Humoral Immunity: B Cell Activation, Affinity... Flashcards
Describe the different antibody functions.
NEUTRALISATION: The antibody works by binding to the docking site or the host entry site of the virus/ bacteria; this prevents viruses from entering cells. It could also bind to the active site of toxins and neutralise the toxins.
OPSONISATION: The antibody will tag the surface of the bacteria, and recruit other cells (such as macrophages) towards it. The macrophage will have Fc receptors on it that is attracted towards the active site. The antibody will induce the macrophage to go through Antibody-Dependent Cellular Phagocytosis (ADCP).
For tumour cells, the antibody will tag it, but since it’s too big to phagocytose, it will instead recruit NK cells to perform ADCC (Antibody-Dependent Cellular Cytotoxicity). NK cells produce chemicals make the tumour cell apoptose.
Antibodies can bind together to form complexes, and agglutinate when they bind to a pathogen, rendering it useless and allowing it to precipitate out of the blood.
COMPLEMENT FIXING: Antibodies can also form complexes with complement molecules, which, in different ways, cause it to phagocytose. In some pathways, the complexes can also form MAC (membrane attack complexes) which punch holes in the membranes, causing the cells to lyse.
Describe the different classes of antibodies.
There are five classes of antibodies, all determines by the five classes of heavy chains. The light chain remains the same in all the classes.
IgG: It has the 4 heavy chain domains, VH-CH1-CH2-CH3. It is stabilised by two disulphide bonds in between the heavy chains.
IgD: It has a longer hinge than IgG, and it stabilised by only one disulphide bond.
IgE: It has 5 heavy chain domains, not 4.
IgA: It is similar to IgG, but the CH3 domain has modification so that it allows interaction with the J (joining) chain. This means that two monomers can join together, and it is then wrapped by secretory component for it to be secreted out to the mucous.
IgM: It is similar to IgD, with 5 domains, but it can form a pentamer with the help of a J chain.
Describe the antigen-dependant part of the B cell lifecycle.
After VDJ recombination of B cells, it will become an immature circulating B cell. However, the lymphoid progenitor cell also gives rise to T cells.
These T cells will migrate to the thymus, where they also undergo their own similar VDJ recombination to form T cell receptors. These T cells receive signals from APCs to become activates, and in turn activate the B cells in the presence of pathogens.
The B cell will then migrate to the germinal centre for a special force of selection.
The germinal centre is split into the light and dark zone.
In the dark zone, affinity maturation (the honing of the variable segment towards a particular antigen) occurs. It involved clonal expansion, where the B cell makes clones of itself, followed by somatic hypermutation, where an enzyme will create mutations in its DNA. It will then select for the B cell receptor that will bind to the pathogen in the light zone. This will go on for several cycles in order to complete affinity maturation.
It will then undergo class switching, as we now know the pathogen so we can switch to the correct heavy chain class. The B cells will then mature into plasma cells and memory cells and circulate in the blood.
Describe two stages of B cell activation.
B cell activation can be split into two stages:
1. T-cell independent B cell activation: this is where the B cell encounters the pathogen first. When the B cell encounters the antigens on the pathogen’s surface, it will undergo clonal expansion. Some of the cells will differentiate into IgM secreting plasma cells, as a first line of defence.
- Some of the cells will travel to the lymph nodes, where they will undergo T-cell dependent
B cell activation. First of all, the B cells need to recognize the antigen, internalize it and present it via MHC Class II (to the T cell?). The second step would be for the B cell to recognize the antigen on a T cell or an APC. To complete the three-step verification process, the T cell produces cytokines that signal to the B cell. This confirms to the B cell that it was not a random activation, so it will start secreting IgG.
The CD40-CD40L ‘handshake’ simply helps the cells in identifying each other.
List the three signals that B cell activation requires.
- Antigen binding to BCRs
- Co-stimulation by activated Th cell specific to same antigen
- Th cell-derived cytokines
Give an overview of what happens in germinal centres.
Naïve B cells are activated by Th cells, and then they will migrate to the germinal centre, undergoing clonal expansion in the dark zone.
An enzyme will cause somatic hypermutations in the variable region, and the cells will migrate to the light zone. With the help of Tfh cells and FDC, it will undergo selection. This will repeat until the affinity is high enough.
Those cells will undergo class switching to select for the correct antibody class, before differentiating to plasma and memory cells.
Describe the basis of affinity maturation.
The receptor that binds the antigen originally is low-affinity.
During the process, an enzyme called AID (activation-induced cytidine deaminase) will cause somatic mutations in the DNA of the variable regions. Another process will select for the high-affinity binding antibody.
Describe, in detail, the process of affinity maturation.
Affinity maturation happens in both the light and dark zone.
The activated B cells will first make clones of itself. Then, AID (activation-induced cytidine deaminase) will induce point mutations in the variable region of the DNA of the B cells. After this, they are all unique B cells (made from identical clones).
After this, they will migrate to the light zone, where the FDC will present the antigen on its molecular surface. The B cells that most closely match will bind to these antigens and present them to Tfh cells. The Tfh cells then send back a survival signal.
From the bunch of cells that underwent somatic hypermutations, since it’s a random process, the cells can end up with a reduced affinity or no change at all. Those cells will apoptose, as they do not receive a survival signal from the Tfh cell.
This cycle occurs several times until the affinity is high enough, and the cells then undergo class switching.
Describe heavy chain class switching.
It only affects heavy chain CONSTANT region.
They have different effector functions – this helps them deal with different pathogens, ranging from viruses to parasites.
There are two types of class switching:
MINOR: Differential splicing (mRNA level) – this is reversible as the DNA is not affected:
- Between IgM and IgD
MAJOR: DNA recombination - irreversible
- IgM to IgG, IgA, IgE
- IgG to IgA, IgE
Describe class switch recombination (CSR).
It requires:
1) a cytokine signal;
2) switch regions (these are similar to RSS, but they are in front of the heavy chain constant region gene segments);
3) AID and DSB (double-stranded break) repair proteins
The recombination occurs between switch regions.
The switching only proceeds downstream (as the upstream gene segments are removed in the process of class switching):
- IgM to IgG, IgA, IgE
- IgG to IgA, IgE
What is the structural difference between membrane-bound and secreted antibodies?
The difference between the secreted form and the membrane-bound form is that the secreted form has a tail piece.
The membrane-bound form has an anchor made up of a hydrophobic transmembrane region and a cytoplasmic tail.
How does the B cell know how to make these forms of antibodies?
It is through differential splicing.
We can see that the Cμ constant region is actually made up of μ1, μ2, μ3, μ4 and a secreted region, followed by a stop codon and a PolyA site. We then have (membrane) M1 and M2, followed by a stop codon and another PolyA site.
For the secreted heavy chains, the DNA region is transcribed into mRNA, and then spliced at the first PolyA site. This results in a sequence that codes for the secreted heavy chain protein.
If you want to include the membrane-bound μ chain, you’d have to splice out the first PolyA site, which ensures that we retain the anchors, so the antibody doesn’t secrete out of the B cell.
