L7. T Cell Receptors and MHC Proteins

Question 1

How can the T cell receptor be expressed compared to a B cell recepetor?

Answer 1

T cell receptors are only expressed on membranes, not as soluble proteins (unlike B cell receptors which can be soluble = antibodies)

Question 2

What are the 2 main categories of T cells and what is their function?

Answer 2

a) T helper cells (CD4 +ve)
>Augment immune responses

b) T cytotoxic cells (CD8+ve)
>Specifically kill infected host cells (Like more specific NK cells, only kill host cells that they recognise specific pathogen infecting)

Question 3

Are the receptor structures on T helper cells (CD4 +ve) and T cytotoxic cells (CD8+ve) the same or different?

Answer 3

Receptor structure on both subpopulations is the same

Question 4

What superfamily of genes are the CD proteins apart of?

Answer 4

Both CD proteins are a part of the immunoglobulin superfamily

Question 5

What part of the T lymphocyte receptor (TCR) allows it to me expressed on the membrane?

Answer 5

C terminus of chains have hydrophobic amino acid residues, anchor into membrane.

Question 6

What 2 chains make up the T lymphocyte receptor (TCR) and how is this similar to an antibody?

Answer 6

> Made of 2 chains: alpha, beta, both members of immunoglobulin super family

> Have structure similar to Fab arm of antigen/ can bind antigen

Question 7

Describe the structure of the T lymphocyte receptor (TCR) and this is similar to an antibody.

Answer 7

> Extracellular domains of the T cell receptor are homologous to the variable and constant regions of immunoglobulins (like Fab arm); so each chain has a constant region which is connected to the outer variable region.

> each V region contains 3 CDRs (hypervariable loops)

Question 8

What are the 2 types of T cell receptor and which is found more commonly in blood?

Answer 8

1) αβ T Cell Receptor (TCR)
>Found more commonly in blood

2) γδ T Cell receptors (gamma, delta chains)
>1-5% of T cell receptors in blood

Question 9

Does γδ T Cell receptors bind to less antigens, why?

Answer 9

Generally these are Less diverse/ can bind to less antigens

Question 10

How is the structure of the variable regions in the alpha and beta chains of TCR similar to antibodies?

Answer 10

> Vα and Vβ domains each have 3 CDRs (1 – 3).

> CDR3 regions of α and β chains are the most variable (same as antibodies)

Question 11

Can TCR not signal like antibodies?

Answer 11

Like antibodies on receptor, T cell receptor itself cannot signal

Question 12

What else must TCR require on he cell surface for signalling to occur?

Answer 12

T cell receptor must have other proteins to have signalling to occur, known as CD3

Question 13

What complex does a TCR form with CD3 and what subunits make this up?

Answer 13

The T cell receptor complex: α and β subunits (TCR), CD3 subunits (ε, δ and γ), together with ζ (zeta) form the complex

Question 14

Why is it important to have CD3 on the cell surface as well as TCRs?

Answer 14

Required for optimal cell surface expression of T cell receptor and signalling

Question 15

How does antigen binding cause signalling in T cells and how is this similar to in B cells, also what is the role of zeta?

Answer 15

> At the cytoplasmic regions of CD3 subunits contain ITAMs (Immunoreceptor Tyrosine Activation Motifs) in their cytoplasmic regions

> CD3 subunits are closely associated to alpha and beta subunits of T cell receptor at surface of T cell meaning when bound to antigen, these motifs can be phosphorylated acting as docking sites for downstream signaling molecules to be phosphorylated to downstream signalling (the same as Igα and Igβ for B cells).

> Zeta amplifies this as itself has 3 ITAMs in humans, which all get phosphyrlated, enhancing the phosphorylation and activation of downstream signaling pathways.

Question 16

How are TCR genes similar to antibody genes?

Answer 16

T cell receptor genes are similar to antibody genes, 2 gene loci

Question 17

Where are the 2 gene loci for TCR genes and what do they encode for?

Answer 17

1) Chromosome 14
>Alpha chain like light chain, made up of 2 exons, V region and many J regions

2) Chromosome 7
>Beta chain like heavy chain of antibody, many V segments, many J segments, and diversity (D) segments,

Question 18

Where do T cells develop and where do they develop receptors?

Answer 18

1) Develop in Bone Marrow

2) Develop receptors in Thymus ( genes in chromosome 14 and 7 are rearranged- somatic recombination)

Question 19

Is somatic recombination the same for TCR as for antibodies, what machinery is used?

