T Cell Receptors and MHC Complex Dr. Diebel 5/7/14

Question 1

B cell can secrete their receptors, antibodies, that can go out and find antigen. How do T cells work?

Answer 1

T cells have to be present at the site of interaction w/ antigen they do not secrete their receptors.

*T cells can only see antigenic determinants show to them by special antigen presenting molecules on the surface of their cells.

Question 2

Th1 cells

Answer 2

• Induced by IL-12, “DC 1”, and IFN-alpha
• This T cell is activated and proliferates in the lymph node. Some of the daughter cells leave and circulate around the body and when they encounter an antigen, say at the site of infection, they secrete lymphokines.
• The most important lymphokine secreted by Th1 cells is IFN-gamma (others include IL-2 and (TFN-beta)) which is pro-inflammatory and a chemotactic agent for blood monocytes and tissue macrophages. These monocytes become classically-activated M1 macrophages. IL-2 helps get CTLs activated by antigen.
• The macrophages release their own cytokines that can intensify inflammation such as TNF-alpha and IL-1.
• In summary the T cell recognizes and the macrophage removes. This system has the capability of damaging local tissue and a good example of this is poison ivy which is a good stimulator of Th1 cells. (contact sensitivity)

Question 3

Th17 cells

Answer 3

• Makes the inflammatory lymphokine IL-17 among others (IL-22)
• Similar to Th1 in that it is involved in inflammation and has been implicated in several autoimmune diseases
• Induced by TGF-beta and IL-21 (also IL-6 and IL-23)
• Make IL-17, IL-21, and IL-22 lymphokines
• Appears to play a role in maintaining the integrity of mucosal surfaces

Question 4

Th2 cells

Answer 4

• Like Th1 cells, when activated they leave the lymph node and circulate through that blood until they encounter their antigen again in the tissues.
• Here they make IL-4 and IL-13 (IL-5,6,10) which attracts and activates macrophages into the alternately activated M2 macrophage that are more involved in healing
• IL-4 and IL-5 serve more as a chemotactic for eosinophils which are cells specialized in killing parasites like protozoans and worms.
• Induced by “DC 2” and IL-4

Question 5

Tfh cells

Answer 5

• After the antigen-presenting DC in the lymph node, some activated T cells can be seen migrating into the follicles of the cortex, where B cells are abundant. These are referred to as follicular helpers, and their role is to help B cells that have recognized antigen become activated and differentiate into antibody-secreting plasma cells.
• They secrete a variety of Th1 and Th2 cytokines and direct B cells to switch from secreting IgM to either IgG, IgA, or IgE which is done heterogeneous so that in the gut it is switched preferentially to IgA versus in the spleen to IgG.
• If Tfh cells can’t communicate correctly w/ B cells you may have difficulty making any antibody class, especially those downstream from IgM.

Question 6

Treg

Answer 6

• Small population of cells that is responsible for suppressing the activation and fxn of all other T cells.
• Most have the phenotype CD4+/CD25+
• They produce TFN-beta and IL-10 and are very potent.

Question 7

Cytotoxic Killer T cell induced apoptosis

Answer 7

There are two ways a CTL can signal a cell to undergo apoptosis.

1. It can engage the ‘death receptor’ Fas (CD95) on the target (CTLs bear the Fas lignad, CD95L). Cross linked Fas activates a latent apoptosis pathway.
2. It can secrete the contents of lytic granules which contain proteases called granzymes, and other proteins called performs which seem to allow the penetration of the granzymes into the target cell. These selective proteases trigger apoptosis.
   - CTLs are activated in the lymph node after contact w/ an antigen-bearing DC. They also require help from Th1 in the form of IL-2 for activation, and conversion in memory cells, IL-21.

Question 8

Memory T cells

Answer 8

The small amount of T cells that remain after the infection is gone or immunization has occurred. They have some attributes of stem cells in that they can replace themselves as well as differentiate into effector cells when re-exposed to low antigen concentrations.

Question 9

T cell subpopulation markers

Answer 9

• B cells are distinguished using antibodies to immunoglobulins to the surface markers CD19 or CD20.
• CD3 is on almost all T cells. CD4 helper. CD8 CTL.
• No relaible way to differentiate between Th1/Th2/Treg although Treg usually have more CD25

Question 10

MHC restriction part I

Answer 10

T cells are restricted in their recognition to antigen on the surface of cells genetically identical to themselves. That is they do not see antigen alone, but only antigen presented to them on the surface of a genetically-identical cell. –> The T cell and the APC must come from individuals who have the same alleles at a group of loci called MHC which code for surface glycoprotein molecules. AKA T cell is antigen-specific and MHC-restricted. The chances of two people having the same is very small so the T cells won’t work in each other’s body.

Question 11

Antigen presentation to T cells

Answer 11

When an antigen enters the body it will cause an innate response and its breakdown products will get ingested by a dendritic cell. Within the endosome proteins are broken down to peptides. The endosomes suss w/ other vesicles which have MHC molecules embedded in their membrane facing in. The endosome fuses to the plasma membrane, thus exposing MHC molecules bearing antigenic peptides to the outside world. DC are the best APC.

