The T Cell Receptor:Structure and Genetic Basis

Jeffrey K. Actor, Ph.D.MSB 2.214, 500-5344

Lecture Objectives:

• Present an overview of T cell receptor (TCR) structure and organization of the genes encoding the TCR chains.

• Understand underlying mechanisms involved in generation of TCR diversity.

• Compare/contrast the B cell receptor with that of the T cell receptor.

The T LymphocyteDealing with intracellular pathogens

• While antibodies are critical for response to antigens present outside of cells, there is need for effective response to antigens inside cells.

• T cells evolved to handle intracellular pathogens: viruses, bacteria and parasites.

• Whereas B cells recognize physical conformations and large molecular shapes, T cells recognize small antigenic determinants expressed on the surface of host cells associated with major histocompatibility complexmolecules.

The T LymphocyteThe “Ringleaders” of the Immune Response

• Regulates immune responses.• Integral in cell mediated immunity.• Critical in B cell-antibody production.

Every mature T cell expresses a TCR with specificity for an antigenic determinant.

• Each T cell has a TCR: a transmembrane heterodimer composed of two disulfide-linked polypeptide chains.

• alpha [] and beta [] chains, or• gamma [] and delta []

• The TCR is expressed on the cell surface in association with co-receptor (accessory) molecules.

• The TCR is not secreted, and remains membrane bound throughout the activation process.

The T Cell Receptor (TCR)

• Each chain (, , , or ) represents a distinct protein with molecular weight between 40 and 60 kDa.

• Most T lymphocytes express alpha [] and beta [] chains on their surface.

• Cells that express gamma [] and delta [] chains comprise only 5% of normal circulating T cells in healthy adults.

The T Cell Receptor

• An individual T cell can express either an or a heterodimer as its receptor, but never both.

• Two polypeptides which span the cell membrane.

The T Cell Receptor

• Each peptide comprised of a constant and a variable region.

• Intra- and inter- disulfide links.

• Antigen binding site is farthest from cell membrane.

• Ag binding site is comprised of physical structure contributed by both peptides.

The T Cell Receptor

The structure of the TCR complex:

- the antigen-binding chains, and - the associated signal CD3 transduction complex

CD3 comprised of , , and chains; (zeta) or (eta) or (theta); (-) and (+) are electrostatic interactions.

The T Cell Receptor Complex

The interaction of TCR, MHC, and linear peptide. The complementarity determining regions (CDRs) of the TCR Variable regions and peptide bound in the peptide-binding groove of an MHC class I molecule are depicted.

TCR Antigen Binding

Genes Coding for TCR

• T cell receptor genes are closely related members of the immunoglobulin gene superfamily.

• Each chain consists of a constant (C) and a variable (V) region, formed by a gene-sorting mechanism similar to antibody formation.

• The repertoire is generated by combinatorial joining of variable (V), joining (J), and diversity (D) genes, and by N region diversification (nucleotides inserted by the enzyme deoxynucleotidyl-transferase).

• V, D, and J genes are mixed together in a more complicated manner than for the Immunoglobulin genes.

• and use only V and J segments.

• and use V, D, and J segments.

• There are more V and V genes (50-100) than Vand V genes (5-10) in the germ line.

Genes Coding for TCR

Germline Genes Coding for TCR

Germline Genes Coding for TCR

Location of and TCR Genes

• The and chain genes are mixed together in one locus. The genes encoding the chain are entirely located between the cluster of V and J gene segments.

• The organization of the chain locus is extremely complex.

• The earliest cell entering the thymus has its TCR genes in the germ line configuration (unrearranged*).

Order of TCR Gene Rearrangement

*( Some rearrangement can occur in bone marrow)

• Both and chain genes then begin to rearrange, more or less simultaneously.

• If the chain genes rearrange successfully, then chain genes also start to rearrange. If both and genes rearrange functionally, no further gene rearrangement takes place and the cell remains a T cell.

• If and/or rearrangements are not functional, then gene rearrangement continues followed by gene rearrangement. In this manner, a product appears, and the cell becomes an T cell.

Allelic Exclusion

• If gene rearrangements are successful, no further rearrangements occur. This is a process called allelic exclusion.

• If all the gene rearrangements are unsuccessful, the second copy of genes on respective chromosomes will begin the rearrangement process.

• If this recombination is not successful, then the cell undergoes apoptosis.

• Recombination of V, D, and J gene segments is coordinated by recombinase-activating genes RAG-1and RAG-2.

