Humoral Immunity Flashcards
Revision: Describe the structure of an antibody (Immunoglobulin)
- Variable region - different
- Constant region - Same for all Ab of the same class
- Antigen binding site
- Light chain - k or lambda
- Heavy chain - 9 different types
Describe the 2 different types of immunoglobulins
- Membrane bound receptor
- secreted antibody
Describe the structure of the antibody
- Heavy chain is made up of 4 parts: Vh, CH1, CH2 and CH3
- The light chain is made up of 2 parts: Vl and Cl
- Disulphide bridges hold the structure together
- The hinge region provides flexibility to the structure
- Glycosolation on CH2 to help with interactions
What is the function of the complementary determining region
- Finger-like structures that are responsible for antigen binding
- On the light and heavy chain
List the functions of antibodies
- Virus and toxin neutralisation
- Opsonisation + ADCP
- Complement fixing/ MAC formation
- Opsonisation + ADCC
What are the different classes of antibodies
- IgG
- IgD
- IgA: Dimer of 2 antibodies
- IgE
- IgM: 5 monomeric molecules
What different heavy chains does each antibody class use
- IgM - u (mu)
- IgD - s (delta)
- IgG - y (gamma)
- IgA - a (alpha)
- IgE - e (epsilon)
What are the molecular weights off each antibody type
- IgM - 900k
- IgD - 180k
- IgG - 150k
- IgA - 385k
- IgE - 200k
What antibody types fixes complements
- IgM
- IgG
What antibody type can cross the placenta
Only IgG
What does the constant region bind to in antibodies
- In IgG it binds top Phagocytes
- In IgM it binds to mast cells and basophils
What is the main function of IgM antibodies
- Main Ab of primary response
- Best at forming immune complexes and fixing complements
- Monomer serves as BCR
What is the main function of IgD antibodies
- The only Ab that isn’t secreted
- B-cell receptor
- Indicated mature b cell
What is the main function of IgG antibodies
- Main Ab of secondary responses
- Neutralises toxins
- Opsonization
What is the main function of IgA antibodies
- Secreted into mucous, tears, saliva and colostrum
What is the main function of IgE antibodies
- Allergy
- Anti-parasites
What is class switching
- Only affects heavy chain constant region
- Allows for different effector function to deal with different pathogens
What are they types of class switching
- Minor class switching - Differential splicing
- Major Class switching - DNA recombination
How doe step B cell know which class to switch to
- Sensing chemicals around them produced by T helper cells
What is required by Class switch recombination to work on heavy chain switching
- Cytokine signal
- Switch regions
- AID and DSB repair proteins
What are the key prerequisite for CSR
- recombination occurs between switch regions
- Switching can only occur downstream
Explain how does Class Switch Recombination works
- When a signal is received to class switch AID and DSB repair proteins are activated
- The switch regions from the unwanted regions are pulled together
- This part is cleaved and formed into a Switch DNA circle
- The VDJ segment is transcribed as normal and the new antibody class is expressed
What are the structural differences between secreted IgG snd membrane bound IgG
- Membrane-bound IgG has a hydrophobic transmembrane region and a cytoplasmic region which allows it to be anchored to the cells’ membrane
- Secreted IgG has a normal tail piece which allows it to be free floating
Explain how differential splicing allows creation of secreted and membrane-bound antibodies
- Polyadenylation at 2 different sites allows for the formation of one variant with tailpiece and another with membrane domains
Revision: What is the difference between Somatic recombination and differential splicing
- Somatic recombination is the alteration of the DNA itself to induce change in expression
- Differential splicing are changes made in mRNA to alter the final product, doesn’t affect original DNA
What are the stages in B cell development
- Antigen independant stage
- Antigen dependant stage
Describe the formation of a B cell in the bone marrow
- Stem cell becomes Pro-B cell
- Pro-B cell undergoes D-J and V-DJ recombination to code in the heavy chain variable region - expressed as the mu heavy chain and becomes a pre-B cell
- Pre-B cell undergoes another V-J recombination to code in the Light chain variable and constant regions and become Immature B cells
- Once they express IgM and IgD from differential splicing they become mature B cells
What happens to B cells when they encounter a pathogen
- Further mature the ability to bind to pathogen to GC
- Class switch to appropriate effecter regions
- Become either IgG or IgA
- Become plasma or memory cells
How many unique B cell receptors do we have
1 billion unique receptors
What gene segments make up the antibody light chain
- V, J and C