Because T cells see antigen only when it is complexed w/ cell-surface MHC molecules, T cells focus their attention on cell surfaces, and do not interact w/ free antigen that is a job for B call and its antibodies.

Question 12

MHC restriction part II

Answer 12

There are two types of MCH I/II. Class I products are on all nucleated cells. Class II products are expressed on the surfaces of DC and macrophage-type cells, B cells, and a few other types of cells which are someway involved in presenting antigenic peptides to Th cells.

When antigen is endocytosed and presented by a DC is associates primarily w/ class II MHC molecules. Th1, 17, fh, reg, and 2 are programmed to recognize peptides of MHC II

Class I molecules best associate w/ peptides that are sampled from proteins synthesized within the cell itself, not taken up by endocytosis. Most peptides would be of normal ‘self’ but antigens could be derived from abnormal molecules from pathogens such as viruses. CTLs are programmed to see antigen in association w/ MHC I molecules.

The dendritic cell is unique in that it allows some peptides from antigens its eaten to leak over to its intrinsic pathway so that it can present them on Class I and II at the same time and called cross presentation.

Question 13

APC cell expression

1. DC
2. Macrophages
3. B cells

Answer 13

1. Constitutively express high levels of MHC II and constitutively express B7 (AKA CD80/86) and other costimulatory molecules
2. Must be activated by phagocytosis to express MHC II and must be activated to express costimulatory molecules
3. Constitutively express MHC II and must be activated by antigen binding to antibody before they express costiumulatory molecules.

Question 14

TCR

Answer 14

Two chains called alpha and beta or gamma and delta and each has a constant and variable portion. T cell makes its receptor out of V, (D), and J regions. The process takes place in the thymus. Both alpha and beta chains have transmembrane domains, unlike surface IgG, in which only the heavy chains are transmembrane. It is a heterodimer unlike antibody which is a homodimer.

Associated w/ the TCR is CD3 which serves to transduce TCR signals for the T cell. What this means is that when a T cell binds the correct antigen + MHC w/ its TCR, the actual signal that turns the T cell on is transmitted by CD3.

When a Th0 binds to a DC, the DC will give the T cell and activating boost by secreting cytokines so it gets two hits one from CD3/TCR complex and one from DC cytokines.

Question 15

CD4 signaling pathway

Answer 15

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Question 16

Affinity of TCR for MHC/peptide

Answer 16

Affinity of the TCR for the MHC/peptide complex is relatively modest. CD4/8 adds to the binding affinity.

Other co-receptors add to binding affinity:

• CD2 binds to LFA-3
• LFA-1 (leukocyte functional antigen (integrin)) binds to ICAM-1 (intercellular adhesion molecule)
• CD28 binds to B7
• CD45R binds to CD22

Question 17

Immunological synapse

Answer 17

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Question 18

TCR gene rearrangement

Answer 18

The α chain, like the Ig light chain, is encoded by V, J, and C gene segments.
Rearrangement leads to a selection of a VJ combination attached to a single C segment.
The α chain has a transmembrane cytoplasmic tail making the TCR α receptor a membrane-bound chain.
The β chain, like the Ig heavy chain, is encoded by V, D, J, and C gene segments.
Rearrangement leads to a selection of a VDJ combination attached to one of two C gene segments.
The β chain also has a transmembrane cytoplasmic tail making the TCR β receptor a membrane-bound chain.

The pre-T cell expresses the RAG1 and RAG2 recombinase genes.
Like in B cells, these enzymes recognize conserved recombination signal sequences flanking the regions of the V, D, and J coding sequences in the DNA.
Like in B cells, the RAG1/2 complex joins together the various DNA segments and “throws away” the intervening DNA information.
Each rearranged DNA sequence will encode a single type of TCR.

Combinatorial joining of the V, D, and J region generates TCRs with a variety of antigen-binding specificities.
Increased diversity is gained by alternative joining of the of the D sequences to give VJ, VDJ, or VDDJ chains.
Somatic mutation is also possible, addition of 0-6 nucleotides can be added to the junctions between the V and D and between the D and J domains.

Somatic hypermutations do not play a role in generating TCR diversity.
Remember that somatic hypermutation is possible in B cells (AID mediated). This leads to enhanced antigen binding by the Ig genes. B cells are continually selected throughout this process.
T cells are only selected in thymus, once they leave the thymus there are no more control mechanisms to guide their TCR binding specificity.
It would be a bad idea to allow somatic hypermutation in T cells  autoimmunity.

Question 19

TCR allelic exclusion

Answer 19

Each T cell will produce a single β chain from just one of the chromosomal loci (allelic exclusion).
Two α chains can be produced by a single cell.
Therefore two αβ TCR sets may be expressed on a given T cell.
However, a single T cell will express a single antigen-binding specificity. How?
There are many MHC molecules that could possibly be expressed and recognized by TCRs.
Most TCRs will fail to recognize self and therefore will not survive the passage through the thymus.
Only those T cells with a TCR that recognizes self will survive – self-restricted T cells.
Only those T cells that also pass negative selection will survive – self-tolerant T cells.
This makes it extremely improbable that a single T cell will express two different types of αβ TCRs that will pass both tests and respond to two different antigens.

