MHC class I and II and antigen processing

Question 1

List the 3 different Antigen-presenting cells (APCs)

Answer 1

• Dendritic cells (the most effective cells
  for the initial activation of naive T cells)
• Macrophages
• B cells

Question 2

Where are dendritic cells mostly found?

Answer 2

The T cell areas of lymph nodes and spleen, under most surface epithelia

Question 3

List the routes of antigen processing and presentation by dendritic cells.
Describe the pathogen, the MHC molecule and the type of naive T cell involved

Answer 3

Question 4

Describe the antigen capture and presentation by dendritic cells.

Answer 4

1. They pick up antigens in peripheral tissues,
2. They migrate to lymph nodes, via afferent
   lymphatics, where they express high levels of adhesion and costimulatory molecules, as well as MHC class II molecules, which allow them to interact with CD4+ TH cells.
3. Once they have migrated, DCs stop synthesizing MHC class II molecules but maintain high levels of MHC class II molecules containing peptides from antigens
4. As they mature, dendritic cells express CCR7 which allows them to localize to the lymphoid tissues.
5. DCs from the skin and the gut have distinctive chemokine receptors, which allow them to selectively recirculate to their own lymphoid organs.
6. As they mature, DCs also increase the expression of key costimulatory molecules, including CD40, CD80 and CD86 (B7-1 and B7-2)

Question 5

What attracts dendritic cells to the lymph
nodes?

Answer 5

• Specialised blood vessels – high endothelial venules (HEV) secrete a chemokine, CCL21 that attracts dendritic cells
• CCL21 also contributes to dendritic cell maturation

Question 6

What is the term for human MHC?

What is the link between the MHC and transplant outcome?

Answer 6

Human leukocyte antigen (HLA)

*Immune responses to transplants are caused by genetic differences between the donor and the recipient
*For transplant compatibility, the most important genetic differences are between MHC-I and MHC-II

Question 7

Role of MHC Class I and MHC Class II
Where are they found?

Answer 7

• Antigen presentation
• MHC class I and class II proteins are found at the cell surface and form a structure that holds antigenic peptides for surveillance by T cells
• MHC class I = recognised by CD8+ cytotoxic T cells
• MHC class II = recognised by CD4+ helper
  T cells

Question 8

What is the major histocompatibility complex?

Answer 8

Large gene complex on chromosome 6 which encodes multiple proteins involved in immune response

Question 9

MHC class I structure

Answer 9

Humans: HLA-A, -B, -C
Tissue distribution: all nucleated cells
Two polypeptides, non-covalently bound:
a glycosylated alpha-heavy chain (45 kDa)
*Inserted in a membrane
*Polymorphic
*a cytoplasmic tail
*three extracellular domains (90aa each), designated alpha 1 (N terminal), a2, and a3

non-covalently associated with Beta2-microglobulin (12 kDa)
*Not inserted in the membrane
*Not Glycosylated
*Not Polymorphic in humans
*essential for expression of MHC class I molecules

Question 10

MHC class II structure

Answer 10

Humans: HLA-DR, -DP, -DQ
Tissue distribution: Professional antigen-presenting cells (dendritic cells, macrophages, B-cells)
Two polypeptides, non-covalently bound
heterodimers of heavy alpha (30–34 kDa) and light beta (26–29 kDa depending on the locus involved) glycoprotein chains
Both Inserted into the membrane
Both Glycosylated
Both Polymorphic

Question 11

Differences between the peptide binding groove of MHC class I and MHC class II

Answer 11

MHC class I bind short peptides that are 8-10 amino acids long (peptide-binding groove has closed ends)

There are no length constraints on peptides
bound by MHC class II (peptide binding groove has open ends), but they are usually 13-24 amino acids long

Question 12

How many alleles are there for each MHC class?

Answer 12

• You have two alleles of each of the MHC class I
  genes (HLA-A, HLA-B, and HLA-C), and three alleles of each of the MHC class II genes (HLA-
  DR, HLA-DQ, and HLA-DP)

Question 13

How will pathogens try and evade immunity?
How successful are they?

Answer 13

Pathogens can evolve to evade immune responses – they will try to avoid making peptides that can be presented
* MHC class I and class II are central to anti-viral immune responses, so why don’t we see many pathogens that have mutated to avoid antigen presentation?
* They do!!!! But, even simple pathogens present multiple peptides and it’s very difficult for them to change all of these.

Question 14

How many alleles are there for each MHC Class I gene?

Answer 14

• HLA-A = 240 alleles
• HLA-B = 470 alleles
• HLA-C = 110 alleles

Question 15

A major difference between MHC class I and class II molecules occurs at the ends of the peptide-binding groove.

Expand on this

Answer 15

Polymorphisms are in the upper peptide-binding part of the MHC protein
* for MHC class I molecules, interactions at the N and C terminals confine the peptide to the cleft;
* for MHC class II molecules, peptides may extend beyond the ends of the cleft.

Question 16

How do the peptides differ between MHC Class I and MHC Class II?

Answer 16

The peptides that bind to MHC class I molecules come from endogenous proteins synthesized within the cell, which are broken down and transported to the endoplasmic reticulum. the internal antigen processing pathways generally produce peptides of an appropriate size to occupy the MHC class I molecule antigen-binding groove

Peptides that bind to MHC class II molecules come from exogenous antigens – proteins that have been internalized by the cell and then degraded. These peptides are less uniform
in size than those that bind to MHC class I molecules and may be trimmed once they have found their way to the MHC class II molecule.