Answer 19

> Somatic recombination of TCR V region genes is the same as that for antibodies (but in the thymus)

> Same recombination machinery as that used by developing B lymphocytes. So, Rag1/2 proteins also used.

Question 20

What are 3 ways TCRs share how they get their diversity like antibodies, at each stage how much variation can there be?

Answer 20

1. Multiple copies of V region gene segment [Vn x Jn/Vn x Dn x Jn]
   >Vn x Jn = alpha chain
   >Vn x Dn x Jn = beta chain
2. α x β chain combination [Vα x Jα] x [Vβ x Dβ x J β] = ~ 6 x 106 different T cell receptors
   >Extra diversity comes from junctional diversity (like in antibodies)
3. Junctional diversity = ~ 2 x 1011 Concentrated in the CDR3s of TCR α and β chains Total diversity = ~ 1018

Question 21

Why are Gamma and delta TCRs are a lot less diverse?

Answer 21

number of V, D, and J gene segments available for γδ TCRs, which is smaller compared to αβ TCRs

Question 22

Why does junctional diversity cause greater variation for T cells than for B cells?

Answer 22

Junctional diversity greater for T cell receptors rather than antibodies as T cells generally have more gene segments than antibody genes.

Question 23

Like with antibodies, why is CDR3 the most variable region in TCRs?

Answer 23

Diversity mostly concentrated at CDR3 loops, CDR1/2 is encoded in germline by just V segments but CDR3 is diverse because as corresponds to VJ (alpha chain) or VDJ (beta chain) join so is much more diverse.

Question 24

What is the main difference in what causes diversity for B and T cell receptors?

Answer 24

The V regions of TCRs do NOT undergo somatic mutation

Question 25

What are three suggestions to why TCRs don’t undergo somatic mutation?

Answer 25

1) Maybe as is too dangerous, when in tissues if they somatically mutate would be too high chance that would recognise own tissue (same as antibodies but as they also need T cell help lowers danger)

2) Maybe don’t need somatic mutation, as is only present on cell surface so doesn’t need to bind with really high affinity… but we don’t know

three) As might not recgonise MHC proteins if mutated

Question 26

What is the differences and similarities between B cell and T cell receptors and their genetics?

Answer 26

> Differences:

1) B cell receptors can be soluble while T cell receptors are always membrane bound.

2) Antibodies have an Fc region while T cell receptors do not.

3) Fab region in TCR made up of alpha and beta chain, while in B cell receptors is made up of light and heavy chain.

4) Antibodies undergo somatic hypermutation and class switching while TCRs don’t

5) TCRs need to be accompanied by CD3 proteins to allow signalling to take place.

> Similarities:

1) Both receptors have Fab like regions, containing CDR1-3 where CDR3 has the most variation so binds to antigen.

2) Both TCRs and antibodies under go, somatic recombination and junctional diversity (just in different places of the body).

3) Both have associated proteins with cytoplasmic domains which contain ITAMs which are phosphorylated when antigen binds, difference is that TCR requires CD3 and zeta with ITAM nearby, while B cell receptor require Ig-alpha/beta

Question 27

What is B cell immunity particularly important against and why?

Answer 27

> B cell immunity is particularly important in defence against extracellular pathogens

> Bind to structures on surface of pathogens or viruses stopping entrance to host cells.

> Normally recognise free (non-cell associated) antigen that hasn’t been processed in anyway (native)

Question 28

What does MHC stand for?

Answer 28

Major Histocompatibility proteins (MHC)

Question 29

What is T cell immunity particularly important against and what mechanism do they need to do this?

Answer 29

> Are important against extracellular pathogens but very important for intracellular pathogen defence.

> How can T cells recognise intracellular antigens? Needs a mechanism where samples of what’s inside a cell are displayed on the surface so a complimentary T cell can recognise this (role of MHC)

Question 30

What is the role of Major Histocompatibility proteins (MHC) in immunity?

Answer 30

> Proteins have to be degraded and processed and presented by MHC receptors to the surface to TCRs.

Protein –> Peptide –> MHC –> cell surface –> T cell recognition

Question 31

What do T cell receptors recognise?

Answer 31

T cells do not recognise native pathogens, so mainly recognise processed proteins (T cells recognize “cell-associated”, processed antigen/ has to have been broken down into peptides)

Question 32

How did MHC proteins come about?

Answer 32

Evolved in higher organisms to display parts of what is inside the cell to the surface, so a T cell with the correct receptor will recognise and respond to this.

Question 33

What does Major Histocompatibility mean?

Answer 33

tissue compatibility

Question 34

What chromosome are the genes for the Major Histocompatibility Complex located on?

