T cell development - Generation of receptor repertoire diversity

Question 1

What events occur in lymphocyte development?

Answer 1

• Commitment - Proliferation - Selection - Differentiation into distinct functional effector subpopulations

Question 2

What can multipotent haematopoietic stem cells give rise to?

Answer 2

Multipotent HSCs give rise to distinct B and T cell lineages, which then commits to a common lymphoid progenitor which can go down to commit to B cell lineage

Question 3

What 2 kinds of mature T cells can you have?

Answer 3

depending on the type of T cell receptor they express: alpha beta t cell receptor gamma delta t cell receptor

Question 4

List the stages of T cell maturation

Answer 4

stem cell pro-lymphocyte pre-lymphocyte immature lymphocyte mature lymphocyte

Question 5

What happens in the journey of T cells through development?

Answer 5

T cell progenitors develop in the bone marrow and migrate to the thymus. Then there are specific signals, such as notch signals provided by the thymic stroma and the progenitors commit to the T cell lineage Notch signals induce the activation of the transcription factor called GATA3 which is essential for lineage commitment. As a result, T cell precursors undergo intense proliferation. There is positive and negative selection in the thymus. Mature T cells migrate to the peripheral lymphoid organs. Activated T cells migrate to sites of infection.

Question 6

What are successive stages in T cell development marked by?

Answer 6

Successive stages in T cell development are marked by changes in surface receptors 1 week after arrival of precursors into the thymus progenitors commit to the T cell lineage Express early markers of the T cell lineage (CD2 in humans eg and Thy1 in mice) Do not express any of the markers that define T cells later in development or after in the periphery (CD3, CD4, CD8) Because of the absence of CD4 and CD8 early developing T cells are called DN (double negatives) At DN stage developing T cells (thymocytes) re-arrange the TCR locus Stages post DN are characterized by the expression of both CD4 and CD8 and later just one or the other

Question 7

What is the T cell receptor?

Answer 7

Upon successful rearrangement and in the periphery (if selected) T cells express high levels of TCR TCR is a heterodimer consisting of two transmembrane polypeptide chains covalently linked to each other by disulphide bonds Two types of T cell receptors: alpha-beta and gamma-delta Each chain has one Ig-like N terminal variable domain (V) and one Ig-like constant domain (C), a hydrophobic transmembrane region and a short signaling cytoplasmic region The V regions of both chains contain short stretches of amino acid sequence that is highly variable between receptors. These regions form the CDRs or complementary determining regions. The 3 CDRs of the alpha chain and 3 of the beta chain form the peptide-MHC binding site

Question 8

The TCR is similar to the immunoglobulin but also has differences- what are some of these differences/similarities?

Answer 8

T cell receptor: alpha and beta chains Ig: has heavy and light chains TCR: has one V domain and one C domain in each chain Ig: Heavy chain- one V domain, 3 or 4 C domains Light chain: one V domain and one C domain TCR: 6 CDRs involved in antigen binding Ig: ALSO 6 CDRs involved in antigen binding

Question 9

What else assists the T cell receptor?

Answer 9

The C regions have cysteines residues that bring the chains together Charged residues in the transmembrane region bind to CD3 and the zeta chain to form the TCR signalling complex CD3 and zeta allow for the transduction of signals upon MHC-peptide binding

Question 10

What are some features of antigens recognised by T cells?

Answer 10

Question 11

Important point to note about the interaction of MHC with TCR?

Answer 11

The MHC also interacts with the T cell receptor, independent of the peptide.

There are polymorphic residues of MHC that recognise the TCR.

This process determines whether a T cell is functional or not

Question 13

What is the MHC?

Answer 13

• MHC is the Major Histocompatibility Complex
• HLA Human Leukocyte Antigen
• MHC class I and MHC class II
• MHC class I molecules present peptide antigens derived from pathogens that replicate inside the cell, such as viruses.
• MHC class II molecules present peptides from pathogens and antigens that are present outside the cell taken up by endocytic vesicles of phagocytic cells.

