L16 - Lymphocyte development: T-cell development

What Structural Features do BCRs and TCRs share?

1. Antigen-binding sites

2. Variable regions

3. Constant regions

4. Transmembrane regions

How do BCRs and TCRs differ in Antigen Binding and Secretion?

• BCR: Bivalent (two identical antigen-binding sites)

  • Immunoglobulins can be secreted as antibodies

• TCR: Monovalent (one antigen-binding site)

  • Never secreted and does not cross-link antigens

What is the Basic Structure of an αβ TCR?

A heterodimer of alpha (α) and beta (β) chains, each containing a variable and constant region

• The TCR resembles a membrane-bound Fab fragment

What are the Two Classes of TCRs, and Which predominates?

1. αβ TCR: Predominant type (over 95%)

2. γδ TCR: Comparatively enriched in ruminants, where it can account for up to 35% of peripheral T cells

Where does TCR Recombination occur, and What Mechanisms generate Diversity?

In the thymus

• Through

  1. Random V(D)J gene-segment recombination

  2. Random nucleotide addition at the joining sites

How is TCR Recombination similar to BCR Recombination?

BOTH use DNA-level rearrangement of V, D, and J gene segments to generate millions of antigen receptor specificities

• BCR rearrangement occurs in the bone marrow, while TCR rearrangement occurs in the thymus

What Two Properties are Evaluated during Thymic Selection?

The goal is to retain functional T cells that are tolerant to self

1. Functionality: Can the TCR recognize antigen presented by self-MHC?

2. Self-reactivity: Does the TCR react too strongly to the body's own antigens?

QUESTION: What Types of Antigens and Epitopes do TCRs recognize?

Most conventional TCRs recognize processed, fragmented peptide epitopes presented by MHC molecules

How does Antigen Recognition differ between BCRs and TCRs?

• BCR: Directly recognizes native antigens, including linear and conformational epitopes

• TCR: Recognizes processed peptide fragments presented by MHC molecules

What Types of Molecules can BCRs recognize?

• Proteins

• Carbohydrates

• Lipids

• Nucleic acids

What are Linear and Conformational Epitopes?

• Linear: Continuous amino acid sequences

  • Do not require the original three-dimensional protein structure

• Conformational: Discontinuous amino acids brought together by protein folding

  • Depend on tertiary or quaternary structure

What are the Two Universal Rules of Immunology?

1. Nothing happens without a receptor–ligand interaction

2. Immune responses are usually mediated by cytokines and chemokines

What is an Antigen-Presenting Cell (APC), and Which Three Cells are Professional APCs?

An APC processes antigens and displays peptide fragments on MHC molecules for T-cell recognition

• Professional APCs

  1. Dendritic cells

  2. Macrophages

  3. B cells

How do Dendritic Cells connect Innate and Adaptive Immunity?

They present processed antigens to T cells, connecting innate immune detection with adaptive immune responses

What is MHC, and What is Its Main Function?

Major histocompatibility complex (MHC) consists of molecules that display peptide fragments for recognition by T-cell receptors

Where are MHC Class I and MHC Class II Expressed?

• MHC I: Nearly all nucleated cells

• MHC II: Primarily professional APCs (dendritic cells, macrophages, and B cells)

How do NK Cells use MHC Class I to Monitor Cells?

MHC I provides inhibitory signals to NK cells

• Reduced MHC I expression, as may occur in infected or cancerous cells, can remove inhibition and permit NK-cell killing

What are the Main Steps of Antigen Processing and Presentation by an APC?

1. Phagocytosis of antigen

2. Lysosomal digestion

3. Peptide loading onto MHC

4. Presentation on the cell surface for TCR recognition

What Structural Feature do MHC I and MHC II share for Antigen Presentation?

BOTH contain a peptide-binding cleft that holds peptide fragments for TCR recognition

What is MHC Polymorphism, and Why is It Beneficial?

MHC polymorphism is extensive genetic variation in MHC molecules among individuals

• Different MHC variants present different pathogen peptides → increasing the diversity of immune responses across a population

How does MHC Diversity differ from BCR/TCR Diversity?

