The T cell receptor (TCR) is a T cell surface receptor that recognizes antigens presented by MHC molecules. It is a heterodimer composed of either α and β chains or γ and δ chains, and which interacts with another T cell surface component CD3. αβ T cells are usually highly specific (adaptive immunity), while γδ T cells usually recognize broader antigen classes (innate immunity). TCR chains each contain two 60-75 amino acid domains: the amino-terminal variable (V) domain and the constant (C) domain. The V domain contains three hypervariable regions, similar to antibody complementarity determining regions.
The two chains are connected at a cysteine residue by a disulfide bond. The transmembrane region is 21 or 22 amino acids and positively charged, anchoring the TCR into the cell membrane and promoting TCR-CD3 interaction. Lastly, the cytoplasmic tail (carboxy-terminal end) is 5 to 12 amino acids. αβ and γδ TCR heterodimers resemble an Fab fragment attached to the cell membrane; due to this structural homology, they are included under the immunoglobulin superfamily.
αβ TCRs react only with peptide antigens. However, γδ TCRs can react to glycolipids and phospholipids (nonpeptides) presented by CD1. Furthermore, some γδ cells react with unprocessed peptide antigens not even presented by an MHC. Also, the most common γδ TCRs within a given species bind the most common pathogens that species encounters. These γδ TCRs directly binds microbes, and the γδ cells might directly kill pathogens.
Both innate and adaptive immunity rely heavily upon binding of the TCR to a peptide-MHC complex. γδ cells are characterized as part of innate immunity due to their rapid responses, low specificity and ability to bind free antigens. αβ cells, conversely, are clearly part of adaptive immunity. In addition to these functional differences, γδ cells and αβ cells are also structurally different. Allison, Garboczi and their coworkers found that a γδ receptor specific for non-presented microbial phospholipids has a deep binding cleft. Also, separate research found that γδ TCRs have a 111° bend between their V and C regions while αβ TCRs have a 147° bend.
In B cells, membrane-bound antibodies associate with the Ig-α/Ig-β heterodimer to form the B cell antigen receptor. In T cells, the T cell receptor associates with its fellow membrane-bound component CD3 to form the TCR-CD3 membrane complex. Please note that CD3 is another component of the cell surface on T cells — TCR and CD3 are together on the T cell surface and interact together to transduce signals into the T cell. CD3 is essential for membrane expression of the TCR and for signal transduction. All CD3 chains contain a negatively charged amino acid (aspartic or glutamic acid) which interacts with one or two positively charged transmembrane TCR residues. Please note that B and T cell accessory molecules only participate in signal transduction after antigen interaction and do not actually influence antigen interaction.
Alone, the T cell receptor has low affinity for the peptide-MHC complex. For this reason, additional accessory molecules known as coreceptors or cell-adhesion molecules strengthen the bond between a T cell and a target or antigen-presenting cell. CD4 and CD8 are coreceptors which also distinguish cytotoxic T cells from helper T cells; CD4 binds Class II MHC molecules and CD8 binds Class I MHC molecules. Once bound, CD4 and CD8 also activate signal transduction. Additional coreceptors include CD2, LFA-1, CD28 and CD45R, all of which bind independently to other ligands on target or antigen-presenting cell. Once these coreceptors bind, the TCR scans for its appropriate peptide-MHC complex. Once TCR binds, membrane expression of cell-adhesion molecules is activated to tighten the intracellular bond. Cell-adhesion molecules play a role in tissue graft acceptance or rejection. If the grafted tissue bears MHC molecules which are similar to the host, then T cells will still respond to the MHC molecules.
