Lecture 12: T Cell Receptor Rearrangement and Function

1961-2: Jacques Miller hypothesized that T cells are generated in the thymus and are critical for adaptive immunity based on thymectomy experiments, where removal of the thymus affected immune function.
Hypothesis: Similar to B cells, T cells utilize a clonal recognition structure.
1981-3: Groundbreaking work by Alan Harris, Jim Golding, Ellis Reinherz, and Jim Allison characterized the structure of TCR.
1984-5: Mark Davis and Tak Mak isolated and mapped the β-chain TCR genes in both mice and humans.

Structural Features:
- Polypeptides with linked chains.
- Resembles a membrane-bound Fab, consisting of TCRα and TCRβ chains (or alternately TCRγ and TCRδ).
- Variable Regions: Responsible for recognizing antigens.
- Constant Regions: Provide structural and functional stability.
Gene Rearrangement:
- Both TCRs and BCRs undergo RAG-dependent somatic gene recombination to create variable regions.
Unique Characteristics of TCR:
- Contains only one antigen binding site.
- Does not generate a soluble form (unlike antibodies).
- Does not undergo somatic hypermutation or class switching.

Comprises roughly ~90% of T cells.
TCRs generated from gene loci encoding the TCR α chain (Chromosome 14) paired with the TCR β chain (Chromosome 7).
Recombination Process:
- Involves Variable (V), Diversity (D), and Joining (J) segments leading to diverse variable regions with specificity.
- Regulated by the RAG-1/2 complex, which acts on the flanking recombination signal sequences (RSS).
The TCR β chain recombines D-J segments before pairing with the V segment, during which P- and N-nucleotides are added via Artemis and terminal deoxynucleotidyl transferase (TdT).

Comparative Analysis with Immunoglobulins (BCRs):
- TCRs exhibit a greater number of gene segments contributing to recombination (combinatorial diversity).
- Enhanced junctional diversity potential through the use of N- and P-nucleotides.
- TCR pairs (α and β) have higher N- and P-nucleotide addition compared to BCR light chains (where TdT is downregulated at rearrangement).

Accounts for ~10% of T cells, originating from gene loci that encode TCRγ (Chromosome 7) and TCRδ (Chromosome 14).
Noteworthy: TCRδ chain sequences are located within the TCRα locus, and their rearrangement deletes the TCRδ genes.
Allows for the incorporation of two D segments, contributing to increased diversity.

TCR β and δ chains have different RSS spacer sequences flanking the D gene segments.
- This allows for the potential to recombine without a D segment, or with more than one segment, although the latter is not commonly observed in TCR β locus.
D-D Rearrangements: Frequent at the TCR δ locus, which leads to increased diversity in antigen recognition.

Allelic Exclusion: This prevents the rearrangement of more than one successful TCR β or γ chain in mature T cells.
TCR chains are expressed as transmembrane disulfide-linked polypeptides, with no soluble forms produced.
Association with CD3 Complex Proteins:
- TCR chains associate with proteins: CD3γδε (linked genes on Chromosome 11) and CD3ζ (Chromosome 1).
- Dimers form: γε, δε, and ζζ, essential for TCR transport out of the endoplasmic reticulum (ER).
- CD3 complex contains immunoreceptor tyrosine-based activation motifs (ITAMs).

CD4 Co-Receptor: Composed of one chain with four domains, containing a hinge in the middle.
CD8 Co-Receptor: Consists of linked α and β chains.
Both are vital for binding to MHC class II (CD4) and MHC class I (CD8), respectively.

Mechanism: TCR:MHC clustering with CD4/CD8 helps stabilize the interaction and leads to recruitment and activation of Lck (a kinase).
Phosphorylation: Lck phosphorylates ITAMs in the cytoplasmic tails of the CD3 complex.
ZAP70 activation: ZAP70, a kinase, is recruited and phosphorylated, facilitating downstream signaling through phospholipase C gamma (PLCγ), yielding transcription factor activation (specifically AP-1, NF-kB, and NFAT-1) which promotes cell survival and proliferation.

TCR:MHC signaling alone is insufficient for full T cell activation; this leads to anergy (non-responsiveness).
Costimulation (Signal 2):
- B7 molecules (CD80 and CD86) are upregulated on activated antigen-presenting cells (APCs).
- These bind to CD28 on T cells, amplifying the signal by 20-30-fold.
- Src kinases (Lck, Fyn) phosphorylate the cytoplasmic tail and recruit PI3K.
Cytokine (Signal 3):
- IL-2 is a critical growth factor, promoting T cell survival and proliferation, upregulated downstream of activation signals.

Pertains to various cytokines (including IL-2, IL-4, IL-5, IL-7, IL-12, IL-23, and IFNγ).
The IL-2 receptor associates with JAK1 and JAK3, leading to STAT 5 activation (predominantly), as well as STAT1 and STAT3.

Mechanism:
- TCR recognizes intracellular antigens presented by MHC/HLA; BCR recognizes extracellular antigens.
Antigen Type:
- αβ TCR mainly interacts with peptides; γδ TCR interacts with native structures.
Composition:
- TCR consists of 2 chains (αβ or γδ); BCR consists of 4 chains (2 light chains - λ, κ; and 2 heavy chains - μ, α, δ, ε, γ).
Binding Sites:
- TCR has 1 binding site; BCR has 2 binding sites for antigens.
Soluble Forms:
- TCR does not produce soluble forms; BCR produces antibodies (soluble forms).
Germline Rearrangements: Both TCR and BCR undergo V(D)J gene rearrangements, which are RAG1/RAG2 dependent.
Junctional Diversity: Both have junctional diversity, but BCR has fewer N-nucleotides.
Somatic Hypermutation: Not present in TCR, but present in BCR.
Associated Molecules: TCR is associated with CD3 complex and CD4 or CD8, while BCR is associated with CD79a and CD79b.