• BCR/TCR: Generated through V(D)J recombination

  • Producing millions of receptor specificities

• MHC: Inherited genetic variation

  • With thousands of variants across populations; an individual's MHC genotype is fixed for life

QUESTION: Which MHC Class does CD4 recognize, and What Type of T Cell expresses CD4?

CD4 recognizes MHC class II and is expressed by helper T cells

QUESTION: Which MHC Class does CD8 recognize, and What Type of T Cell expresses CD8?

CD8 recognizes MHC class I and is expressed by cytotoxic T cells

What are the Main Functional Differences between CD4+ and CD8+ T Cells?

Memory trick: CD4 × MHC II = 8; CD8 × MHC I = 8.

• CD4+ helper T cells: Coordinate immune responses

• CD8+ cytotoxic T cells: Kill infected or abnormal cells

What are the Primary and Secondary Lymphoid Organs, and What are Their Functions?

• Primary lymphoid organs: Bone marrow and Thymus

  • Sites of lymphocyte development and maturation

• Secondary lymphoid organs: Lymph nodes, Spleen, and Mucosa-associated lymphoid tissues

  • Sites where lymphocytes encounter antigens and initiate adaptive immune responses

What is the Main Function of the Thymus in T-Cell Development?

The thymus is the primary lymphoid organ where T-cell precursors undergo

• TCR rearrangement

• CD4/CD8 lineage commitment

• Selection for functional, self-tolerant receptors

What are the Three Main Stages of T-Cell Development based on CD4/CD8 Expression?

Sequence: DN → DP → SP

1. Double negative (DN): CD4−, CD8−

2. Double positive (DP): CD4+, CD8+

3. Single positive (SP): CD4+ or CD8+

What happens to T Cells during Co-Receptor Selection?

1. Developing T cells progress from expressing neither CD4 nor CD8 to expressing both

2. Finally to expressing only one → becoming either CD4+ helper or CD8+ cytotoxic T cells when leaving the thymus

What is Positive Selection, and Why is It Necessary?

Positive selection preserves developing T cells capable of recognizing the body's own MHC molecules

• This ensures that mature T cells can later recognize peptides presented by self-MHC

What happens to T Cells during Positive Selection based on Their Recognition of Self-MHC?

• Weak-to-moderate recognition: Receives a survival signal

• Insufficient recognition: Dies by neglect

• Strong recognition of self-peptide–MHC: Can trigger negative selection

What is Negative Selection, and Why is It Necessary?

Negative selection eliminates strongly self-reactive T cells before they leave the thymus, helping prevent autoimmunity

What are Tissue-Specific Antigens (TSAs) and Medullary Thymic Epithelial Cells (mTECs)?

• TSAs: Antigens normally associated with particular tissues, such as insulin from the pancreas

• mTECs: Specialized thymic epithelial cells that express tissue-specific antigens to help identify self-reactive T cells

What is Aire, and What is Its Role in Negative Selection?

Aire is a transcription factor that allows mTECs to express tissue-specific antigens normally found outside the thymus

• Helping identify T cells that react strongly to self

What are the Possible Outcomes of Negative Selection?

• Strong self-reactivity: T cell undergoes apoptosis or may develop into a regulatory T cell (Treg)

• No strong self-reactivity: T cell can survive and mature

What is the Main Function of Regulatory T Cells (Tregs)?

1. Suppress immune responses

2. Help maintain self-tolerance, reducing the risk of autoimmunity

What is Central Tolerance, and How does It differ from Peripheral Tolerance?

• Central tolerance: Elimination or control of self-reactive lymphocytes during development in primary lymphoid organs

• Peripheral tolerance: Additional mechanisms that control self-reactive lymphocytes outside primary lymphoid organs

How can Failure of Thymic Selection contribute to Autoimmunity?

If strongly self-reactive T cells escape thymic selection and are not adequately controlled by peripheral tolerance, they may attack the body's own tissues

What are the Four Main Outcomes of T-Cell Maturation in the Thymus?

Mature T cells:

1. Have a functional rearranged TCR

2. Are committed to a CD4+ or CD8+ lineage

3. Can recognize self-MHC through positive selection

4. Have undergone selection against strong self-reactivity through negative selection

What is a Naïve T Cell, and Where does It travel after Leaving the Thymus?

