Humoral immunity - antibodies and the life cycle of B cells Flashcards
Describe the antibody/immunoglobulin
2 heavy chains
- Have 4 domains (can be divided into 5 classes and then sub-classes as above, 9 classes all together)
- Vh = variable heavy
- Ch1, Ch2, Ch3 = constant heavy
2 light chains
- Have 2 domains
- VL = variable light
- CL = constant light
Variable region: 1 light and 1 heavy chain - target system of the antibodies (different for each, high specificity)
Constant region - rest of antibody - same for all antibodies of same class
What are the 2 forms of antibodies?
- Membrane bound = B cell receptor
- Secreted = final fully functional form of the antibody secreted by mature plasma cells
* Prior to this, it is anchored on the membrane B cells for weapon development
The difference between them is the secreted form has a tailpiece, and the membrane bound form has a transmembrane region and a cytoplasmic tail as a anchor
Where does the antigen binding and biological activity happen on the antibody?
Variable light and variable heavy = antigen binding
Constant region plays a part in the biological activity of antibody
Describe the different chains in the antibodies
All the chains are amino acids - start with NH3+ group and end with COO- group
Heavy and light chains are held together by disulphide bonds between cysteine and amino acid residues in the chains
There are also intramolecular disulphide bonds to stabilise each of the domain
What is the hinge region and CHO region on the antibody?
There is a hinge region between CH1 and 2 domains to provide flexibility
CHO region is to promote interaction between antibodies and other immune cells
What are the complementarirtiy determining regions?
CDR = where the antibody interacts with antigens
How does virus and toxin neutralization combat pathogens by a antibody?
- prevents the pathogen from entering the cell.
The variable fragment can bind to active sites of toxins produced by the pathogens and neutralises them
How does opsonization and ADCP combat pathogens by a antibody?
= Tagging of the pathogen so that it becomes more visible to other immune cells like macrophages and NK cells.
ADCP is performed by macrophages to engulf smaller pathogens
How does opsonization and ADCC combat pathogens by a antibody?
performed by NK cells instead which releases chemicals to induce and apoptosis - generally for infected or cancerous cells
How does complement fixing/MAC formation combat pathogens by a antibody?
antibodies are able to form new complexes of pathogen and antibody and fixes complement - leading to series of events which promotes inflammation, fibro cytosis and formation of membrane attack complexes (MAC) which causes cell lysis
What are the different classes of antibodies?
Compare and contrast different types of antibodies
What is heavy chain class switching?
Occurs rapidly after activation of mature naïve B cells, resulting in a switch from expressing IgM and IgD to expression of IgG, IgE, or IgA
- Only affects heavy chain constant region
- Enables different effector functions to deal with different pathogens
What is the difference between major and minor heavy chain class switching?
Minor: mRNA level - between IgM and IgD
Major: DNA recombination between switch regions
- IgM to IgG, IgA, IgE
- IgG to IgA, IgE
- Major requires class switch recombination (CSR) involving:
- cytokine signal
- swtich regions
- AID and double-stranded break repair proteins
How does the B cell know which class to switch to?
By sensing chemicals around them produced by T helper cells
- Eg. An increase in IL-4 will lead to class switch to IgG1, IgG4 and IgE
What is somatic recombination?
= alteration of genetic information on the DNA level.
Processes include:
- V(D)J recombination
- Tdt nucleotide addition
- Somatic hypermutation
- Class switching
What is alternative splicing?
= changes made on the mRNA level.
- 2 identical copies of mRNA in the B cell will be altered differently to produce 2 different protein products like IgM and IgD
- Membrane bound and secreted immunoglobins
How does a stem cell become a mature B cell?
B cell starts in bone marrow, from stem cells to Pro-B cell.
- The DNA of the pro-B cell will undergo D–>J and V–>DJ recombination to permanently code in the heavy chain region and will be expressed with a ‘u’ constant region–> Pre-B cell (cannot differentiate further due to not knowing what the pathogen is at this time)
- The Pre-B cell will then undergo another V-J recombination to permanently code in the light chain variable and constant region to become immature B cells. These cells express IgM and mature over time
- They can add additional diversity through a junction of flexibility and P and N nucleotide addition
- Once they express IgM and IgD on their surface through alternative splicing of their mRNA they will become mature B cells (AKA resting and naive) and circulate between the bloodstream, spleen and lymph nodes.
