Genetics of Antigen Receptors

Question 1

What is clonal selection of B cells?

Answer 1

Progenitor cells = large amount of naive lymphocytes circulating in the body
Recognition of a threat by one of them = expansion of that specific clone

Question 2

How do B cells recognise foreign cells, and effect a response?

Answer 2

Naive antigen - receptor converted to soluble antibody form, to recruit effector molecules using the Fc

Question 3

How do T cells recognise foreign cells, and effect a response?

Answer 3

Peptide-MHCs, mediated by cell contact or cytokine secretion

Question 4

Each T cell and responsibility?

Answer 4

Th1 - intracellular pathogens e.g. virus for macrophage killing
Th2 - extracellular parasites so recruit eosinophils, mast cells, IgE
Th17 - inflammatory, extracellular bacteria/fungi, recruit neutrophils
Tfh - localise to B cell follicles
Treg - tolerance/immune regulation
Tc - cytotoxic

Question 5

Gene rearrangement?

Answer 5

Generates binding diversity in B cell Igs, where variable and joining gene segments are rearranged to give different transcripts

Question 6

Encoding CDRs in light chain?

Answer 6

CDR 1 and 2 encoded in V kappa

Rest by final joint of the joining segment and variable segment - join between Jk and Vk within CDR3

Question 7

Encoding CDRs in heavy chain?

Answer 7

CDR1 and 2 in V regions

V, D and J all form CDR3

Question 8

Combinatorial diversity?

Answer 8

Needed for broad immune protection - this refers to combining V, J and D segments to give about 2*10^6 combos

Question 9

Junctional diversity?

Answer 9

Responsible for the rest of the diversity needed - J and D joins are imprecise as sequences don’t match exactly

Question 10

How are gene segments joined?

Answer 10

Flanked by recombination signal sequences, RSSs, made up of a heptamer and a nonamer separated by a 12 or 23 base pair spacer

12 always joins to 23,, ensuring that V joins to J in light chains, and V joins to D joins to J in heavy

Nonamers line up complementing each other, heptamers are cleaved and intermediate DNA (spacer) excised

Question 11

Where does imprecision arise in joining?

Answer 11

Resolution of coding ends
Joining of coding ends

In heavy chains: resolution of hairpin coding ends leaves nicks which unfold and form palindromic P-nucleotides, allowing random nucleotides to be added by TdT

Question 12

Non-productive join?

Answer 12

When N-nucleotides are added to the heavy chain in a multiple of 3, leading to a frameshift

Question 13

Sources of diversity in T and B cell receptors?

Answer 13

Variable segments - higher in light chains/alpha
Diversity segments - higher in Ig (BCRs)
T cells have higher junctional diversity from imprecise joining, as TdT is active at all joints
T cells have greater diversity overall

Question 14

Reading frame corrections

Answer 14

1/3 chance that adding or removing nucleotides will cause a frame shift
Correction only possible in TCRs, in beta chains (VDJ joins)
If V-D join goes out of frame, downstream D-J join may produce a complementary insertion/deletion to fix it

Also helps to explain higher junctional diversity in TCRs

Question 15

Why is combinatorial diversity higher in TCRs?

Answer 15

Even though both BCRs and TCRs have similar number of V gene pairs possible

There are different J segments in TCR frame
All included in encoding the CDR3 region
=greater diversity

Question 16

Overall, why is T cell diversity higher?

Answer 16

D segments read in all reading frames
TdT activity at all junctions = imprecise joining
More J gene segments in CDR3 region
Frame shift corrections

Question 17

Define clonality?

Answer 17

Only one antigen specificity expressed

Question 18

Allelic exclusion?

Answer 18

Two copies of the alleles for light and heavy chain are inherited - gene recomb is linked to lymphocyte development so only on functional copy is used (suppression of the other after gene recomb)

Question 19

Isotypic exclusion?

Answer 19

Suppresses expression of the light chain alleles after rearrangement of the functional copy

Question 20

Order of rearrangement in developing B cell?

Answer 20

H chain first, D-J then V-DJ, on first chromosome
L chain rearranged on first chromosome
Rearrangement stops

Question 21

Proteins involved in B cell development and linkage to stage of rearrangement?

Answer 21

RAG 1 /2, expressed during both rearrangements and not during pre-B cell proliferation
TdT - expressed up until small pre-B cell stage, during heavy chain rearrangements
Surrogate light chain components - stop for light chain rearrangement

Question 22

How are RAG genes controlled in B cells?

Answer 22

On during heavy chain recomb
Viable heavy chain binds surrogate light chain - switches off RAG
Proliferation occurs
Then light chain rearrangement

Question 23

Proteins involved in T cell development and linkage to stage of rearrangement?

Answer 23

TdT strongly active throughout
Same RAG pattern
Also uses surrogate alpha chains

Question 24

Alternative RNA processing in BCRs?

Answer 24

Switch from membrane to secreted Ig form when B cells are activated
Uses alternative poly(A) sites
This uses the second poly(A) splice site in the last C exon to exclude the cytoplasmic region, used in secreted IgM

Question 25

C region exons in BCRs?

Answer 25

All B cells express a single C exon (e.g. Cy in IgG), which controls the constant region binding properties and therefore effector function

Question 26

IgM and IgD C region co-expression?

Answer 26

Uses alternative RNA processing
Cmu encodes IgM, Cdelta IgD
If second splice site of IgM is used, IgD exons are added through splicing, usually in mature B cells

Question 27

Isotope/class switching

Answer 27

Irreversible recombination between switch regions in the allele to give a different Ig

Question 28

Diversification in the germinal centre?

Answer 28

Secondary antibody repertoire
Somatic hypermutation and class switching

Question 29

Somatic hypermutation? In B cells

Answer 29

Targets mutations in V regions for varied antigen binding properties
A selection process then narrows down those with greatest affinity to improve response over time