Humoral Immunity: B Cell Activation, Affinity

Question 1

Life cycle of B cells

Answer 1

stem cells undergo heavy and light chain recombination to become immature B cells, and there is also junctional flexibility and P and N nucleotide addition, contribute to B cell antibody diversity

the B cell is now mature and circulates

Mature recirculating B cell encounters a pathogen or TCR signalling, migrates to lymph node germinal centre to undergo affinity maturation and class switching – improves affinity to the specific pathogen. Also it will switch the heavy chain constant region to the correct antibody class to deal with the pathogen.

The B cells will differentiate to plasma cells, secreting antibodies

B cells also differentiate into memory B cells circulate in the body and don’t differentiate into plasma cells until the pathogen is encountered again.

Question 2

antibody functions?

Answer 2

1. virus and toxin neutralisation, prevents pathogen-host binding
2. opsonisation + ADCP, antibody binds to pathogen and “tags” it, and macrophages bind to the antibody via the Fc receptor and phagocytose the microbe
3. Complement fixing/MAC formation (CDC)
   - complement fixing = agglutination as more and more antibody binds to the pathogen
   - MAC punches holes holes in tumour/infected cells, causing them to lyse.
4. Opsonization + ADCC
   - NK-induced apoptosis, occurs with tumour cells because they are too big for macrophages to ingest

Question 3

Different classes of antibodies

Answer 3

type of heavy chain determines antibody class (gamma, epsilon, delta, alpha, miu)

IgG

• 4 heavy chain domains – 1 variable and 3 constant (CH1, CH2 and CH3)
• stabilized by 2 disulfide bonds between the heavy chain
• fixes complement and crosses placenta
• most abundant in the serum
• main Ab of secondary responses: neutralize toxins and opsonisation

IgE

• 5 domains
• mast cells, basophils
• allergy, anti-parasites

IgA

• similar to IgG but the third CH3 domain has a modification allowing interaction with the joining chain so 2 monomers can join together, then its wrapped by a secretory component for it to be secreted out to the mucus.
• secreted into mucous, tears, saliva, colostrum
• breastfeeding

IgM

• forms a pentamer with the help of the j (joining) chain
• main Ab of primary response, forming immune complexes & fixing complement
• monomer serves as BCR

IgD
-BCR; indicates mature B cells, only Ab not secreted

Question 4

how are B cells activated?

Answer 4

The T-cell independent stage - B cell encounters the pathogen, specifically the antigens expressed on the surface. Undergoes clonal expansion, some will differentiate into IgM secreting plasma cells – first line of defence.

Some cells will migrate into the lymph nodes, where they undergo T cell dependent B cell activation. First of all, the B cell needs to recognise the antigen, internalise it and present it via MHC class II. Then it will be recognised by the TCR and also a dendritic cell to verify it. Then the t-helper cells produce cytokines.

Naïve B cells activated by T helper cells and migrate into the germinal centre while undergoing clonal expansion.

germinal centre split into the dark and light zone.

• clonal expansion: B cell makes clones of itself
• somatic hypermutation: an enzyme creates mutations in the DNA
• selection: selecting the BCR that binds to the pathogen the best

These 3 happen in a cycle – affinity maturation

Class switching – switching to the correct class
Differentiation to plasma cells and circulation in the blood

Question 5

explain how clonal expansion and affinity maturation are related

Answer 5

clonal expansion occurs as the B cell that binds the closest to the pathogen will expand

-these B cells will then undergo affinity maturation in the GC

Question 6

Affinity maturation and class switching in Germinal Centre

Answer 6

the AID enzyme will cause point mutations in variable region DNA (somatic hypermutation), makes all the B cells unique to one another - point mutations means the B cells can have reduced affinity, improved affinity, or no change at all, and reduced affinity means they die by apoptosis as they have no help from the FDC or Tfh

then they migrate to the light zone, where FDC present antigens on their surface, and the B cells with a high affinity BCR on their surface grab as much antigen as possible and present to the Tfh (the only T cells that can enter the germinal centre) which give the B cell a survival signal

The cells go through several cycles of affinity maturation before they reach a level where they are able to undergo class switching - correct class is selected for before differentiation into plasma cells

low affinity antibody - high affinity antibody via many rounds

memory cells also formed

Question 7

Heavy chain class switching

Answer 7

Only affects heavy chain CONSTANT region

Different effector functions – deal with different pathogens

Minor: Differential splicing (mRNA level)
IgM and IgD

Major: DNA recombination
IgM to IgG, IgA, IgE
IgG to IgA, IgE

Question 8

signalling for class switching?

Answer 8

CD40L on T cell interacts with CD40 on B cells + cytokine signalling

Question 9

Class switch recombination (CSR) - the steps

Answer 9

1) Cytokine signal; 2) Switch regions; 3) AID and DSB repair proteins

Recombination between switch regions

Switching only proceeds downstream
IgM to IgG, IgA, IgE
IgG to IgA, IgE

e.g. Alternative splicing causes the B cell to make IgM and IgD, then changes occur and B cells switch to making IgA1

Question 10

Membrane and secreted Ig

Answer 10

Secreted form has a tail piece, whereas the membrane bound has an anchor made up of hydrophobic transmembrane region and a cytoplasmic tail

Question 11

how are membrane and secreted Ig made?

Answer 11

differential splicing of mu (μ) constant regions

Membrane M1 M2 region and S secreted region – these regions are spliced out accordingly