A naïve T cell is a mature T cell that has NOT yet encountered its specific antigen

• It circulates through secondary lymphoid organs, including lymph nodes, spleen, and mucosa-associated lymphoid tissue

QUESTION: How many epitopes can an BCR bind?

ANSWER → Two

• An immunoglobulin is bivalent, meaning it has two antigen-binding sites that recognize the same epitope

• TCR recognition is monovalent, unlike the bivalent immunoglobulin

QUESTION: Which Types of Epitopes can BCRs recognize?

ANSWER → Linear and conformational epitopes

• Why: BCRs can directly bind native antigens, including structures created by protein folding

QUESTION: What are the Two Types of Epitopes based on Their Structure?

ANSWER → Linear (continuous) & conformational (discontinuous)

• Why: Linear epitopes consist of consecutive amino acids, while conformational epitopes depend on protein folding

QUESTION: Which MHC Class does CD4 or CD8 recognize?

ANSWER →

• CD4 → MHC II

• CD8 → MHC I

Why: These coreceptors help T cells recognize the appropriate peptide–MHC complex

QUESTION: During T-Cell Development in the Thymus, What is the Main Purpose of Positive Selection?

a) To activate T cells that recognize a foreign pathogen 

b) To preserve T cells capable of recognizing self-MHC molecules 

c) To eliminate T cells that strongly recognize self-antigens 

d) To generate diverse T-cell receptors through gene rearrangement

ANSWER → b) To preserve T cells capable of recognizing self-MHC molecules

• Why: Positive selection ensures T cells can recognize self-MHC molecules, which is necessary for future antigen recognition. Choice C describes negative selection

QUESTION: What Structural Feature is shared by Both αβ TCRs and Immunoglobulins (Antibodies)?

a) Antigen-binding sites located in the constant regions 

b) Two antigen-binding sites per receptor molecule 

c) Two heavy chains and two light chains 

d) Antigen is bound by CDR3 of antigen receptor

ANSWER → d) Antigen is bound by CDR3 of the antigen receptor

• Why: Both receptors have variable regions containing complementarity-determining regions (CDRs)

• CDR3 is particularly important for antigen-binding specificity.

TCR/BCR vs. MHC Diversity

	TCR/BCR	MHC
How is it created	V(D)J recombination	Inherited
Number of specificities	Millions	Thousands
Plasticity	Dependent on exposure history	Fixed for life

TABLE 1. BCR vs. TCR Comparison

Feature	BCR	TCR
Found on	B cells	T cells
Structure	2 heavy + 2 light chains	αβ or γδ heterodimer
Binding sites	2 (bivalent)	1 (monovalent)
Variable region	Determines specificity	Determines specificity
Constant region	Determines antibody class/function	Structural region
Antigen recognition	Directly binds native antigen	Recognizes processed peptide–MHC
Antigen types	Proteins, carbohydrates, lipids, nucleic acids	Primarily peptides
Epitope types	Linear and conformational	Linear peptides
Secreted form	Antibody	Never secreted
Receptor diversity	V(D)J recombination	V(D)J recombination
Development	Bone marrow	Thymus

Key concept: BCRs directly recognize antigens, while conventional αβ TCRs recognize processed peptides presented by MHC molecules.

TABLE 2. TCR Types and Generation of Diversity

Feature	αβ TCR	γδ TCR
Chains	Alpha + beta	Gamma + delta
Relative abundance	Predominant type	Less common in many species
Veterinary significance	Main focus of lecture	Enriched in ruminants
Antigen recognition	Primarily peptide–MHC	Can differ from conventional αβ recognition

TABLE 2a. Mechanisms of Initial TCR Diversity

Mechanism	Description
Somatic recombination (V(D)J)	Randomly rearranges receptor gene segments
Combinatorial diversity	Different combinations of V, D, and J segments
Random nucleotide addition	Adds nucleotides at gene-segment junctions
Junctional diversity	Additional receptor variation produced at joining sites
Location	Thymus
Result	Millions of different TCR specificities

Remember: Somatic recombination and V(D)J recombination refer to the same process. Random nucleotide addition increases diversity further.