• The enzymes recognize specific DNA signal sequences consisting of a heptamer, followed a spacer of 12 or 23 bases, and then a nonamer (Recombination Signal Sequences).

• If either RAG gene is impaired or missing the homologous recombination events are abolished, giving rise to severe combined immunodeficiency (SCID).

Process of Recombination

Rearrangement of the T-cell receptor genes.

Recombination of V, D, and J gene segments is coordinated by recombinase-activating genes RAG-1 and RAG-2.

If either RAG gene is impaired or missing the homologous recombination events are abolished, giving rise to severe combined immunodeficiency (SCID).

Generation of Diversity

Ig TCRNumber of V gene pairs 3.4 x 106 5.8 x 106

Junctional Diversity ~3 x 107 ~2 x 1011

Total Diversity ~1014 ~1018

•Greater diversity in TCR compared to B cell Ig receptor.

No Somatic Mutation in TCR

• Unlike immunoglobulin genes, genes encoding TCR do not undergo somatic mutation. There is no change in TCR affinity during activation, differentiation, and expansion.

More on T Lymphocyte Development

• T lymphocytes develop in the thymus. • In the thymus, the cells develop both a CD4

and a CD8 marker. They lose one marker prior to leaving the thymus. – The cells with a CD4 marker are called helper T

cells (Th cells). – The CD8 positive cells that develop are cytotoxic T

cells (Tc cells).

Genesis of Mature T Lymphocytes

95% apoptosis

5% exported to periphery

(double positive)

(single positive)

Thymic Selection

No interaction = CD4+CD8+ cell dies

Interaction = Positive SelectionMHC + self

or MHC + non-self

++, CD4+CD8+ cell interacts with Thymic

epithelial cell

++, CD4+CD8+ cellinteracts with interdigitating cell

High affinity interaction = Deletion

Low affinity interaction = Survival

Commitment CD4+ or CD8+

Double positive in thymus, leave as committed single positive cells

T Helper Cells

• Different phenotypic populations exist.– TH1, TH2, TH17, Treg ……and more…..

• All express the CD4 molecule.• Aid effector T lymphocytes in cell-mediated

immunity. • Aid antigen-stimulated subsets of B cells to

proliferate and differentiate toward antibody-producing cells.

• Regulatory role for tolerization events.

T Cytotoxic Cells

• T cytotoxic cells (CTLs) express CD8 and are cytotoxic against tumor cells and host cells infected with intracellular pathogens.

T Regulatory Cells

• T regs are a population of T Helper cells (CD4+CD25+), characterized by TGF-βsecretion, that also serve as regulators of response.

Natural Killer T Cells

• Natural killer T cells (NKT) share properties of both T cells and natural killer (NK) cells. – NKT cells are different from NK cells.

• These cells recognize lipid and glycolipid antigen.

• Have a limited TCR repertoire.

“Unconventional” Antigen Presentation to T Cells

• Lipids/glycolipids • Superantigens

Non-Classical Ag PresentationLipids and Glycolipids

CD1 is a Surface glycoproteinwhich can present lipids/glycolipids to T cells.

• Non-MHC encoded, Non-polymorphic

• Expressed in association with 2-microglobulin

• Binds hydrophobic region of lipid, exposing polar region for T cell interaction

• Can present to or T cells, and NKT cells.

“Unconventional” Antigen Presentation to T Cells (cont’d)

• Lipids/glycolipids• Superantigens

Superantigens• Superantigens bind directly to

T-cell receptors and MHC, without processing.

• “Presented” by MHC II, but not in peptide groove

• Involves direct interaction to V region of TCR; activates any T cell expressing specific V TCR segment

• Non-specific activation of large number of T cells

• Various organisms have superantigens in makeup– Staphylococcus, rabies

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

Fig 47.1 Garland Science

Case #47, Geha and Notarangelo:

“Toxic Shock Syndrome”

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

VDJ

VJ

CC

Fig 47.1 Garland Science

Superantigens

Comparison of B Cell and T Cell

Receptors

B Cell vs T Cell Receptors

BCRs and TCRs SHARE these properties: integral membrane proteins, present in thousands of identical

copies exposed at the cell surface made before the cell ever encounters an antigen encoded by genes assembled by the recombination of DNA allelic exclusion ensures only one receptor with a single

antigenic specificity demonstrate N region addition during gene rearrangement have a unique binding site to recognize antigenic determinant

(epitope) binding depends on complementarity of the receptor with the

epitope binding of antigen occurs by non-covalent forces

B Cell vs T Cell Receptors

BCRs and TCRs DIFFER in these properties: structure genes that encode them type of epitope to which they bind TCRs do not somatically mutate TCRs do not undergo isotype switching TCR gene recombination exhibits far greater junctional

diversity than Ig genes TCRs are never secreted from the T cell

Comparison: BCR vs TCR

Clinical Vignette:Dysfunction in Gene Rearrangement

• Omenn syndrome is characterized by generalized erythematous skin rash, lymph node enlargement, hepatosplenomegaly, shift in immunoglobulin isotypes, and evidence of combined immune deficiency.

