Structural Basis for Self-Peptide/MHC Recognition by
Autoimmune T Cell Receptors in Multiple Sclerosis
(recognition of self)
Structural Basis for Self-Peptide/MHC Recognition by
Autoimmune T Cell Receptors in Multiple Sclerosis
(recognition of self)
Structural Basis for Tumor Antigen Recognition by
CD4+ T Cells
(recognition of mutated self)
Myelin proteins as targets for CD4+ T cells
PLP and MBP
MAG
CNPase
MOG
• Proteolipid protein (PLP)
Myelin basic protein (MBP)
• Myelin oligodendroglia glycoprotein
(MOG)
• 2’3’ Cyclic nucleotide 3’
phosphodiesterase (CNPase)
• Myelin-associated glycoprotein
(MAG)
•
All the above self-antigens are able to
induce EAE, an animal model of MS.
Myelin proteins as targets for CD4+ T cells
PLP and MBP
MAG
CNPase
MOG
• Proteolipid protein (PLP)
Myelin basic protein (MBP)
• Myelin oligodendroglia glycoprotein
(MOG)
• 2’3’ Cyclic nucleotide 3’
phosphodiesterase (CNPase)
• Myelin-associated glycoprotein
(MAG)
•
All the above self-antigens are able to
induce EAE, an animal model of MS.
Immunology of Multiple Sclerosis
•Susceptibility to MS is associated with certain MHC class II
haplotypes, most notably HLA-DR2.
•HLA-DR2 haplotype comprises two alleles (HLA-DR2a and HLADR2b) that are co-expressed on the cell surface and are thought to act
in concert in conferring susceptibility to MS.
•In humans, an immunodominant T cell epitope has been identified in
MBP, corresponding to residues 84–102.
•MBP 84–102 can be presented by both DR2a and DR2b to T cells
from MS patients, but in completely different ways.
Properties of T Cell Receptor 3A6
•TCR 3A6 was isolated from an HLA-DR2+ MS patient.
•TCR 3A6 recognizes MBP 84–102 in the context of HLA-DR2a.
•Mice transgenic for TCR 3A6 and HLA-DR2a develop spontaneous
CNS inflammation, demonstrating the pathogenic potential of this TCR.
•TCR 3A6 displays considerable degeneracy in peptide recognition, as
determined using combinatorial peptide libraries.
Questions
•
Do autoimmune TCRs engage peptide/MHC in the same manner
as anti-microbial TCRs, or do fundamental differences in binding
mode exist that may allow autoreactive T cells to escape negative
selection?
•
What is the structural basis for TCR degeneracy? TCR degeneracy
underlies the “molecular mimicry” hypothesis for autoimmune
disease.
Binding of TCR 3A6 tetramers to MBP/DR2a and superagonist/DR2a complexes
MBP/DR2a immobilized
self-peptide
HSP/DR2a immobilized
irrelevant peptide
K38/DR2a immobilized
superagonist peptide
•TCR 3A6 binds peptide/MHC with low affinity (Kd>100 mM) but high specificity.
•Nevertheless, 3A6 T cells are activated by nanomolar concentrations of MBP peptide.
•Superagonist peptide stimulates 3A6 T cells ~10,000-fold more efficiently than MBP peptide.