Answer 34

Chromosome 6

Question 35

What are MHC proteins also referred to as and why?

Answer 35

Also known as HLA molecules in humans (human leukocyte antigen- discovered on these cells). e.g. HLA-A, HLA-B, HLA-C, 3 gene loci coding for 3 different MHC proteins.

Question 36

When were MHC proteins discovered?

Answer 36

MHC proteins discovered during research on graft rejection.

Question 37

What must antigen presenting cells have on their surface?

Answer 37

Antigen presenting cells have to express MHC proteins

Question 38

Why is it hard to find matching MHC proteins in humans?

Answer 38

> Very polymorphic (many forms of MHC proteins in human population)

> e.g. ~5000 alleles of HLA-B locus. Alleles may differ by up to 20 a.a. substitutions. Is hard to find people with exact matches for transplant (matching MHC proteins on the same loci as much as they can)

Question 39

What is the main role of MHC proteins?

Answer 39

Major role in antigen presentation and initiation of T cell responses

Question 40

What is MHC restriction?

Answer 40

MHC restriction refers to the phenomenon where T cells will only recognize an antigen if it is presented on the surface of a cell by self-MHC proteins. In other words, T lymphocytes are ‘restricted’ to recognizing antigens when they are displayed in the context of the individual’s own MHC molecules. This means that the ability of a T cell to recognize an antigen is ‘restricted’ by whether a self-MHC molecule can bind and present that antigen

Question 41

How was MHC restriction showed?

Answer 41

> Experiments with inbred mouse strains and virally infected cells (Very inbred mice share the same MHC proteins)

> Strain A and B immunized with virus, T cells form these mice were isolated and cultured in vitro with cells infected with the same virus

a)T cells from mice strain A and put in cells with virus, the virus was killed

b) T cells from mice strain B, T cells could not kill infected cells from mouse A even though mouse B was immunized against the same virus (MHC restriction)

Question 42

Why does MHC restriction (T cells will only recognise antigen if it’s presented to by a cell which expressed MHC proteins from self) occur and how was this proven?

Answer 42

> As the TCR recognises both antigen (foreign peptide) + MHC

> Proved by X-ray crystallography:
(1) MHC binds peptide
(2) TCR recognises complex of foreign peptide + self-MHC

Question 43

How can a TCR recognise and bind to the complex of an MHC presenting a processed antigen (peptide)?

Answer 43

1) CDR1 and CDR2 of T cell receptor bind self MHC
>germline-encoded, so less variable (as encoded by just V)

2) CDR3 binds peptide
>Variation introduced by somatic recombination and junctional diversity (as encoded by V(D)J)

Question 44

How could somatic hypermutation occurring in T cell receptors cause issues with MHC proteins?

Answer 44

If T cell receptor genes underwent somatic hypermutation, then might lose recognition of self MHC as don’t want CDR1-2 to mutate.

Question 45

What are the 2 classes of MHC protein, what cells are they found on, and what is their purpose?

Answer 45

1) MHC class I
>Expressed by all nucleated cells
>Present peptides derived from endogenous proteins to cytotoxic (CD8) T cells
>Endogenous, is synthesised from the cell that displays it(but hsow infection or cancer as would be expressing foriegn peptides if infectious or cancerous)

2) MHC class II
>Expressed by certain leukocytes (dendritic cells, B cells, macrophages). = antigen presenting cells starting adaptive immune response by presenting to T cells
>More restricted expression pattern
>Present peptides derived from exogenous proteins to helper (CD4) T cells (Things taken up from outside the cell)

Question 46

Name 3 properties of MHCI and MHCII that are similar in structure?

Answer 46

1) Membrane proximal domains are Immunoglobulin-like
>immunoglobulin fold,” a type of protein domain consisting of a sandwich of two beta-sheets. This fold is stabilized by a disulphide bond.

2) Membrane distal domains bind peptide

3) Membrane distal domains contain polymorphisms

Question 47

Describe the structures of 1) MHCI 2) MHCII proteins, go into detail about their subunits..

Answer 47

1) MHCI
>2 chains
a) Alpha3 is intrinsic in membrane
b) Beta2-microglobulin stabilises the molecule in the membrane
>Polymorphisms of alpha1 and 2 are clustered in the peptide binding cleft (furthest away from membrane)

2) MHCII
>2 chains both of which traverse the membrane and are both polymorphic.
>Domains close to membrane are immunoglobulin like, bind to membrane, both alpha2 and beta2 do this.
>Domains distal from membrane are most polymorphic and bind to peptides, both alpha1 and beta1 do this.

