Antigen Recognition by T-lymphocytes/ Generation of Diversity in the T cell repertoire

Question 1

What is the definition of an antigen?

Answer 1

Proteins, carbohydrates and lipids that are capable of binding to B-cell receptors, T-cell receptors and innate immune receptors (PRRs e.g lectin receptors, scavenger receptors, toll-like receptors)

Question 2

What is an epitope?

Answer 2

An epitope is a small portion (e.g small peptide) of the antigen that the TCR and BCR (antibody) can bind to

Question 3

Can the immune system recognise the whole antigen?

Answer 3

No normally it is only the epitope that will result in an adaptive immune response

Infection and vaccination usually induce polyclonal T cell and B cell responses as multiple epitopes can be recognised on an antigen

Question 4

What is a key difference between B cells and T cells in terms of recognising antigens?

Answer 4

B cells will recognsie unprocessed antigens which will then cause proliferation of B-cell clones and produce Abs just like original B cell

T cells do not recognise unprocessed (native antigens) and therefore they will not proliferate or produce cytokines

Question 5

How are antigens taken up by APCs?

Answer 5

• Phagocytosis/ Pinocytosis
• Membrane Ig receptor mediated
• Complement receptor mediated
• Fc receptor mediated phagocytosis

Question 6

What are the main immune cells which present antigens?

Answer 6

APC’s which express high levels of MHC Class II can efficiently induce T cell responses

• Monocytes
• Macrophages
• Dendritic Cells
• B-cells

Question 7

Where do monocytes reside and what do they become?

Answer 7

Monocytes are blood circulating immune cells which will differentiate into either macrophages or dendritic cells

Question 8

What is the difference between dendritic cells and macrophages? Which cell is better at being an APC and which cell is better at being a phagocyte?

Answer 8

• They are both highly phagocytic cells that will induce a strong T-cell response and inflammation. (important protection against mycobacterium tuberculosis)
  
  • Macrophages better at phagocytosis and killing pathogens (higher NO production)
  • Dendritic cells better at migrating to lymph nodes (via CCR7) and presenting antigens to T cells

Question 9

Where do macrophages and dendritic cells mainly reside?

Answer 9

• Rare in peripheral blood, mainly enriched in mucosal tissues

Question 10

How do B-cells internalise antigens?

Answer 10

Ig receptor mediated internalisation of antigens (opposed to phagocytosis)

(However primary function of B-cells is to make antibodies via plasma cells, however still good at antigen presentation to other cells)

Question 11

What are the two types of antigen processing?

Answer 11

Exogenous and Endogenous Antigen processing

Question 12

Describe the endogenous/ intracellular pathway

Answer 12

1. Infected (virally or malignant) nucleated cell (can include APC) will produce viral/malignant proteins in the cytosol
2. These will be ubiquitinated and targeted to the proteasome which will recognise proteins and degrade them into peptides
3. Peptides will be imported into the ER lumen via a transporter called TAP protein where there is MHC I production
4. Tapasin will link MHC I molecules to the TAP protein allowing antigens to be easily captured by MHC I molecules as they enter
5. MHC I with succesfully captured peptides will leave the ER and enter the golgi where they form exocytotic vesicles
6. Exocytotic vesicles will fuse with the plasma membrane which can be recognised by CD8+ cytotoxic T cells

Question 13

Describe the exogenous/ extracellular pathway?

Answer 13

1. Dendritic cells/ phagocytes will ingest extracellular microbes e.g bacteria by phagocytosis/ receptor mediated endocytosis to form an endosome/phagsome
2. Endosome will fuse with lysosome to form a phagolysosome which will break antigens down into its peptides
3. MHC II proteins are produced ER and held by an invariant chain to stabalise the MHC II
4. The invariant chain contains a sequence called CLIP which binds to the peptide binding groove
5. MHC II will leave the ER and enter the golgi where it forms an exocytotic vesicle
6. Exocytotic vesicle will fuse with the late-endolysosome where the antigenic peptides are
7. Late endolysosome has a low pH, invariant chain will be broken down leaving CLIP in the MHC II groove
8. HLA-DM has a high affinity for CLIP removing it from the MHC II molecule
9. Antigenic peptides will bind to MHC II complex leaving the late endo-lysosome and fuses with the plasma membrane
10. Infected APC can now be recognised by TCRs on CD4+ T cells

Question 14

What do MHC I and MHC II activate respectively?

Answer 14

MHC I will activate CD8+ Cytotoxic Cells

MHC II will activate CD4+ T helper cells

Question 15

Where is MHC Class I + II expressed?

Answer 15

• MHC I = Expressed on all nucleated cells (including APCs) (so only cell that doesnt is RBCs)
  
  • Binds short peptides
  • Antigens from cytosol (+ cross presentation)
• MHC II = Expressed on APCs and activated T cells
  
  • Binds long peptides
  • Antigens from phagosomes and endosomes

Question 16

What is the tissue specific expression of MHC like?

Answer 16

Epithelial cells of the thymus will express MHC II at high levels (only exception)

Question 17

What is the structure of MHC Class I?

Answer 17

• Large structural part containing alpha 3 domain and beta 2 microglobulin supporting the peptide binding cleft
• Peptide binding cleft containing alpha 1 and alpha 2 domain

Question 18

What is the structure of MHC Class II?

Answer 18

• Large structural part with alpha 2 and beta 2 domains supporting peptide binding cleft
• Peptide binding cleft with alpha 1 and beta 2 domains

Question 19

What are the MHC isotypes?