Construction of a stabilized TCR–MBP–MHC class II complex
Peptide (MBP)-TCR
Refold a chain and
pep-b chain together
MBP-TCR/DR2a complex
Mix MBP-TCR and CLIP/DR2a together in the
presence of HLA-DM (peptide exchanger)
at pH 5.8 / 37°C
-s-s-
Purify by
Anion Exchange
& Gel Filtration
Purify by
Anion Exchange
& Gel Filtration
mAU
mAU
1500
1000
800
1000
HLA-DM
600
400
500
200
0
0
0.0
0.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
5.0
10.0
15.0
20.0
ml
mAU
1000
800
600
-s-s400
MHC class II (HLA-DR2a)
Refold a chain, b chain,
and CLIP (class II
associated invariant chain
peptide) together
200
0
0.0
5.0
10.0
15.0
ml
mAU
400
300
Purify by
Anion Exchange
& Gel Filtration
200
100
0
0.0
mAU
1000
mAU
1200
800
1000
600
800
600
400
400
200
200
0
0.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
ml
0
0.0
5.0
10.0
15.0
20.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
ml
TCR 3A6/MBP/HLA-DR2a
(autoimmune)
TCR HA1.7/HA/HLA-DR1
(anti-microbial)
Vb
Vb
a1
N
a1
C
C
N
b1
Va
TCR 3A6/MBP/HLA-DR2a
(autoimmune)
b1
Va
TCR HA1.7/HA/HLA-DR1
(anti-microbial)
Vb
Vb
a1
N
a1
C
C
N
b1
Va
TCR 3A6/MBP/HLA-DR2a
(autoimmune)
b1
Va
TCR HA1.7/HA/HLA-DR1
(anti-microbial)
3A6/MBP/DR2a
172.1/MBP/I-Au
HA1.7/HA/DR1
Ob.1A/MBP/DR2b
Comparison of MBP/HLA-DR2a and MBP/HLA-DR2b complexes
MBP 87-104
PVVHFFKNIVTPRTPPPS
PVVHFFKNIVTPRTPPPS
P1
P4 P6
DR2a (DRB5*0101)
DR2b (DRB1*1501)
P9
Peptide register is shifted by three residues.
TCR-contacting residues in the MBP/DR2a and MBP/DR2b complexes
Interactions of TCR 3A6 with HLA-DR2a
a1
b1
Interactions of autoimmune vs. anti-microbial TCR with MHC
TCR 3A6/MBP/HLA-DR2a
TCR HA1.7/HA/HLA-DR1
Interactions of TCR 3A6 with MBP peptide
•No hydrogen bonds or salt bridges between 3A6 and MBP
•Interactions are restricted to van der Waals contacts
•Suboptimal shape and chemical complementarity
•Structural degeneracy is consistent with functional degeneracy
Proliferative testing of mouse transgenic 3A6 T cells with
human DR2a cells using combinatorial peptide libraries
P-1
8.0
6.0
4.0
P5
16.0
14.0
12.0
10.0
8.0
6.0
2.0
4.0
2.0
0.0
0.0
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
Y
P1
12.0
10.0
8.0
6.0
4.0
2.0
0.0
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
Y
P6
12.0
10.0
8.0
6.0
4.0
2.0
0.0
A
Y
P7
W
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
Y
16.0
25.0
P2
20.0
P7
14.0
12.0
10.0
15.0
8.0
10.0
6.0
4.0
5.0
2.0
0.0
0.0
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
Y
P3
15.0
10.0
5.0
0.0
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
20.0
W
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
8.0
4.0
2.0
Y
W
Y
P9
6.0
5.0
W
P8
A
10.0
0.0
C
7.0
6.0
5.0
4.0
3.0
2.0
1.0
0.0
Y
P4
15.0
A
0.0
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
Y
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
Denotes a.a. of native MBP ligand
Y
Proliferative responses to predicted superagonists of
TCR 3A6
14000
PROLIFERATION (cpm)
12000
FFKNIVTPRT: MBP 89-98
WFKLIPTTKL: AGONIST 1
WFKLILTPKL: AGONIST 2
10000
8000
6000
4000
2000
0
10-10 10-9
10-8
10-7
10-6
10-5
10-4
10-3
10-2
10-1
100
101
102
103
mg/ml
•Agonist peptides were deduced from combinatorial decapeptide
libraries
•Agonists are at least 10,000-fold more potent than the MBP selfpeptide
Interactions of TCR 3A6 with MBP peptide
Prediction: TCRs that recognize the N-terminal portion
of peptides are more degenerate (cross-reactive) than
TCRs targeting the central portion. Why?
Superposition of peptides bound to human MHC class II molecules
Is the unconventional binding
mode utilized by TCR 3A6
broadly characteristic of
autoreactive TCRs?
Properties of TCR E8
•TCR E8 was isolated from tumor-infiltrating CD4+ T cells of a melanoma patient.