Question 17

List the disease risk-associated HLA’s and their associated disease

Answer 17

Question 18

List the different types of pathogens and:
* where they’re degraded
*relevant MHC class
*relevant T cells
*Effect on presenting cells

Answer 18

Question 19

How are large proteins degraded into the small peptides that bind MHC class I?

Answer 19

• The proteasome is a cytoplasmic structure that digests proteins in the cytosol to generate short peptide fragments
  – the mechanism for removing poorly folded or damaged proteins
  – these have been tagged for destruction by the addition of ubiquitin
  – hence, produces peptides form all internal proteins

The proteasome cannot distinguish between self and non-self-proteins

Question 20

How do peptides produced in the cytosol get to empty MHC class I in the ER?

Answer 20

TAP transporters
* Transporter associated with antigen processing - TAP
* TAP are endoplasmic reticulum membrane proteins
* transport peptides from cytoplasm into ER lumen
* also associates with newly synthesised class I molecules and helps load peptides into their groove
* some proteasome and TAP chains are encoded within MHC region (class II) - TAP1 and TAP2 genes

Question 21

MHC proteins do not adopt a stable conformation until complexed with peptide in their binding groove

How is it stabilized before binding?

Answer 21

Stable complexes must be formed before the MHC can exit the endoplasmic reticulum

Chaperones stabilise the complex and assist with
refolding in the endoplasmic reticulum

Calnexin is the first of these chaperones. Calnexin acts as a chaperone to keep the binding site open

Question 22

What happens after Transporters move peptides to the ER

Answer 22

MHC class-I alpha chains (a) are initially held in the endoplasmic reticulum associated with the chaperone protein calnexin. Once released from calnexin they bind to b2-microglobulin to form a complete MHC class-I molecule.
After combining with b2- microglobulin, they are join a peptide-loading complex, consisting of tapasin (Tps), ER-protein-p57 and calreticulin (CRT). Tapasin also associates with the TAP transporters (I and II).

The peptides may be trimmed by ER-associated aminopeptidases (ERAAP). The MHC class-I molecule with a bound peptide is finally released from the peptide loading complex, to be transported to the plasma membrane

Question 23

What happens if not peptide is bound to MHC Class I

Answer 23

MHC class I molecule complexes lacking peptide is unstable, ensuring that only functionally useful complexes are available for interaction with TCRs

Question 24

Describe the uptake of extracellular proteins for antigen
presentation by MHC class II

Answer 24

Question 25

Describe MHC Class II molecules are produced and transported (before antigen loading)

Answer 25

1) MHC class II molecules are produced in the ER, complexed to a polypeptide called the invariant chain (Ii) which stabilizes the complex and prevents the inappropriate binding of antigen.
2) This complex is transported from the Golgi to an antigen processing compartment (the MIIC compartment), specialized for transporting and loading MHC class II molecules with characteristics of both endosomes and lysosomes.

Question 26

Describe how Class II molecules are loaded with exogenous peptides

Answer 26

1) Exogenous antigens reach the MIIC compartment.
2) The Ii chain is cleaved by cathepsins into small fragments, one of which, termed CLIP (class II-associated invariant peptide), is located in the groove of the class II molecule until replaced by peptides destined for presentation.
2) The exchange of CLIP for other peptides is orchestrated by HLA-DM, an MHC class-II-like chaperone protein
3) HLA-DM binds to the ab-CLIP complex to stabilize
it until it has bound a suitable antigenic peptide
4) MHC-encoded molecule, HLA-DO associates with DM, regulates peptide loading
5) Class-II/peptide complexes are released from the multivesicular bodies as tubular/vesicular structures that fuse with the plasma membrane

Question 27

What happens to cell lines lacking HLA-DM

Answer 27

In cell lines lacking HLA-DM, the class II molecules are unstable and the cells no longer process and present proteins. Their class II molecules end up at the cell surface occupied by CLIP fragments of the invariant chain

Question 28

Why is the invariant chain important for MHC Class II?

Answer 28

Stable complexes must be formed before the MHC can exit the endoplasmic reticulum

BUT
MHC class II proteins do not encounter their peptide antigens until they have left the ER, and must not bind the intracellular peptide fragments they might encounter in the ER

Solution – invariant chain (Ii) binds in their groove, acts as a chaperone and directs MHC class II complexes towards the endosomal compartment of the cell MHC proteins do not adopt a stable conformation until complexed with peptide in their binding groove

Question 29

Describe cross-presentation

Answer 29

• Naïve cytotoxic T cells must be primed by professional
  antigen-presenting cells, usually dendritic cells
• Dendritic cells are known to ingest external antigens,
  process them and then present via MHC class II
• This will not allow the priming of CD8+ T cells
• Dendritic cells have a range of specialised processes to
  allow processing and presentation of external antigens
  through MHC class I

Question 30

What might determine cross-presentation
pathway?

Answer 30

• Size of ingested particles may determine which cross-presentation pathway is used
• Smaller particles: phagosome-cytosol pathway
• Larger particles: vacuolar pathway
• Large particles might also require the recruitment of ‘extra’ membrane from ER, bringing ER proteins with it

The proteasomes found in dendritic cells differ from
those in other cells due to the presence of MHC-encoded subunits
This can alter the repertoire of peptides that they generate

Question 31

How can autophagy also play a role in antigen
presentation

Answer 31

• Autophagy allows controlled degradation & recycling of cellular components
• Degradation occurs in lysosomes
• Resulting peptides can be presented by MHC Class
  I and MHC Class II