Genes encoding α, β, γ and δ chains are expressed only in T cells. Functional TCR genes are formed by rearrangements of V and J segments in α and γ genes, and of V, D and J segments (like IgH) in β and δ genes. The α, β, γ and δ genes also include C segments which do not rearrange. The C region contains a long exon encoding much of the C domain, followed by shorter exons encoding the connecting, transmembrane and cytoplasmic regions. In humans, α and δ gene segments are located on chromosome 14, while β and γ gene segments are located on chromosome 7. In mice, the α, β and γ gene segments are located on chromosomes 14, 6 and 13. In both mice and humans, δ gene segments are located between V and J α segments. The location of the δ gene segments is important: functional rearrangement of the α gene segments deletes Cδ, so that a T cell cannot co-express αβ TCRs with γδ TCRs. The table below describes human and murine TCR genes, and counts both pseudogenes (nonfunctional mutants) and functional genes.
| # of gene segments | ||||||||||
| Gene | Xsm | V | D | J | C | Overview | ||||
| Human α Chain | 14 | 54 | 61 | 1 | Jα region is enormous, with over 75 segments over 50kB. | |||||
| Mouse α Chain | 14 | 80 | 80 | 1 | ||||||
| Human β Chain | 7 | 67 | 2 | 14 | 2 | 30-50 V segments and two near-identical repeats of D, J and C segments. | ||||
| Mouse β Chain | 6 | 20 | 2 | 2 | 2 | Contains two almost identical repeats of D, J and C segments. | ||||
| Human γ Chain | 7 | 14 | 5 | 2 | Contains two almost identical repeats of J and C segments. | |||||
| Mouse γ Chain | 13 | 7 | 3 | 3 | Contains three different functional J-C repeats. | |||||
| Human δ Chain | 14 | 3 | 3 | 3 | 1 | δ gene segments are located between α gene V and J segments. | ||||
| Mouse δ Chain | 14 | 10 | 2 | 2 | 1 | |||||
Like the antibody L chain, the α chain is encoded by V, J and C segments. Like the antibody H chain, the β chain is encoded by V, D, J and C gene segments. Rearrangement of the TCR gene segments results in α VJ joining and β VDJ joining. After the rearranged TCR genes are transcribed, the α and β chains are expressed as a disulfide-linked heterodimer on the T cell membrane. Immunoglobulins can be bound or secreted, but TCRs are only membrane-bound. The TCR constant region, as shown in the figure above, includes constant (C), connecting, transmembrane and cytoplasmic sequences. Also, as opposed to the multiple different immunoglobulin C gene segments encoding different isotypes, α DNA has only one C segment and β DNA has duplicated J and C segments.
Pre-T cells expresses the recombination activating genes RAG-1 and RAG-2, as do pre-B cells. The RAG-1/2 recombinase recognizes conserved recombination signal sequences (RSSs) flanking each V, D and J gene segments in non-recombined TCR DNA. RAG-1/2 catalyzes V-J and V-D-J joining via the same deletions and inversions which occur in immunoglobulin genes. First, Rag-1/2 knicks one DNA strand between the coding and signal sequences and excises the resulting DNA loops. SCID mice (which lack B and T cells) are defective for a gene which repairs double-stranded DNA breaks; as a result, in SCID mice, the Ab and TCR D and J gene segments are not rejoined. RAG-1/2 only recombines TCR genes in T cells and Ig genes in B cells. This specificity is due to different recombinase regulatory systems, and uniquely configured chromatin in each lineage that allows recombinase to access only appropriate sites.
δ genes are located between the V and J segments of the &alpha gene. Thus, when the α gene recombines, the δ DNA is excised. In a phenomenon known as allelic exclusion, this mechanism prevents αβ TCRs and γδ TCRs from co-expressing. Another instance of allelic exclusion involves the two duplicate β gene J and C clusters. Having two duplicates means that at least one functional rearrangement is likely to take place. Once a functional rearrangement occurs, then rearrangement of the other cluster is inhibited. However, more than one α allele can undergo functional recombination; while this can lead to different α chains being expressed on the same cell surface, only one allele is MHC restricted and therefore functional. The mechanisms of TCR diversity are described below.