TABLE 3. MHC Class I vs. MHC Class II

Feature	MHC Class I	MHC Class II
Expression	Nearly all nucleated cells	Primarily professional APCs
Antigen source	Primarily intracellular proteins	Primarily extracellular proteins
Antigen processing	Cytosolic pathway	Endosomal/lysosomal pathway
Recognized by	CD8+ T cells	CD4+ T cells
T-cell subset	Cytotoxic T cells	Helper T cells
Additional role	Provides inhibitory signals to NK cells	Presents antigens to helper T cells

Memory trick: CD4 × MHC II = 8; CD8 × MHC I = 8.

The intracellular/extracellular distinction is a useful general immunology connection; the lecturer primarily emphasized MHC distribution and CD4/CD8 matching.

TABLE 4. CD4+ vs. CD8+ T Cells

Feature	CD4+ T Cell	CD8+ T Cell
Name	Helper T cell	Cytotoxic T cell
Coreceptor	CD4	CD8
MHC recognition	MHC II	MHC I
Main function	Coordinates immune responses	Kills infected or abnormal cells
Final developmental stage	Single positive	Single positive

Key concept: T cells become committed to the CD4+ or CD8+ lineage during development in the thymus.

TABLE 5. Stages of T-Cell Development in the Thymus

Stage	CD4	CD8	Key Concept
Double negative	−	−	Immature thymocyte
Double positive	+	+	Expresses both coreceptors; undergoes selection
Single positive	+ or −	− or +	Committed to helper or cytotoxic lineage

Remember: DN → DP → SP. The lecturer said you do not need to memorize the exact anatomical location of each stage.

TABLE 6. Positive vs. Negative Selection

Feature	Positive Selection	Negative Selection
Main purpose	Ensure T cells recognize self-MHC	Prevent strong self-reactivity
What is tested?	Ability to recognize self-peptide–MHC	Strength of recognition of self-antigens
Survival outcome	T cells with sufficient recognition survive	Strongly self-reactive T cells are eliminated or diverted
Failure outcome	Death by neglect	Apoptosis or possible Treg development
Importance	Produces functional, MHC-restricted T cells	Promotes central tolerance

TABLE 6a. TCR Recognition Strength and Outcome

TCR Recognition	Outcome	Explanation
No/insufficient recognition	Death by neglect	Cannot adequately recognize self-MHC
Weak-to-moderate recognition	Positive selection	Receives survival signal
Strong recognition	Negative selection	Potentially self-reactive
Strong recognition with appropriate developmental signals	Possible Treg development	Helps suppress immune responses

Memory trick: Positive selection = keep the useful T cells. Negative selection = remove dangerous self-reactive T cells.

TABLE 7. Aire, mTECs, and Central Tolerance

Component	Definition	Main Function
mTECs	Medullary thymic epithelial cells	Express tissue-specific antigens in thymus
TSAs	Tissue-specific antigens	Represent proteins normally expressed in other organs
Aire	Autoimmune regulator transcription factor	Promotes expression of tissue-specific antigens by mTECs
Negative selection	Removal of strongly self-reactive T cells	Prevents potentially harmful T cells from maturing
Tregs	Regulatory T cells	Suppress immune responses and help maintain tolerance

TABLE 7a. How Aire Supports Central Tolerance

1. Aire in mTECs

   • Promotes tissue-specific antigen expression

2. Self-Antigen Presentation

   • Developing T cells encounter self-antigens

3. Negative Selection

   • Strongly self-reactive T cells are identified

4. Central Tolerance

   • Reduces the risk of autoimmunity

Key concept: Aire allows developing T cells to be tested against antigens from tissues throughout the body, even though development occurs in the thymus.

TABLE 8. Central vs. Peripheral Tolerance

Feature	Central Tolerance	Peripheral Tolerance
Location	Primary lymphoid organs	Outside primary lymphoid organs
T-cell location	Thymus	Peripheral tissues and secondary lymphoid organs
Timing	During lymphocyte development	After lymphocytes leave primary organs
Main purpose	Eliminate/control self-reactive cells during development	Control self-reactive cells that escape central tolerance
Mechanisms emphasized	Negative selection; Treg development	Additional control of self-reactive cells
Failure can lead to	Autoimmunity	Autoimmunity

Remember: Central tolerance is not perfect, so peripheral tolerance provides another layer of protection.