Answer 19

• Main MHC Class I are HLA-A, HLA-B and HLA-C
• MHC Class II are split into HLA-DM, DO, DP, DQ and DR

Question 20

How does the TCR receptor exist?

Answer 20

Exists in a complex with accessory molecules such as CD3

Question 21

What are similarities between the B-cell receptor and T-cell receptor?

Answer 21

• Belongs to an Ig superfamily
• TCR also has Fab like fragment of BCR
• Large diversity
• Single specificity

Question 22

What are the differences in the B-cell receptor and the T-cell receptor?

Answer 22

• T cell receptor has a lower affinity to their antigens compared to B-cell receptors
• T-cell receptor cannot be released and remains on T-cell surface, BCRs can become circulating antibodies
• TCR has no Fc fragment
  
  • So not cellular function
• TCR only has a single binding site
  
  • Unlike the BCR that has multiple binding sites
• TCR only has two classes (αβ and γδ)
  
  • BCR has 5 classes (IgM, IgD, IgG, IgE, IgA)

Question 23

What differs in the mechanisms of creating receptor diversity between B-cells and T-cells?

Answer 23

B cells can generate receptor diversity by:

1. Before antigen stimulation = somatic recombination
2. After antigen stimualtion = somatic hypermutation

T-cells can generate receptor diversity by:

1. Before antigen stimulation = somatic recombination
2. After antigen stimulation = NONE!!!!

Question 24

Can T-cell receptor diversity be stimulated after antigen stimulation?

Answer 24

No T-cell receptor diversity can only be stimulated after antigen stimulation as they lack somatic hypermutation (explains the lower affinity of the TCR compared to BCR)

Question 25

What are the signals needed for naive T-cell activation?

Answer 25

1. Peptide-MHC 2. Co-stimulation (**CD28** on T-cell and B7 family (**CD80/CD86**) on APC) 3. Cytokine secretion

Question 26

What is the level of importance of the three signals?

Answer 26

* **Signal 1 + 2** → activate naïve T-cells * **Signal 3** is not required for naïve T-cell activation but it important for amplification of the response and determining the **T-cell phenotype**

Question 27

Expand on the three signals that activate naive T-cells

Answer 27

* The main signal **(Signal One)** is delivered from the **APC** by a **peptide-MHC complex** to the **TCR** * The **_co-stimulatory signal_** **(Signal Two)** is delivered from the **APC** by germline-encoded accessory receptors such as the **B7 family (CD8- and CD86)** = although many of these receptors are not fully characterised or understood * Lastly **(Signal Three)** is formed of **cytokines** secreted by the APC to determine the **T-cell phenotype**

Question 28

What do IL-2, IL-4 and IL-23 secreted by the APC determine?

Answer 28

The APC will secrete these cytokines to determine the T-cell phenotype * IL-2 promotes TH1 cells * IL-4 promotes TH2 cells * IL-23 promotes TH17 cells

Question 29

Describe the immunological synapse

Answer 29

* Region of contact and communication between T cell and APC required for T cell activation * Surrounding integrins and accessory molecules (CD80 and CD86 from APC)

Question 30

What co-receptor is expressed depending on the type of T-cell?

Answer 30

Either CD4 (T-helper cell) or CD8 (cytotoxic T-cell)

Question 31

What is the difference between Th1 and Th2 cells?

Answer 31

**Th1 cells** will produce cytokines that **activate macrophages** which kill intracellular pathogens more efficiently than those without cytokines **Th2 cells** will produce cytokines that **activate B-cells** which will produce plasma cells that produce antibodies which neutralise circulating viruses or prevent bacterial colonisation

Question 32

Why are there negative regulators of antigen presentation?

Answer 32

An overly vigourous immune response is harmful to the host and so negative regulators provide an immune checkpoint to limit T-cell activation

Question 33

What are examples of negative regulators of antigen presentation?

Answer 33

CTLA4 (Cytotoxic T-Lymphocyte-Associated Protein 4) PD-L1 (Programmed Death Ligand 1)

Question 34

How do PD-L1 and CTLA-4 inhibit T-cell function?

Answer 34

PD-L1 will bind to and activate PD-1 which is expressed on T-cells leading to production of SHP-2 which dephosphorylates TCR signalling molecules = inhibiting T-cell activation CTLA-4 competes with CD28 on the TCR to bind co-stimulatory molecules on the APC (CD80/86)

Question 35

Describe how T-cells are programmed and self antigens are removed

Answer 35

* T-cells arise from the thymus, which is a ‘school’ for T-cells * T-cells are exposed to self-antigens and tested for reactivityT-cells that can’t bind to self-antigen are deleted = **POSOTIVE SELECTION** * These T-cells are useless because they won’t protect against pathogens * T cells that bind self-antigen-MHC too strongly are also deleted à **NEGATIVE SELECTION** * These T-cells are dangerous because they are too self-reactive

Question 36

What are T-regualtory cells?

Answer 36

Subset of CD4+ T cells which carry an additional CD25 molecule and express the transcription factor FOXP3 Regulating any potential self-reacting T-cells by inducing their apoptosis or deletion

Question 37

How can pathogens impede antigen presentation?

Answer 37

* **Herpes Simplex Virus** * Produces a protein which binds to and inhibits TAP * Prevents viral peptide transfer to ER * **Adenovirus** * Produce protein which binds MHC class I molecule * Prevents MHC Class I molecule from leaving ER