When does a cell become a pre B-cell?
- when it can express a full heavy chain with a unique variable region
It will also express a light chain placeholder that forms a pseudo antibody with the heavychain
What happens when a B cell becomes activated when it encounters a pathogen?
The B cell will further fold its ability to bind to the pathogen through affinity maturation in the GC
They will then undergo class switching to the appropriate and effective regions to treat that specific pathogen - either become IgG or IgA
The majority of these cells will further develop into professional plasma cells that secrete the antibody that they code for
What happens to B cells after an infection?
Some of the B cells will remain memory B cells
How can Ig get variable fragment diveristy?
VDJ and VJ recombination
- Gene segments are arranged in different combinations to generate many Ig sequences
What does VDJ and VJ recombination mechanisms need?
Recombination signal sequences (RSS) = conserved sequences upstream or downsream of gene segments
- made up of turns consititng of heptamer and nonamer with a 12 or 23bp spacer
What is the one-turn/two-turn rule?
= recombination occurs only between a segment with a 12 bp spacer and a 23 bp spacer
- prevents the D fragments or the V and J fragments from accidentally recombining
What is combinatorial diversity?
= how many antibodies can we get from using all the VDJ segments of heavy chain and VJ segments of light chain
- not all antibody diveristy comes from this
What is junctional flexibility?
generates several productive combinations that encode alternate amino acids at the coding joint
Adding P nucleotides and N nucleotides will additionally add diversity
What is allelic exclusion?
Two copies of each Ig gene - one from mother and one from father
Antibody genes are different - only one heavy chain allele and one light chain allele is expressed
Order of rearrangement: heavy>kappa>lambda, 1st allele than 2nd
- If not successful cell will undergo apoptosis
What is the antigen-dependent life cycle of B cells?
- B cells are activated from the helper T cells.
- These activated B cells will then migrate to the Germinal centre where it will undergo affinity maturation (to improve their affinity for attacking antigens)
- This process involves clonal expansion and somatic hyper mutation which occurs in the dark zone. Then the cells migrate to the light zone to undergo selection. This process is repeated several times
- They will then undergo class switching once they know what antigen they are fighting
- The B cells will then differentiate to plasma cells - secreting antibodies while still maintaining some B-cell receptors on their surface
What is the T-cell indepdendent part of B-cell activation?
When the pathogen invades, the B cells are partially activated to process the antigen
The B cells will then make clones of itself through clonal expansion
- Some will secrete IgM
- The others will migrate to the lymph node to wait for T-cell activation
What is the T-cell dependent part of B-cell activation?
T-cell dependent B-cell activation requires a triple-verification process to ensure the B cells don’t get activated by mistake
- First, the B cell will encounter the pathogen and internalise the antigens
- The antigen is then presented on the surface of the B cell via it’s MHC Class 2 receptor
- Then it has to be activated by a T cell (which has already been activated by the same pathogen)
So the pathogen will be engulfed by the dendritic cells, presented on the surface and the T helper cells would detect this and be activated, which would activate the B cell in turn
CD40 and CD40L confirms that the cell is a T-helper cell and nothing else
The third signal it requires are the 2 cytokines produced
Compare T-cell independent and dependent B cell activation
What is affinty maturation?
= to fine tune antibody affinity to antigen
- After differentiation and diversity, there will be many naïve B cell’s not exposed to antigen yet
- Each B cell has a unique BCO receptor on the surface
- The best fit for the pathogen that will fit the unique receptor will make clones of itself = clonal expansion
- Then the clones will undergo affinity maturation - to improve the affinity of antibody to the antigen
What is the difference between a low-affinty antibody and a high-affinity antibody during affinity maturation?
Where does affinity maturation take place?
In the germinal centre of the lymph node, which are circular cell clusters
- Made up of light and dark zones
- Clonal expansion takes place in dark zone, followed by affinty maturation and selection
- follicular dendritic cells will present antigens on the surface the B cells have to compete for limited amount of antigens and then present to Tfh cells to give survival signal
- Class switching takes place in the light zone with the B cells with the highest affinity