| Mechanism | Overview |
| Alternative Joining | In addition to combinatorial joining, δ genes can alternatively join. Although impossible with antibodies, there are functional (VDDDJ)δ and other processed δ genes. |
| Combinatorial Joining | This simply refers to the various ways that α V and J segments and β V, D and J segments can combine. |
| Junctional Flexibility | The junctions between gene segments are prone to nucleotide addition during rearrangement. |
| Nucleotide Addition | In addition to junctional flexibility, palindromic sequences known as p-region nucleotide additions can be added between gene segments. Also, nucleotides can be added at the ends of TCR genes via n-region nucleotide addition. |
| Step | Overview |
| MHC Restriction | Cytotoxic T lymphocytes (CD8+ T cells) only kill infected cells with a self MHC haplotype. T cells do not kill infected cells with non-self MHC haplotypes, nor bind free antigens. This phenomenon — MHC Restriction — was identified in a classic 1974 experiment by Zinkernagel and Doherty (which led to the 1996 Nobel Prize). Using cytotoxic T lymphocytes (CTLs) specific for virally infected cells, immunologists found that the CTLs lysed target cells in vitro but did not bind free virus antigens. Zinkernagel and Doherty built upon this, realizing that CTLs are also specific only for virally infected cells presenting a self MHC molecule. |
| Heterodimers | Another study was performed to isolate the T cell receptor. Researchers created monoclonal antibodies to various T cell clones, then isolated antibodies which bound a specific clone population. Assuming that the T cell clones differed only in their T cell receptors, it was concluded that these antibodies bound the T cell receptors. This approach illuminated that T cell receptors are heterodimers, and the two chains were labeled α and β. |
| αβ and γδ | αβ heterodimers were isolated from the membranes of various T cell clones. Antibodies were generated to these heterodimers; some antibodies bound to only one clone population, while other antibodies bound all the clone populations. This suggested that T cell receptors contained variable and constant regions, which is logical due to their antigen specificity. Later, a second T cell receptor heterodimer was identified, and its two chains were labeled δ and γ. Most T cells expressed αβ heterodimers, but depending on the organ there can be just as many or more δγ T cells. |
| TCR cDNA | Immunologists Hedrick and Davis next isolated the genes encoding the T cell receptor. After isolating mRNA from T cells, the researchers eluted only mRNA associated with membrane-bound polyribosomes (as opposed to free cytoplasmic ribosomes). This step removed ∼97% of total cellular mRNA. Next, reverse transcriptase synthesized labeled cDNA probes from these mRNA samples. The following step was DNA subtractive hybridization: they hybridized labeled B cell mRNA to the cDNA probes; unhybridized labeled cDNA was thus unique to T cells. After eliminating ∼97% of cellular mRNA, this step removed ∼98% of cDNA probes. |
| TCR Genes | Approximately ten cDNA probes remained, and it was assumed that within these probes were the genes encoding the T cell receptor. The probes were used to identify genomic DNA in T cells and other cells; a certain region of DNA was found to rearrange in T cells but not in any other cells. This region was putatively the T cell receptor gene: it encoded a membrane-bound protein, was expressed only in T cells and rearranged only in T cells. It was later found that the cDNA clone encoded the β chain — subsequent research identified α, γ and finally δ chain genes. |
T cells — also known as T lymphocytes or thymocytes — are broken into two classes: CD8+ T cells, which are T cells which express CD8 cell surface proteins; and CD4+ T cells, which express CD4 cell surface proteins and are called TH cells. All T cells contain a T cell receptor (TCR), which is activated only by antigens embedded in an MHC complex. TcRs and antibodies are similar in that they both specifically bind antigens, but they are critically different because TcRs only bind MHC-associated antigens and an antibody will bind a free (floating) or membrane-bound antigen. When a TcR is activated by an antigen-MHC complex, its associated T cell will release a plethora of cytokines. Cytokines are used by the immune system to communicate within itself and with other tissues. To fully understand T cells, please read about T cell development upon completion of this article.
