L5 - Humoral responses Flashcards
Steps in B cell activation and differentiation
- B cells develop inbone marrow - precursor rearranges immunoglobulin genes to create BCR
- cytokines bind and promote survival
- migration to lymph nodes to wait to be activated
what are BCRs
B-cell receptors are cell surafce Immunoglobulins- Ig
same structure as an antibodyb but has a transmembrane region to tether it
what part of BCRs are involved in intracellular signalling
BCR alone cannot generate signal
Immunoreceptor tyrosine-based activation motifs (ITAMs)
on Iga and Igb hetrodimer
phosphorylation of tyrosine residues on activation cause intracellular signal
= P13kinase
what are antibodies made of (chains and regions)
2 identical L chains + 2 identical heavy chains
each light chain has:
- 1 variable region VL
- 1 consrant region CL
each heavy chain has:
- 1 variable region VH
- 3 constant regions CH1,CH2 and CH3
hinge protein in between
what is in the Variable regions of the heavy and light chains that binds to antigens
hypervariable/complimentatiry-determining regions
V region is made of beta-pleated sheets with loops between them
loops bind to antigens
how many CDRs regions /hypervariable per chain per antigen binding site (remeber 2 identical binding sites per antibody)
3
so 6 per side of Y
what is the orginal ‘germline theory’ for number of antibodies produced and why is this wrong
1 gene per antibody
not enough genes in genome to code for all possible antibodies
= let alone rest of bodies proteins
what is the answer to limited genome and unlimited antibodies
VDJ recombination
what are the 2 chromosones that code for light chains (heavu chain coded for by a different one)
lambda and kappa
every antibody is 1 kappa/lambda and 1 heavy chain
describe VDJ recombination - simple
light chain composed of smashing togther random gene segments :
1 V gene + 1 J gene
heavy chain made of:
1 V gene + 1 J gene + 1 D gene
constant regions join to the new gene and do not partake in recombination = effector function
what is the role of Recombination site sequnces (RSS) in VDJ recombination
short DNA sequnces that flank the regions - V.D and J
site of RAG recruitment and where it cuts
what is the 12/23 rule in VDJ recombination and how does it ensure thinsg work fine
There are 12 or 23 bases between RSSs
a 12 RSS can only bind to a 23 RSS
heavy chain V + J genes are ONLY 23 spacers
cannot come together
= ensures inclusion of 12 spacer D gene in the middle
name of the enzyme complex that undergoes somatic recombination
V(D)J recombinase
Includes:
1. lymphocyte-specific RAG-1 and RAG-2 endonuclease
- Ku70 and Ku80
role of RAG endonucleases
bind to RSS serqunces
recruit andother RSS
cleave between RSS and coding sequnce removing the non-chosen segments
= leaves hairpin loop
jucntional diversity mechansim - following on from RSS cleavage by RAG
- hairpin loops bound by Ku70/80 and recruit artmesis
- cleaves hairpin loop leaving open ends
- TDT binds - Terminal deoxynucleotide transferase
- randomly adds and removes nucleotides to gap until complimentartity found
= junctional diversity at CDR3
- DNA ligase heals nick
why is there increased diversity via junctional diversity at CDR3 compared to the other hypervariable regions
CDR1 and 2 coded by a single segment - a V region
no junctional diversity can occor here due to no cleavage by RAGs
= CDR3 includes V + J (and a D if the heavy chain)
= chance for random nucleotides to be added at end of gene segments
summary of VDJ corecomination: combinatorial diversity + jucntional diversity
combinatorial:
multiple V,D and J genes combine
any heavy chain can combine with any light chain to generate antigen-binding site
junctional:
random nucleotides added to ends of genes
what can B cells do AFTER encountering an antigen to produce higher affinity antibodies
somatic hypermutation
= T cells once matured never change affinity
what is somatic hypermutationn and what enzyme is most involved
B-cells continously undergo 1 or more amino acid changes in the hypervariable regions
AID enzyme - activation induced cytidine deaminase
where does somatic hypermutation take place
germinal centres in lymph nodes
1. Light zone:
somatic hypermutation site and rapid division of B-cells
- Dark zone:
affinity maturation - cells compete to identify who has highest affinity
Describe affinity maturation in steps (thinkl helper T cells)
- advantageous mutations in B cells bind and present antigen better
- T follicular helper (Tfh) cells produce selecting cytokines
- cytokines promote growth and survival
B cells can be activated by T-cell dependant or independant pathways - describe the differences
B cells always need 2 signals to be activated
dependant - protein antigens:
- BCR binds to antigen and presnets it via MHC 2
- T cells bind via MHC increasing expressiuon of CD40 leukin
= survival signal
independant - non-protein antigen:
1. antigen binding to BCR
2. binding of PAMP to TLR
= produces NFkB transcription factor
why do T-cell independant activations of the B cells look like innate responses
No memory cells produced
No affinity maturation or somatic hypermutation
No class switching of antibodies
= short lived plsams cells that cannot clas siwtch or undergo affinity maturation
= T cells provide the signals needd for all these things via cytokines and bidning of CD40
term for chaning the Fc region of an antibody
class switching
= no change in affinity (Fab) but changes effector functiom
