T Cell development and generation and of repertoire diversity

Question 1

What are the stages of haematopoesis?

Answer 1

Commitment
Proliferation
Selection
Differentiation in distinct subpopulations

Question 2

What progenitor do lymphocytes (B and T cells) come from?

Answer 2

Common lymphoid progenitor which commits into B or T cell lineage (forming pro B or pro T cell)

Question 3

What happens in the proliferation stage of haematopoesis?

Answer 3

Pro B or pro T cell proliferates intensely after receptor rearrangement before selection

Question 4

What happens in the selection stage of haematopoesis?

Answer 4

Types of receptors on specific cells selected

Question 5

What drives the commitment of precursor in T or B cells?

Answer 5

c-KIT

IL7 and IL3 cytokines later needed to T cell lineage

Question 6

What are the 3 cells that a common lymphoid progenitor commited to T cell lineage forms?

Answer 6

alpha beta or gamma delta t cell from pro t cell

some t cells progenitor may branch out earlier before becoming a pro T cell instead into iINNATE LYMPHOID CELLS which accumulate in cells

Question 7

What factors control the development from a T cell precursor into a pro-T cell?

Answer 7

Notch 1 and GATA 3

Question 8

Describe the process of T cell maturation/development?

Answer 8

Stem cell reprecursor move early from bone marrow to thymus and becomes pro-lymphocyte

Pro-lymphocyte becomes committed T cell lineage due to notch signals from thymic stroma into pre-lymphocyte in T lineage.

Notch signals activates transcription factor GATA 3 for commitment of T cell and rearrangement of T cell receptor.

Once receptor has developed, T cell proliferate intensely.

Thymic selection occurs and T cell can leave thymus untl activated by antigen where it can carry out effector function

Question 9

Where do pro-lymphocytes becomes committed to T cell lineage?

Answer 9

In the thymus

Question 10

What are the 2 regions of the thymus?

Answer 10

Cortex (darker)

Medulla (lighter)

Question 11

What cells are in the thymus?

Answer 11

network of epithelial cells and lymphocytes

Question 12

Why does the thymus not increase in size during intense proliferation of T cells?

Answer 12

Many T cells fails negative selection due to rearrangement of T cell receptor

Question 13

How do you know which stage a T cell is in in its development?

Answer 13

Changes in surface receptors indicate level of development

Question 14

Describe the change in surface receptors of T cell during its development/maturation process?

Answer 14

T cell progenitors (committed to T cell lineage in thymus) express CD2 and Thy1 so far DON’T EXPRESS NORMAL MARKERS OF T CELLS FOUND IN PERIPHERY (SUCH AS CD3, CD8, CD4) so currently double negative T cells

Rearrangement of T cells receptor on thymocyte as it becomes a pro T cell to express both CD4 and CD8 (double positive)

Later stages and selection causes single positive T cells (loses one of receptors)

Question 15

What is a t cell receptor?

Answer 15

Heterodimer - 2 transmembrane polypeptide chains covalently linked by disulfide bonds (either an alpha and beta or a gamma and delta)

Question 16

What are the regions on a TCR?

Answer 16

N terminal variable domain (V)
Constant domain (C)
Hydrophobic transmembrane region
Short signalling cytoplasmic region

Question 17

What varies in the V region of TCRs?

Answer 17

Short amino acid chains that are variable between receptors

Question 18

What does the variable region of TCRs form?

Answer 18

Complementary determining regions (CDRs) forming the MHC binding site (aka antigen binding site)

Question 19

Compare similarities and differences structurally between immunoglobulins and TCRs?

Answer 19

TCR made of alpha and beta chain
Ig made of heavy and light chain

Both have 3 CDRs (one for each chain)

TCR has one V and one C domain on each chain
Ig has one V domain and 3-4 C domains on heavy chain and one V domain and one C domain on light chain

Somatic mutation, secreted form and isotope switching only in Ig’s

Question 20

What components make up a TCR signalling complex?

Answer 20

TCR and CD3 and zeta chain

Question 21

How is the TCR signalling complex formed?

Answer 21

C regions of TCR have cysteine residues which are charged amino acids. They bring the chains together and also bind to CD3 and zeta chain (receptor interacts with accessory molecules) forming signalling complex.

Once MHC peptide binds to TCR, transduction of signal takes place (downstream signals)

Question 22

Which type of peptides do T cell recognise?

Answer 22

short LINEAR peptides (not full confirmational peptides) , so must be cleaves and processed and presented on MHC

Question 23

What peptides do CD4 T cell recongise? and why

Answer 23

Peptides from extracellular spaces since these are the peptides expressed on MHC class 2 which CD4 recongises

Question 24

What peptides do CD8 T cell recognise?

Answer 24

Peptides from intracellular spaces since these are he peptides presented on MHC class 1 which CD8 recognises

Question 25

What is the molecular principle of MHC restriction?

Answer 25

MHC also interacts with TCR independent of peptide showing TCR is functional (which is why during selection they are made sure to bind to self MHC)

Question 26

What is MHC?