T cells expressing CD4 surface protein are called helper T cells (aka TH cells, TH cells, effector T cells or any variation with the word ‘lymphocyte’ in place of ‘cell’. TH cells begin as TH0 cells, but then differentiate into either TH1, TH2 or TH17 cells. TH1 and TH2 cells cross-regulate each other. However, an individual TH cell only produces one cytokine; thus, TH1 and TH2 effects are on the level of the entire body.
| TH1 Cells | TH1 cells are inflammatory cells which secrete: IL-2; IFN-γ, which inhibits TH2 proliferation and interferes with IL-4 effects; and TFN-β, which activates macrophages. In less technical words, TH1 cells activate macrophages and stimulate T cell responses. |
| TH2 Cells | TH2 cells are helper T cells which secrete: IL-4, which interferes with IFN-γ effects; IL-10, which inhibits IFN-γ synthesis; and IL-5, which stimulates B cell and eosinophil growth and differentiation. |
| TH17 Cells | Located on mucosal surfaces, TH17 cells express CD4+ surface proteins (they are CD4+) and fight bacterial infections. TH17 cells secrete IL-17, an important inflammatory cytokine, and IL-22, a cytokine inducing production of antibacterial defensins. TH17 differentiation (and maintenance) is stimulated by IL-23 and is distinct from TH1 and TH2 cell production; TH17 differentiation is inhibited by IFN-γ and IL-4. |
| TS Cells | Separate from TH cells are CD4+CD25+ suppressor T (TS) cells. TS cells have a subpopulation of regulatory T (Treg) cells which suppress immune responses. This is critical to prevent autoimmune diseases, and to help control the damaging immune mechanisms (such as inflammation) from overperforming. |
There are multiple diseases related to TH1 and TH2 cells. Experimental Allergenic Encephalomyelitis (EAE) is caused by a faulty TH1 response to myelin basic proteins of the central nervous system. Leprosey results from an inappropriate TH2 cell activity and is carried by a dominant allele. Allergies are causes by TH2 responses that lead to preferential IgE production. Also, as AIDS progresses, TH1 cells become TH2 cells, and TH17 cells rapidly disappear into the gut.
Virgin T cells migrate from the thymus through the blood, eventually flowing into capillaries at a lymph node and then through post-capillary venules (PCVs) into the node itself. T cells sometimes activate and exit lymph nodes to patrol the bloodstream. These circulating, activated T cells have two fates: extravasation to an infection site (detailed) below, following by reversion to a memory state and flow through the lymph to the nearest lymph node; or, if no extravasation occurs, reversion to a memory state after a few days of circulation, followed by crossing out of blood through PCVs and then flow to the nearest lymph node.
T cells expressing CD8 surface protein are called cytotoxic T cells (aka TC cells, TC cells or any variation with the word ‘lymphocyte’ in place of ‘cell’). Cytotoxic T cells recognize surface markers on other cells in the body that label those cells for destruction. In this way, TCs help to keep virus-infected or malignant cells in check. Also, manipulation of antigen presenting TC cells is important for immune system targeting of tumor cells.
| Next Steps | Study T cell maturation, activation and proliferation and differentiation. |
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Traveling along chemical signals, thymocyte precursors migrate via blood from the bone marrow to the thymus. These cells have not yet rearranged their T cell receptor (TCR) genes and thus lack the T cell receptor (let alone CD3, CD4 or CD8); still lacking any characteristics of thymocytes, these immature T cells begin to divide furiously before individually undergoing four stages denoted double negative (DN) 1-4, named as such because the cells still lack CD4 and CD8 (CD4-CD8-). The four different DN steps — taking a total of ∼3 weeks — are described below, followed by the double-positive state (CD4+CD8+) and finally mature single-positive CD4+CD8- or CD4-CD8+ cells.
