid CTL recognition
• Generate point mutations or insertions/deletions to disturb– Peptide binding to MHC– CTL recognition
• Only involve the antigentic peptide region
– Antigen processing• Can involve peptide flanking region
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Immune escape via antigen processingHIV-1 Nef epitope VPLRPMTY (Milicic et al. JI, 2005, 4618)
IP
IP
CP
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Summary
• The most important players (MHC, TAP and proteasome) in the MHC class I pathway have co evolved to a share a common C terminal pathway specificity
• We can predict (up to a degree) proteasomal cleavage • TAP binding motif characterized in a weight matrix
– Binding mostly determined by the N1-3 and C terminal amino acids
• Proteasome produces and TAP transports precursor T cell epitopes of length 8-13 amino acids
• Epitope trimming in the ER by several amino peptidases (ERAP)
• We still do not understand everything– Many more important players are involved in the class I path way