Answer 26

Major histocompatibility complex

Question 27

What is the MHC found in humans?

Answer 27

HLA (human leukocyte antigen)

Question 28

What peptides do MHC class 1 present?

Answer 28

Peptide antigen derived from pathogens that replicate inside the cell e.g viruses

Question 29

What peptides do MHC class 2 present?

Answer 29

Peptides from pathogens and antigens that are present outside the cell taken up by endocytic vesicles of phagocytic cells

Question 30

What binding site is present on the conserved region of MHC class 2?

Answer 30

CD4 binding site

Question 31

What binding site is present on the conserved region of MHC class 1?

Answer 31

CD8 binding site

Question 32

Which part of the TCR does CD4 or CD8 depending on MHC class bind to?

Answer 32

Conserved domain

Question 33

Why are there many different MHCs?

Answer 33

MHC molecules are polymorphic and polygenic (many genes and many variants of each gene)

Question 34

Define polymorphic

Answer 34

Multiple variants of each gene

Question 35

Define polygenic

Answer 35

Several different MHC class 1 and class 2 genes

Question 36

How many peptides can bind to MHC?

Answer 36

Only one at a time but can bind diff peptides (due to high level of diversity)

Question 37

When do MHC molecules acquire the peptide they present?

Answer 37

Whilst being assembled inside the cell

Question 38

How do peptides bind to MHC molecules?

Answer 38

Peptides have similar structural features that promote binding

Question 39

What other molecule apart from pathogenic peptide does MHC present?

Answer 39

Self peptides

Question 40

Which cells express MHC class 1?

Answer 40

All body cells apart from erthyrocytes

Question 41

What is the pathway of antigen processing and presentation on top of MHC class 2?

Answer 41

Phagocytosis (dendritic cells and macrophages) Internalised protein is processed in endosomal/lysosomal vesicles Synthesis and transport of MHC class 2 molecules via golgi to vesicles to endosomes MHC class 2 release CLIP molecules and exchanges for peptide molecule - association of MHC class 2 and processed peptide (peptide has affinity to binding groove) Expression of peptide MHC complex on cell surface

Question 42

What are the simplified steps of TCR rearrangment?

Answer 42

Germ line alpha chain DNA (contains multiple V, J etc... variants) Rearranged alpha chain DNA made (variants selected) + Germ line beta chain DNA Rearranged beta chain DNA made Rearrnaged beta chain and alpha chain form alpha beta heterodimer (TCR)

Question 43

How are T cells produced with many different TCRs (diverse reportire) with specificities?

Answer 43

Variable domains made from combination of a group of sequences in the germ line Choose which D and J fragments then binds to form rearranged product

Question 44

How many TCRs does each T cell have?

Answer 44

Only one TCR so specific to one antigen

Question 45

Are TCRs secreted?

Answer 45

No, immunoglobulins are

Question 46

When does a T cell become an effector T cell?

Answer 46

When it finds a complimenatry foreign antigen

Question 47

Why is it unlikely that a T cell will bind to a self antigen

Answer 47

Due to randomness of process of selection in rearrangment of germ line genes (VDJ reassortment)

Question 48

Which enzyme rearranges gene segments (VDJ)?

Answer 48

Rag 1 and Rag 2

Question 49

Apart from alpha beta receptors, what other TCRs are made from gene reassortment?

Answer 49

gamma delta

Question 50

What order does gene reassortment go in?

Answer 50

V fist, then J and last C(constant) to join it all together

Question 51

Does alpha or beta chain have a D segment?

Answer 51

Beta chain

Question 52

Does the biosynthesis of TCRs mediated by Rag 1 and Rag 2 (for gene reassortment) require antigens?

Answer 52

No this is antigen independent

Question 53

Does TCRalpha or TCRbeta o through gene reassortment first? and what happens to the other

Answer 53

TCR beta TCR alpha rearrangement is halted

Question 54

How much gene reassortment has to happen to produce a TCR?

Answer 54

Repeated multiple times until productive rearrangment made

Question 55

What is junctional diversity?

Answer 55

Nucleotides made be added or removed during gene reassortment creating new sequences at junctions

Question 56

What mediates junctional diversity?

Answer 56

TdT terminal deoxynucleotidyl transferase

Question 57

Why is junctional diversity important?

Answer 57

another source of diversity in TCRs

Question 58

What happens to the beta chain until alpha chain is also made?

Answer 58

Beta chain is exported with temporary alpha chain(pre-TCR) to cell surface to signal completion of beta chain rearrangement after which alpha chain rearrangment finishes

Question 59

What is the point of pe-TCR?

Answer 59

Beta selection Pre-TCR signals suppression of RAG genes temporarily - no more rearrangement by allelic exclusion ensuring only one TCR beta is expressed

Question 60

What happens after succesfull pre-TCR signalling step by step?

Answer 60

Further beta chain rearrangement stopped Induce expression of CD4 and CD8 Initiate alpha chain rearrangement

Question 61

What are the 2 ways diversity is given to TCRs?

Answer 61

Gene reassortment and junctional diversity