| Stage | Phenotype | Overview | ||
| DN1 | c-kit+ | CD25- | CD44high | Double-negative DN1 cells enter the thymus and proliferate as they become DN2 cells. |
| DN2 | c-kit+ | CD25+ | CD44low | TCRβ genes begin rearranging first, followed by TCR γ and δ (but not α) genes by ∼14 days. |
| DN3 | c-kit- | CD25+ | CD44- | In DN3 cells, TCR γ, δ and β rearrangement progresses. Immature thymocytes not expressing Notch proteins do not mature past DN3. At the transition from DN2 to DN3, γδ thymocytes become mature, undergoing very little more change; γδ cells frequently remain double-negative, and never become CD4+. DN3 αβ thymocytes halt proliferation, and β chains combine with a 33kD pre-Tα chain (aka gp33) and associate with CD3 to form the pre-T cell receptor (pre-TCR). The pre-TCR activates the following processes:
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| DN4 | c-kit- | CD25- | CD44- | The DN4 state occurs quickly after β rearrangement completes in DN3 cells. CD4 and CD8 coreceptors begin expression, leading to the double-positive state (CD4+CD8+) |
| DP | CD4+ | CD8+ | The double-positive state (DP) involves rapid proliferation. This leads to a large population of T cell clones with identical TCR β chain rearrangements. Once proliferation stops, RAG-2 expression is activated and TCR α chain rearrangement occurs. This leads to tremendous diversity, as each TCR β chain rearrangement is now bound to a unique α chain rearrangement. | |
| CD4+CD8-/CD4-CD8+ | DP cells proceed through thymic exclusion (described below), and surviving thymocytes expressing the αβ TCR-CD3 complex mature into single-positive CD4 or CD8 cells. | |||
Thymic selection is a two-step process: positive selection, which induces apoptosis in thymocytes whose TCR cannot bind self MHC molecules; and negative selection, which induces apoptosis in thymocytes which bind self MHC molecules too well or in presence of a self peptide. Positive selection results in MHC restriction, and negative selection results in self-tolerance (meaning the thymocytes will not attack healthy self cells).
| Selection | Overview |
| Positive | Positive selection ensures the T cell only reacts to self MHC (MHC restriction) and takes place in the cortical region of the thymus, with immature thymocytes binding (or not) to MHC molecules on cortical epithelial cells. Upon binding to the MHC molecule, the thymocyte receives a protective signal that prevents apoptosis; if the thymocyte does not bind an MHC molecule, it proceeds with apoptosis. |
| Negative | Occurring after positive selection, negative selection ensures the T cell is does not react to self peptides. Dendritic cells and macrophages bearing Class I and II MHC molecules interact with thymocytes that bind self-antigen-MHC complexes or MHC complexes alone. Binding leads to apoptosis. |
There are two proposed models as to how CD4+CD8+ cells mature into CD4+CD8- or CD4-CD8+ cells: the instructive model and the stochastic model. Neither model has been definitely proven nor disproven. The instructive model postulates that double-positive cells interact with either a Class I or a Class II MHC molecule, and are somehow signaled to differentiate into either CD4 or CD8 cells. The schotastic model postulates that repression of CD4 or CD8 is random and has nothing to do with TCR specificity. Only thymocytes whose TCR and coreceptor bind the same MHC molecule continue to mature.
| Next Steps | Study T cell activation and proliferation and differentiation. |
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The primary response is activation of naive thymocyte by a peptide-MHC complex. ∼48 hours after activation, the thymocyte enlarges into a blast cell and repeatedly divide to form a population of genetically identical cells (clonal expansion). Remember the G proteins described under transduction, and that G proteins help trigger the G1 phase of the cell cycle). IL-2 concentration increases 100x in activated cells, helping induce up to 2-3 daily divisions for 4-5 days as well as thymocyte differentiation into either effector T cells or memory T cells.
| Cell Type | Overview |
| Effector | Derived from naive cells and memory cells, effector T cells have short lives of only a few days or a few weeks and carry out specialized functions including: cytokine secretion; B-cell help, performed by activated CD4+ cells, aka TH cells; and cytotoxic killing, performed by activated CD8+ cells, aka CTLs. Effector thymocytes and naive thymocytes express different cell membrane molecules, leading to different recirculation cycles. |
| Memory | Derived from naive cells and effector cells, memory T cells are long-lived, quiescent cells with heightened reactivity to subsequent antigen exposure. Like naive cells, they are arrested in G0; however, they are activated more easily and by more cell types than naive thymocytes. Their cell surface markers are not distinguishable different from effector cells, although their recirculation cycles are different from naive and effector cells. |
Over 98% of all thymocytes die during positive and negative selection, with the remaining cells entering the circulatory system to differentiate into effector or memory thymocytes. These T cells express two cell-surface proteins, Fas and Fas ligand (FasL), which are both essential for apoptosis via the Fas pathway. Upon activation, thymocytes increase Fas/FasL expression — the result is that over-stimulated cells are killed. This is essential for avoiding over-proliferation of thymocytes, and also for killing any self-reactive cells which avoided thymic selection.