• Geha & Notarangelo, 6th ed: Clinical Companion Case #7

Omenn Syndrome

The skin is bright red and wrinkled from edema and infiltration of inflammatory cells.

Diffuse, scaly rash on the face and shoulders of an infant with Omenn syndrome. Conjunctivitis present.

Case 7 Garland Science

Clinical Vignette: Omenn Syndrome

A: Recombination of V, D, and J gene segments is coordinated by recombinase-activating genes RAG-1 and RAG-2. Many Omenn Syndrome patients have etiology of missense mutations in RAG-1 or RAG-2 gene, with only partial recombinase activity.

•If either RAG gene is impaired or missing the homologous recombination events are abolished, giving rise to severe combined immunodeficiency (SCID, T-, B-).

Q: In Omenn Syndrome there is fault in assembly of gene segments that encode the variable regions of and chains of the TCR. What is the mechanism underlying the defect?

T lymphocytes are involved in regulation of immune response and in cell mediated immunity.

Mature T cells express antigen-specific TCR in a complex with CD3 molecules. The TCR is a disulfide-linked heterodimer composed of either or chains. T cells express either or chain heterodimers, but never both.

T cell receptor genes are closely related members of the immunoglobulin gene superfamily and derive part of their structural diversity form recombination of different V, D, and J gene segments.

During differentiation in the thymus, immature T cells undergo rearrangement of their TCR, and commit to lineage of CD4+ or CD8+ phenotype.

TCR Summary

A child with Severe Combined Immunodeficiency (SCID) has a deficiency in her lymphocyte populations, which include B and T lymphocytes. Regarding normal B and and T lymphocytes, which property is not shared by both the B and T cell receptors?

A. they are made prior to encounters with antigen B. they undergo somatic mutation after antigenic stimulation C. they are encoded by recombined segments of DNA D. they show allelic exclusion for single antigen receptor expression E. they demonstrate N region addition during gene rearrangement

Option B (they undergo somatic mutation after antigenic stimulation) is correct. The B and T cell receptors are integral membrane proteins present in thousands of identical copies exposed on the cell surface, that are available to specifically react with antigen prior to encountering antigen. The receptors are encoded for by a genes comprised of recombined DNA segments. The tremendous binding potential for binding antigens (>1015 different receptors) is in part due gene rearrangement mechanisms that bring together Variable (V), Diversity (D), and Junction (J) gene sequences. However, only the B cell receptor (the immunoglobulin molecule) undergoes somatic mutation after antigenic exposure, thus driving increases in functional affinity of the antibody-antigen reaction.

Element Name: WOMAN; Symbol: WOAtomic Weight: (don't even go there!)

Physical properties: Generally round in form. Boils at nothing and may freeze any time. Melts whenever treated properly. Very bitter if not used well.

Chemical properties: Very active. Highly unstable. Possesses strong affinity to gold, silver, platinum, and precious stones. Violent when left alone. Able to absorb great amounts of exotic food. Turns slightly green when placed next to a better specimen.

Usage: Highly ornamental. An extremely good catalyst for dispersion of wealth. Probably the most powerful income reducing agent known.

Caution: Highly explosive in inexperienced hands.

Element Name: MAN; Symbol: XYAtomic Weight: (180 +/- 50)

Physical properties: Solid at room temperature; gets bent out of shape easily. Fairly dense and sometimes flaky. Difficult to find a pure sample. Aging samples unable to conduct electricity as easily as young samples.

Chemical properties: Attempts to bond with WO any chance it can get; tends to form strong bonds with itself. Becomes explosive when mixed with Kd (Element: Child) for prolonged period of time. Neutralize by saturating with alcohol.

Usage: None known. Possibly good methane source.

Caution: In the absence of WO, this element rapidly decomposes and begins to smell.

Comparison: BCR vs TCR