Structure:

• Extracellular peptide binding cleft
• Ig-like domain
• Cytoplasmic tail

•

•MHC class II has a conserved

CD4 binding site

•MHC class I has a conserved

CD8 binding site

MHC class II molecule has 2 chains

Question 14

Why are MHC molecules polymorphic and polygenic?

Answer 14

The MHC is highly polymorphic. There are multiple variants of each gene within the population.

The MHC is polygenic, it contains several different MHC class I and class II genes. Thus every individual possesses a set of MHC molecules with different ranges of peptide binding specificities

Thus polymorphism and polygeny lead to a high degree of polymorphism

Question 15

What do we know about the polymorphic residue location and MHC-peptide interactions?

Answer 15

• Each MHC has one cleft that binds one peptide at the time but can bind different peptides
• Peptides that bind one MHC share structural features that promote binding
• Acquire peptides while assembled inside the cell
• Peptide-MHC interactions are storable with low off rate
• Very small number of MHC-peptide complexes can activate a T cell
• MHC molecules can bind and display foreign and self peptide
• MHC class II bids to longer peptides than class I

Question 16

What cells are MHC classes expressed by?

Answer 16

MHC Class I is expressed by all cells, except erythrocytes.

MHC Class II is restricted to antigen presenting cells.

Question 17

What is the pathway of antigen processing and presentation on top of MHC class II?

Answer 17

Question 18

What are some properties of the T cell receptor?

Answer 18

• Only one form of TCR is expressed on each T cell.
• This means that each T cell and its daughter cells have only one TCR and one specificity for antigen
• This is a T cell clone
• However, there are an infinite number of different versions of the TCR each with a unique antigen binding site.
• A TCR has only one antigen binding site.
• A TCR is never secreted.

Question 19

What do Rag 1 and Rag 2 genes do?

Answer 19

RAG 1 and 2 genes mediate the recombination events leading to rearrangement

• The T-cell receptor gene segments are arranged in a similar pattern to immunoglobulin gene segments and are rearranged by the same enzymes; Rag 1 and Rag 2
• T-cell receptors concentrate diversity in the third hypervariable region CDR3.
• g:d T-cell receptors are also generated by gene rearrangement.

Biosythesis of the TCR is antigen independent

Question 20

What characteristics do TCR alpha genes have?

Answer 20

—They do not have D gene segments

—They are rearranged only after the TCRβ chain gene locus has been rearranged.

—Successive rearrangements may be attempted until a productive rearrangement has been achieved

Question 21

How does junctional diversity come about?

Answer 21

During the joining of different gene segments, addition (or removal) of nucleotides may create new sequences at junctions.

Mediated by TdT terminal deoxynucleotidyl transferase .

for extra diversity

Question 22

What do we know about allelic exclusion?

Answer 22

As a result of signalling through the pre-TCR - allelic exclusion

—Signalling through the pre-TCR suppresses expression of the RAG genes.

—So, no more rearrangement at this stage, this is allelic exclusion.

—Allelic exclusion ensures that only one TCRβ chain gene is expressed.

—These events together are known as β-selection

—

Question 23

What does successful signalling of a PreTCR lead to?

Answer 23

Successful signaling of a PreTCR:

• Halts further b chain rearrangements
• Induces expression of CD4 and CD8
• Initiates alpha chain rearrangement

Question 24

Summary

Answer 24

• T cells come from HSC via common lymphoid progenitors and originate in the bone marrow
• Migrate and colonise the thymus where they commit to the T cell lineage and develop into naïve T cells
• Development of T cells occurs in close contact with thymic stromal cells (cortical epithelial cells at DN stages)
• The diverse TCR interacts with the polymorphic MHC molecule in a close relationship involving self and later foreign peptides
• Initial developmental stages focus on the generation of an antigen T cell receptor that is produced by gene rearrangement and junctional diversity
• The rearrangement of the TCR is a step-wise process controlled by checkpoints (2)
• Successful rearrangement of the b chain and signals through the pre-TCR lead onto allelic exclusion of the beta chain
• Alpha chain rearrangements coincide with the transition of T cells to the CD4 CD8 DP stage prior to the events of selection