What gene segments make up the antibody heavy chain
- V, D, J and Cmu
How does V (D) J recombination happen to produce unique heavy and light chains
Different gene segments are alternatively spliced together to make unique heavy and light chains
Where are the gene loci encoding for Ig located in the genome
- There are 2 light chain loci and one for the heavy chain
- Lamda on chromosome 22
- Kappa on chromosome 2
- Heavy chain on chromosome 14
Describe the process of VJ recombination
- V and J gene segments are chosen at random
- The gene segment is transcribed
- The transcript is then polyadenylated and spliced
- The transcript is translated to a amino acid sequence
- The sequence folds to become a protein
Describe the process of VDJ recombination
- V, D and J gene segments are chosen at random
- The gene segment is transcribed
- The transcript is then polyadenylated and spliced
- The transcript is translated to a amino acid sequence
- The sequence folds to become a protein
- The C segments determines what types of antibody it is
What are recombination signal sequences in V(D)J recombination mechanisms
- They are conserved sequences upstream or downstream go gene segments
- ‘Turns’ consisting heptameter and nonamer with 12 or 23 bp spacer
Why do we have the one-turn/two-turn rule
- Recombination only occurs between a segment with a 12bp spacer and a 23bp spacer
- Prevents V and J fragments from accidentally recombining
What other ways does antibody diversity come from
- Junctional flexibility
- P-nucleotide addition
- N-nucleotide addition
- Combinatorial association of heavy and light chains
- Somatic hypermutation during affinity maturation
How is junctional diversity created
- Occurs during VDJ recombination via junctional diversity
- P and N nucleotide addition before end joining
- Also produces non-productive rearrangements
Describe the mechanism how which VDJ recombination occurs
- RAG 1 and 2 bind the turn ends and form a major hairpin structure
- Cleavage occurs at the turn ends to produce minor hairpins between the DNA strands
- More enzymes repair the ends and joins the ends together
Describe the hairpin opening and closing mechanism
- the hairpins are repaired by enzymes such as Artemis and endonuclease
- The enzymes mess around with the free DNA ends by adding or removing nucleotides
- The ends are joined together by other enzymes
How does Artemis cause junctional diversity
Artemis chooses at random what nucleotide to cut which changes the nucleotide code for future repairs
How does allelic exclusion work
- Two copies of each Ig gene – one from the mother and one from the father
- In other cells, both genes are expressed
- Antibody genes different – Only one heavy chain allele and one light chain allele is expressed
- Order of rearrangement: Heavy>kappa>lambda; 1st allele then 2nd
- Ensure each B cell makes one type of antibody
Describe the Antigen dependant phase of B cells
- T cell activates B cell by presenting pathogen antigen
- B cells migrate to germinal centre where it undergoes affinity maturation
- Affinity maturation involves: Clonal expansion, Somatic hypermutation and selection
- They undergo class switching to be more appropriate for the infection
- Become plasma and memory cells
What are the 2 stages of B cell activation
- T cell dependant phase
- T cell independent phase
Describe the T cell independent stage
- When a pathogen invades some B cells bind to antigens and become partially activated
- Undergoes clonal expansion
- Some clones secrete IgM and become the first defence
- Other clones migrate to lymph nodes awaiting T cell activation
Describe how B cells are activated in the lymph nodes
- Requires triple verification process
- Encounter rte pathogen and internalise antigen
- Antigen presented on B cell surface
- Activated by T cell which the same pathogen has activated
- CD40/CD40L handshake ensures it is a T cell
- Cytokine signalling is needed as well and the B cell undergoes affinity maturation and class switching
Describe the signal transduction pathways present in B cells
- BCR binding > Activation of tyrosine kinase
- Signal transduction pathway for cell proliferation, differentiation and survival
How does affinity maturation occur
- Aims to fine tuner antibody affinity to antigen
- After clonal expansion, the clones undergo affinity maturation
- Mutations occur in variable region genes and select for antibodies with the highest affinity for the antigen
- Takes place in germinal centre of lymph node
Describe the selection stage in the light zone
- Follicular Dendritic cells present antigen on the cell surface
- B cells compete to bind to antigen and present it to Tfh cells
- Cell with reduced affinity is apoptosed
- Survived b cells migrate back to the dark zone to repeat processes until affinity is high enough