Question 20

Positive and negativ selection

Answer 20

Only about 2% of the cells survive this process.

Question 21

A T cell must

Answer 21

A T cell must:
Not recognize “self”. Cannot bind so firmly to a self structure (MHC alone, or MHC loaded with a ‘self’ peptide) that the T cell becomes activated; this would be autoimmunity.
Not recognize free antigen (which is antibody’s job).
Recognize antigenic peptide plus self MHC.

The repertoire is selected within the thymus.

Question 22

What are MHC molecules?

Answer 22

MHC = Major Histocompatibility antigens.
Also known as the HLA (human leukocyte antigen) complex in humans.
Originally recognized for their involvement in rejection of tissues exchanged between two unrelated organisms.
MHC molecule play an essential role in antigen recognition and presentation to the immune system.

The MHC complex contains more than 100 separate genes that are subdivided into Class I, Class II, and Class III MHC molecules.
Class I MHC molecules present antigen to CD8+ T cytotoxic T cells.
Class II MHC molecules present antigen to CD4+ helper T cells.
Other MHC proteins have been grouped together as the Class III MHC molecules.
Diverse group of proteins
Complement proteins
TNFα and TNFβ
Various enzymes, heat shock proteins, antigen processing molecules, etc

Question 23

MHC subclasses

Answer 23

There are three “classical” Class I gene loci
MHC Class I gene A encoding HLA-A
MHC Class I gene B encoding HLA-B
MHC Class I gene C encoding HLA-C

Each of the Class I gene products associates with another peptide known as the β2 macroglobulin, when expressed on the cell surface.

Each Class II MHC gene locus encodes two peptides, an α subunit and a β subunit that are expressed together on the cell surface.
There are three major Class II gene loci.
MHC Class II gene DP encoding DP αβ
MHC Class II gene DQ encoding DQ αβ
MHC Class II gene DR encoding DR αβ

Question 24

Each locus of MHC is polymorhphic

Answer 24

There are many alleles for each classical Class I and Class II locus.
This polymorphism gives us our unique identity, permits the recognition of self versus non-self, is an impediment to organ transplantation.
Affect the ability to make an immune response.
Affects the resistance or susceptibility to infectious diseases.
Affects the susceptibility to autoimmune diseases and allergies.

MHC Class I polymorphisms:
1448 A alleles
1988 B alleles
1119 C alleles
Most of the polymorphisms are in the cleft region (antigenic binding site).

MHC Class II polymorphisms:
DR locus
2 DR α alleles
860 DR β1 alleles
~71 DR β2-9 alleles
DQ locus
47 DQ α1 alleles
126 DQ β1 alleles
DP locus
17 DP α1 alleles
134 DP β1 alleles

Question 25

Medical significance of the polymorphisms

Answer 25

HLA-B27 individuals are 90x more likely to develop ankylosing spondylitis (destruction of the vertebral cartilage).
Also linked to psoriasis, inflammatory bowel disease, and Reiter’s syndrome.

HLA-DR2 individuals are 130x more likely to develop narcolepsy.
Also linked to multiple sclerosis, hay fever, and SLE.

HLA-A3/B14 individuals are 90x more likely to develop hemochromatosis (too much iron adsorption which can lead to internal organ damage).

HLA-DQ2/GQ8 is linked to Celiac disease.

HLA-DR3 is linked to diabetes mellitus type I, Grave’s disease

HLA-DR4 is linked to rheumatoid arthritis and diabetes mellitus type 1

Question 26

Lectins and Mitogens

Answer 26

Lectins are proteins made by many life forms including us, plants, and invertebrates. They have affinity for certain sugars – usually ones that the organism they come from doesn’t itself have. They may function as innate ‘immune’ molecules, gumming up the surfaces of foreign invaders.

Some of them bind to and stimulate T and B cells, and they are thus very useful in the research and clinical labs. For example, phytohemagglutinin, PHA, a bean lectin, stimulates all T cells to divide because it binds to CD3. Because it stimulates T cell mitosis it is called a mitogen. So does another bean lectin, concanavalin A, Con A. Pokeweed mitogen (PWM) stimulates both T and B cells (again, nonspecifically) to divide. And antibody to CD3 can be a T cell mitogen, too.

The usefulness of these agents is considerable. For example, to see a person’s karyotype
(metaphase chromosome picture) you need dividing cells. Take some blood leukocytes, add
PHA, and in three days or so you have all the lovely mitotic figures you could want, derived
from activated T cells. Or if you want to see if someone’s T cells can divide normally, or make IL-6, but you don’t know what antigens she’s immune to, just add some Con A to their blood cells; it will stimulate all helpers and CTL, without regard to their antigen-specificity.