| Next Steps | Study the T cell receptor. |
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One of the central mechanisms of the immune system is thymocyte activation, clonal expansion and differentiation (into either effector or memory cells). T cells are activated by binding of the TCR-CD3 complex to a processed antigen peptide bound to a Class I (CD8 cells, aka cytotoxic T cells) or a Class II (CD4, aka helper T cells) MHC molecule. A cascade of biochemical events is initiated, inducing the resting thymocyte to proliferate and differentiate. Induction occurs in two steps: initiation and signal generation, described below. This leads to expression of various gene products, listed below by how early they are expressed after initiation.
| Event | Overview | ||||||||||
| Initiation | The TCR-CD3 complex binds the peptide-MHC complex, bringing the thymocyte and the antigen-presenting cell together. Next, CD4 or CD8 coreceptors bind invariant regions of the MHC molecule. At this point, the tyrosine kinase p56Lck is brought close to the cytoplasmic tails of the TCR. p56Lck is essential for initiation of TCR signaling, and in a resting thymocyte it is sequestered from the TCR in a lipid raft. Upon binding of the coreceptors to their ligands, however, the lipid raft moves to the TCR so that p56Lck can phosphorylate the ITAMs of the TCR complex. Phosphorylated tyrosines in the ITAMs of the CD3 ζ chain bind and activate ZAP-70 and other molecules, which catalyzes phosphorylation of various membrane-associated adaptor molecules. Phosphorylated membrane-associated adaptor molecules aid recruitment of signal transduction pathway mediators. Initiation triggers a litany of signal transduction pathways, described immediately below. | ||||||||||
| Transduction |
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Naive thymocytes are those which have not yet encountered a peptide-MHC complex. Arrested in G0, naive thymocytes have condensed chromatin, minimal cytoplasm and little transcriptional activity. They continuously recirculate through blood, lymph an lymph nodes. To activate, they need additional costimulatory signals to those described above. Signal 1 is the initial interaction between TCR-CD3 and peptide-MHC. Signal 2 is is provided by thymocyte CD28 and CD152 interaction with B7 proteins on the antigen-presenting cell. B7 proteins are constitutively expressed on dendritic cells, and in activated macrophages and activated B cells. B7 binds two proteins — CD28 and CD152 — which are both found on thymocyte membranes as disulfide-linked dimers. Binding of CD28 induces the cell to activate, while binding of CD152 represses activation. CD152 has a much higher B7 affinity than CD28, and its expression is activated by binding of CD28. Thus, CD152 is essentially a braking mechanism that maintains homeostasis; CD152 knockout mice have enlarged lymph nodes, enlarged spleen and die 3-4 weeks after birth. In the absence of a costimulatory signal, an unresponsive state called clonal anergy ensues (as opposed to clonal proliferation).
| Gene Group | Overview | ||||||||||||||||||||||||||||||||||||||||||
| Immediate | Immediate genes are expressed within ½ hour of antigen recognition, and encode mostly transcription factors.
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| Early | Early genes are expressed within 1-2 hours of antigen recognition. and encode mostly cytokines.
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| Late | Late genes are expressed more than 2 days after antigen recognition, and encode various adhesion molecules.
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In essence, activation occurs when a dendritic cell simultaneously binds itself to a TH‘s antigen receptor (primary signal) and to its CD28 receptor (secondary signal). This signals to the dendritic cell that the antigen is foreign (dangerous) and that the next encountered cytotoxic thymocyte must be activated. Other times, dendritic cells are directly activated by an antigen via toll-like receptors and activate cytotoxic thymocytes — this is a critical example of how innate immunity activates adaptive immunity.
| Next Steps | Study T cell clonal expansion and differentiation. |
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