Question 48

Wat is the binding grove for MHC proteins made of?

Answer 48

Grove made of beta pleated sheets, alpha helices form sides of domains in both

Question 49

Describe the difference in structure of binding grove for 1) MHCI 2) MHCII

Answer 49

1) MHCI bind peptides 8-10 a.a. long (more restricted in length)
>N and C-termini of peptides bind to invariant sites at ends of the groove.
>Two or three “anchor residues” on the peptides bind to “specificity pockets” formed by polymorphic residues at the base of the groove
>Fairly closed restricting the size of peptides which can bind.

2) MHCII bind peptides ~ 13-18 a.a. long (longer)
>Peptide backbone interacts with conserved residues that line the base of the groove.
>“Anchor residues” on the peptide bind to “specificity pockets” formed by polymorphic residues.

Question 50

What is a funny analogy used to remember how MHC binds to peptides?

Answer 50

> MHC protein is the roll, the peptide is the sausage

> MHCI has smaller sausage (peptide)

> MHCII accommodates bigger sausage (peptide) which can extend past the groove.

Question 51

Can MHC proteins bind to a wide range of peptides, why?

Answer 51

A particular MHC molecule (allele) can bind a wide range of related peptides due to diverse genes

Question 52

Do MHC proteins undergo somatic recombination, if not how do they get their diversity?

Answer 52

The diversity in the MHC proteins are all inherited, no somatic recombination

Question 53

Describe in 5 steps how a virus-infected cell would present endogenous antigen (protein made within the cell to cytotoxic T cell (CD8+ve)

Answer 53

1. Misfolded proteins will be mainly derived from virus
2. In response to interferon stimulation another subunit is made to join the proteosome to form an immunoproteasome
3. Proteins are broken down very efficiently, need to get out of c cytoplasm where are made and into the ER where MHCI proteins are present.
4. Transported into ER by specialised transporter named as TAP transporter (driven by ATP hydrolysis)
5. In ER, peptides bind with MHCI proteins, MHCI proteins is transported to surface and T cell with correct receptor will recognise the self MHC and hopefully the peptide too.

Question 54

What is a proteosome?

Answer 54

Proteosome: multi-subunit complex that breaks down misfolded proteins, when infected from a virus these misfolded proteins will be from the virus

Question 55

What is an immunoproteasome and what induces its formation?

Answer 55

> Immunoproteasome: induced by interferon stimulation

> An extra subunit converts proteosome to immunoproteasome, much more efficient at producing peptides at the correct length for MHCI presentation.

Question 56

What is TAP and how does it work?

Answer 56

Peptides transported to ER by ATP-hydrolysis driven transporter, TAP (transporter associated with antigen presentation).

Question 57

Describe in 4 steps how macrophage/dendritic cell/B cells present exogenous antigen and presents it to T helper cell (CD4+ve)

Answer 57

1. Antigen taken up by phagocytosis or endocytosis
2. In phagolysosome: acidification in vesicles promotes unfolding & proteolysis, breaking protein into peptides.
3. Peptides associate with MHC II in the endocytic compartment.
4. MHCII-peptide transported to cell surface for recognition by helper T cell

Question 58

Where does association with peptides occur for MHCII and MHCI proteins

Answer 58

For MHCII the association with peptides occurs in acidic/ endosomsal/ lysosomal compartment (of phagocyte e.g.)(MHC Class II molecules are synthesized in the endoplasmic reticulum (ER) and transported to the acidic compartments), for MHCI occurs at the ER

Question 59

What is cross-presentation?

Answer 59

Cross-presentation = exogenous peptide (taken up by dendritic cell) associating with MHCI.

Question 60

Why is cross-presentation sometimes needed in an immune response, give an example?

Answer 60

> Sometimes a Dendritic cell needs to induce a naïve cytotoxic T cell to respond to an antigen but dendritic cell is not infected

> E.g. dendritic cell might take up a necrotic cell (dying), proteins made to proteins in proteosome, transported to ER where associates with MHCI which can be presented to naïve cytotoxic T cells to activate them to kill other cells presenting this protein but also does not kill the dendrite

Question 61

What is an overview of the process of cross-presentation?

Answer 61

> Sometimes a Dendritic cell needs to induce a naïve cytotoxic T cell to respond to an antigen but dendritic cell is not infected

> This is done by cross-presentation: Some dendritic cells present exogenous (taken up by dendritic cell) peptide associated with MHCI to cytotoxic T cells(as T helper respond to MHCII and Cytotoxic to MHCI)