•TCR E8 recognizes a mutated melanoma antigen presented by HLA-DR1.
•The tumor-specific antigen is a peptide from triose phosphate isomerase (TPI 23–37)
with a single amino acid substitution (Thr28Ile).
•TCR E8 therefore recognizes a neo-antigen derived from a self-antigen.
•T cell stimulation is enhanced >100,000-fold for mutated TPI 23–37 relative to the
wild type peptide.
Stimulation of tumor-infiltrating E8 T cells by wild type and mutant TPI 23–37
From Pieper et al. (1999) J. Exp. Med. 189, 757-765
Conformations of wild type and mutant TPI23-37 bound to
HLA-DR1
a1
b1
P-3 P-2
P-1
P1
P2
Gly23 Glu24 Leu25 Ile26 Gly27
P3
P4
P5
P6
P7
P8
P9
P10 P11 P12
Thr28/ Leu29 Asn30 Ala31 Ala32 Lys33 Val34 Pro35Ala36 Asp37
Ile28
Construction of stabilized TCR E8–TPI–DR1 complex
Peptide (TPI)-TCR
Refold a chain and
pep-b chain together
TPI-TCR/DR1 complex
Mix TPI-TCR and CLIP/DR1 together in the
presence of HLA-DM (peptide exchanger)
at pH 5.8 / 37°C
-s-s-
Purify by
Anion Exchange
& Gel Filtration
Purify by
Anion Exchange
& Gel Filtration
mAU
mAU
1500
1000
800
1000
HLA-DM
600
400
500
200
0
0
0.0
0.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
5.0
10.0
15.0
20.0
ml
mAU
1000
800
600
-s-s400
MHC class II (HLA-DR1)
Refold a chain, b chain,
and CLIP (class II
associated invariant chain
peptide) together
200
0
0.0
5.0
10.0
15.0
ml
mAU
400
300
Purify by
Anion Exchange
& Gel Filtration
200
100
0
0.0
mAU
1000
mAU
1200
800
1000
600
800
600
400
400
200
200
0
0.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
ml
0
0.0
5.0
10.0
15.0
20.0
5.0
10.0
15.0
20.0
25.0
30.0
35.0
ml
Overall structure of the E8/mutTPI/HLA-DR1 complex
Interactions between TCR E8 and mutant TPI peptide
TCR-induced conformational changes in peptide/MHC
Free TPI/DR1
Bound TPI/DR1
Docking topologies of TCRs
HA1.7/HA/DR1
anti-foreign
E8/TPI/DR1
anti-mutated self
3A6/MBP/DR2a
anti-self
Docking topologies of TCRs
HA1.7/HA/DR1
anti-foreign
E8/TPI/DR1
anti-mutated self
Affinity: HA1.7 > E8 > 3A6
3A6/MBP/DR2a
anti-self
CDR3 positions on peptide/MHC
HA1.7/HA/DR1
anti-microbial
E8/TPI/DR1
anti-tumor
3A6/MBP/DR2a
autoimmune
Position of TCRs along peptide antigen
HA1.7/HA/DR1
anti-foreign
E8/TPI/DR1
anti-mutated self
P5
3A6/MBP/DR2a
anti-self
P5
CARB
Yili Li
Lu Deng
Ries Langley
Patrick Brown
Yiyuan Yin
Gang Xu
Yuping Huang
Jessica Lue
Lesley Teng
NINDS/NIH
Roland Martin
Jacqueline Quandt
NCI/NIH
Suzanne Topalian
Conformational changes in TCR E8
Conformational changes in MBP/HLA-DR2a upon binding TCR 3A6
Free MBP/DR2a
Bound MBP/DR2a
Conformational changes in self-peptide/MHC upon TCR binding
MBP/DR2a after binding 3A6
Sc = 0.62
MBP/DR2a before binding 3A6
Sc = 0.50
•TCR-induced adjustments in MBP/DR2a increase interfacial shape complementarity.
•However, energetic cost of distorting MBP/DR2a from ground state reduces affinity.