Download CellPath Savage TCR Ig Re FINAL

Immunology:
Antigen recognition in the adaptive immune system
(TCR and Ig Rearrangement)
Peter A. Savage, Ph.D.
Assistant Professor
Department of Pathology
[email protected]
PATH 30100 01 Cell Pathology / Immunology (Spring 2016)
Monday, April 11, 2016
Murphy • Travers • Walport
Janeway’s Immunobiology
Seventh Edition
Copyright © Garland Science 2008
Features of antigen recognition by Igs and TCRs
the Ig serves as both
an antigen sensor and
an effector molecule
the TCR serves
solely as an
antigen sensor
A single peptide-MHC complex can trigger cytokine secretion by T cells
single peptide-MHC
labeled with
quantum dot
on APC surface
antigen-specific
T cell
“digital” production
of cytokine triggered
by single pMHC
“A single peptide-MHC ligand triggers digital cytokine secretion in CD4+ T cells”
Huang et al., Immunity 39:846, 2013
Structure of the T cell antigen receptor (TCR)
hypervariable
CDR3 loops
Recognition of peptide-MHC complex by TCR
hypervariable CDR3 loops
sit directly over peptide
Identification and characterization of the TCR protein
Immunized Balb/c mice with T-cell lymphoma C6XL derived from C57BL/Ka strain
T-cell lymphoma is presumed to express a unique TCR on it’s surface
Generated hybridomas producing antibodies reactive to C6XL cells
One antibody, clone 124-40, reacted with C6XL cells but did not react with normal spleen cells
Identified disulfide-linked cell-surface heterodimeric protein (the TCR ?!?!?)
“Tumor-specific antigen of murine T-lymphoma defined with monoclonal antibody”
Allison et al., Journal of Immunology 129:2293, 1982
Identification and characterization of the TCR protein
Two-dimensional SDS-PAGE analysis of cell extracts
125I-labeled cell surface antigens
C6XL extract immunoprecipitated
using 124-40 Ab
C6XL extracts
protein migrating
below the diagonal is
disulfide-linked
found to be a
heterodimer comprised
of 39 and 41 kDa chains
splenic
T cells
splenic
B cells
thymocytes
bone marrow
cells
“Tumor-specific antigen of murine T-lymphoma defined with monoclonal antibody”
Allison et al., Journal of Immunology 129:2293, 1982
ab TCR locus (germline configuration)
V(D)J recombination of the a and b TCR genes
TCR gene segments are flanked by
recombination signal sequences (RSS)
The 12/23 rule governs recombination fidelity
of RSS elements during V(D)J recombination
Induced asymmetry of the (RAG1-RAG2)2 complex
underlies the molecular mechanism of the 12/23 rule
“Molecular mechanism of V(D)J recombination from synaptic RAG1-RAG2 complex structures”
Ru et al., Cell 163:1138, 2015
V(D)J recombination of antigen receptors
V(D)J recombination of antigen receptors
coding joints
signal joints
Steps in maturation and selection of T lymphocytes
T cells attempt to re-arrange functional in-frame TCRa and TCRb chains to generate a stable TCRab heterodimer
TCRab-expressing thymocytes then undergo thymic education (positive selection, negative selection, lineage commitment)
rearranged TCRb first
pairs with invariant
pre-Ta chain
Structure of antibodies
Structure of Ig V domain
during B cell responses,
somatic hypermutation of the
CDR loops drives
affinity maturation
Features of the major isotypes (classes) of antibodies
J chain induces
multimerization
of IgA and IgM
Binding of an antigen by an antibody
antigen-binding site can recognize soluble macromolecules in their native conformation
Structures of full-length antibody molecules
Harris et al, 1992 &
1998
Saphire et al, 2001
(anti-HIV)
Immunoglobulin locus (germline configuration)
V(D)J recombination of the heavy and light chain Ig genes
Ig gene segments are flanked by
recombination signal sequences (RSS)
The 12/23 rule governs recombination fidelity
of RSS elements during V(D)J recombination
Steps in maturation and selection of B lymphocytes
Steps in maturation of lymphocytes
self-reactive
B cells undergo:
1. receptor editing
2. deletion
3. anergy
Features of the major isotypes (classes) of antibodies
J chain induces
multimerization
of IgA and IgM
Generation of hybridomas and monoclonal antibodies
Nobel Prize in Physiology and Medicine, 1984
development of monoclonal antibody technology
immune theory
production of monoclonal antibodies
Properties of antibodies and T cell antigen receptors (TCRs)
Potential diversity of Ig and TCR antigen receptors
Differences between Ig and TCR receptors
Case 7: Omenn Syndrome
A defect in V(D)J recombination results in severe immunodeficiency
The case of Ricardo Reis: a bright red rash betrays an immunodeficiency
By day 10 after birth, had 10 loose bowel movements per day
By day 21, developed rash over entire body
Normal weight and size
Scaly rash and blisters on face, trunk, extremities
Spleen and liver not enlarged
Ricardo’s parents had had two other children, a boy and a girl, who had died
after the onset of a similar rash at 1 month of age
Parents were first cousins of Portugese extraction
White blood cells had high % of eosinophils and monocytes, but few lymphocytes
Low levels of serum IgG, IgA, IgM but high levels of IgE
Dermis infiltrated by large numbers of eosinophils, lymphocytes, macrophages,
with large numbers of cells around blood vessels
Condition worsened in hospital, developed bacterial and fungal infections
Blood was devoid of B cells and contained only activated T cells
Sequencing revealed hypomorphic R229Q (Arg to Gln) mutation in RAG2 gene
While studies were carried out, Ricardo developed pneumonia and died of
respiratory failure
Case 7: Omenn Syndrome
A defect in V(D)J recombination results in severe immunodeficiency
Case 7: Omenn Syndrome
A defect in V(D)J recombination results in severe immunodeficiency