Transcript The Major Histocompatibility Complex

TCR Identification
February 13, 2007
The elusive TCR
“The T-cell receptor-the molecule in the T-cell membrane
capable of specifically binding antigen-is the San Andreas
Fault of immunology: here the plates rub against each other,
shifting gradually and unnoticed, and a major “earthquake”
may occur at any moment.”
Klein 1982
“Few issues in science have proved as difficult to resolve and
as subject to contention as the nature of the TCR.”
Jean Marx (writer for Science) 1983
“The study of the T cell receptor has been the subject of many
studies, many of which have led to diverse and controversial
findings. At the present time, the consensus is that the
receptor is not conventional surface Ig as found on B cells.”
Unanue and Benacerraf 1984
Initial lack of success in
identifying the TCR
• IgT-antibodies against an Ig-like
molecule on the surface of T cells
• Identify/clone by homology to
immunoglobulin genes
IgT
Identifying the TCR
• Two overall approaches
– cDNA cloning
– anti-TCR antibodies, protein
• Leading laboratories
– cDNA:Hedrick/Davis (mouse), Tak Mak
(human)
– Protein: Kappler/Marrack,Allison,
Trowbridge, Reinherz
Isolation of cDNA clones encoding
T cell-specific membraneassociated proteins
Steve Hedrick, David Cohen, Ellen
Nielsen, and Mark Davis
Laboratory of Immunology
NIAID, NIH
Cytochrome c
• Model protein with isolates from many different
species to study the fine specificity of T cell
recognition
• Ron Schwartz’s laboratory at NIH was a leading
investigator studying cytochrome c
• Canonical response was B10.A mice against pigeon
cytochrome c (PCC)
• A dominant epitope 91-104 of PCC was identified that
was I-Ek restricted
• A heteroclitic response of the same T cells was
observed with moth cytochrome c 91-103
• The heteroclitic response was due to the MCC
peptide being one AA shorter than PCC, which
resulted in an optimal I-Ek anchor residue at P9
• The response to MCC is oligoclonal and utilizes V3
and V11 exclusively
Sequence of TM86 clone reveals an immunoglobulin like molecule
LOCUS
1109237A
123 aa
DEFINITION T cell receptor VDJbeta 1.9.2.
ACCESSION 1109237A
VERSION 1109237A GI:224610
DBSOURCE prf: locus 1109237A;
linear VRL 26-JUN-1996
state: germ;
taxonomy: Virus.
KEYWORDS T Cell Receptor V beta Gene; Mouse; Seq Determination; 450bp;
96AAs; Use Limited Repertoire of Gene; Somatic Diversification at
V-D-J Region; DNA Blot; Single Gene Subfamily; Variability Plot;
Hydrophobicity; beta Sheet Potential.
SOURCE
Murid herpesvirus 1 (Murine cytomegalovirus)
ORGANISM Murid herpesvirus 1
Viruses; dsDNA viruses, no RNA stage; Herpesviridae;
Betaherpesvirinae; Muromegalovirus.
REFERENCE 1 (residues 1 to 123)
AUTHORS Barth,R.K., Kim,B.S., Lan,N.C., Hunkapiller,T., Sobieck,N.,
Winoto,A., Gershenfeld,H., Okada,C., Hansburg,D., Weissman,I.L. and
Hood,L.
TITLE The murine T-cell receptor uses a limited repertoire of expressed V
beta gene segments
JOURNAL Nature 316 (6028), 517-523 (1985)
PUBMED 2412120
COMMENT cDNA.
FEATURES
Location/Qualifiers
source
1..123
/organism="Murid herpesvirus 1"
/db_xref="taxon:10366"
Region
30..123
/region_name="IGv"
/note="Immunoglobulin domain variable region (v)
subfamily; members of the IGv subfamily are components of
immunoglobulins and T-cell receptors; cd00099"
/db_xref="CDD:28982"
ORIGIN
1 vslclvetal mntkitqspr ylilgranks leceqhlghn amywykqsae kppelmflyn
61 lkqlirnetv psrfipecpd ssklllhisa vdpedsavyf cassqtggnt evffgkgtrl
121 tvv
Blast search of TM86
identifies it to be V1,
but cytochrome c
responses are all V3!
Cloning of the human TCR
Complete primary structure of a heterodimeric T-cell receptor deduced from cDNA
sequences.
* Saito H,
* Kranz DM,
* Takagaki Y,
* Hayday AC,
* Eisen HN,
* Tonegawa S.
Nature. 1984 Jun 28-Jul 4;309(5971):757-62.
Two related, but distinct, cDNA clones have been isolated and sequenced from a
functional murine cytotoxic T-lymphocyte clone. The genes corresponding to these cDNA
are expressed and rearranged specifically in T cells and both have similarities to
immunoglobulin variable and constant region genes. It is concluded that these genes
code for the two subunits of the heterodimeric antigen receptor on the surface of the T
cell; its complete deduced primary structure is presented.
PMID: 6330561 [PubMed - indexed for MEDLINE]
Transfection of these two receptors does not recapitulate
the specificity of the 2C TCR!
The Major Histocompatibility
Complex-restricted on T cells in
Mouse and Man: Identification of
Constant and Variable Peptides
John Kappler, Ralph Kubo, Kathryn
Haskins, Charles Hannum, Michele
Pigeon, and Pippa Marrack
National Jewish Hospital, Denver
Brad McIntyre and Jim Allison
UT, Smithville-> Berkeley->MSK
Ian Trowbridge
Salk Institute, La Jolla
Major players in the protein
identification of the TCR
• Kappler and Marrack
– Generated monoclonal antibodies against the
TCR of ovalbumin specific T cell hybridomas (i.e.
DO.11.10)
• Jim Allison
– Generated a monoclonal antibody against a T cell
tumor line
• Ian Trowbridge
– Found an anti-TCR in a mouse anti-human T cell
monoclonal antibody-retrospectively
• Ellis Reinherz
– Monoclonal against an alloreactive human clone
A T cell name REX
“REX line was obtained from
a patient with T cell
lymphoblastic lymphoma
which gave rise to
spontaneously growing
subclones as isolated by
limiting dilution technique.”