adaptive immunity - b cells Flashcards
brief description of the 3 mechanisms of humoral immunity
Abs promote pathogen neutralization, opsonization, complement activation
what is IgM?
Immunoglobulin M, an antibody class that serves as a receptor on naïve B cells
name properties of B cells
- antigen-specific
- ## clonotypic
what are plasma cells?
-Activated and differentiated B cells.
the main antibody-secreting cells
what are plasmablasts?
B cells in a lymph node that already show some plasma cells features but don’t secrete (or not a lot of) antibody
what do B cells do once they are activated?
secrete BCR (antibody)
what happens during clonal selection?
activated B cells undergo proliferation and differentiation until they become plasma cells
what is B cells signal 1? (survival signal)
antigen
through what vessel do b cells enter and exit lymph node?
enter via HEV
exit via efferent lymphatics
in what cases do naive B cells die?
after a few month if they haven’t encounter signal 1 (antigen)
does b cell bind antigen on MHC?
No! binds antigen DIRECTLY
what are the 3 ways how antigen end up in lymph node?
- from pathogens via afferent lymphatics
- has been opsonized via complement pathway
- transported via activated conventional DCs & transferred to follicular DCs
what are subcapsular sinus macrophages?
SCS macrophages: lymph node resident macrophages that express complement receptors on their surface to bind the complement on the opsonized antigen.
retain antigen in the lymph node.
what does it mean that SCS macrophages have low endocytic and degradative activity?
they don’t phagocytose antigens, they keep them on their surface in the lymph node
what does B cell bind to specifically?
epitope on antigen
what are B cell’s co-receptor for signal 1? what do they bind?
CD19 and CD21: bind complement (they are not necessary but enhance signaling)
what happens in BCR when it encounters antigen?
Igalpha and Igbeta domains associated to BCR become phosphorylated (well their ITAM motifs get phosphorylated)
what happens to BCR-Ag complex after phosphorylation of ITAMs?
it gets endocytosed! (internalized) with the Ag
3 main outcomes of signal 1
- Transcription factors are activated -> gene transcription
- Survival signal
- Cytoskeletal reorganization
what happens to the internalized BCR-Ag complex?
gets processed and presented on MHC so it can interact with TCR
why is antigen processing by B cell considered exogenous pathway?
because it’s the cell that decides when to endocytose.
If it’s the pathogen that wanted to enter the cell, it would be endogenous pathway.
what are TD vs TI antigen?
thymus-dependent vs thymus-independent (highly repetitive molecules, ex LPS and cross-link on BCR)
what are the difference between TD and TI?
TD-antigen provides signal 2 by an activated CD4+ Tfh cell via BCR.
TI-antigen provides signal 2 by TLR signaling!
can TD or TI lead to generation of memory B cells?
only TD antigens are specific Ab and can provide memory
what are the 2 parts of signal 2?
- Signal from pMHC that has bound to TCR & co-receptor on TFH cell
- Signal from CD40 on B cells that has bound to CD40L on TFH cell
what are the outcomes of signal 2?
- signaling & activation of transcription factors
- Leads to activation, proliferation, differentiation: antibody secretion
what cytokine is always secreted in high amounts by TFH to help with proliferation?
IL-12
what is linked recognition?
for signal 2. the same antigen (can be different epitope) must be presented to T cell by DC AND by B cell, but via a different route
where are SCS macrophages located?
subcapsular sinus. where they encoutner Ag
what is in the T cell zone?
T cells: they get activated by DCs there
what’s in the B cell zone?
it is where B cells encounter Ag (signal 1) and undergo later stages of proliferation and differentiation.
where follicles and germinal centers are found.
what happens at the T-B border?
B cell’s signal 2
what happens in the follicle vs in the germinal center?
follicle: activation and development of B cells
germinal center: B cell proliferation and differentiation
what changes happen to B cell after signal 1?
increased expression of pMHC II, increased expression of chemokines receptors to target to T-B border
what are activated B cell’s 2 options?
- form primary focus in subcapsular region
- migrate to follicle to form germinal center
what happens in primary focus?
B cells go near subcapsular region and become plasmablasts
what antibodies can be secreted by plasmablasts?
the basic IgM
difference between naive B cell and plasmablast?
naive B cell has surface IgM but can not secrete antibody.
plasmablast has surface Abs and can secrete them, and can proliferate
what other than plasmablast can come out of primary focus?
IgM MEMORY B cells that produce IgM
where can primary focus form?
- near subcapsular region
- in interfollicular regions
- medullary cords
how long does it take for primary focus to be apparent after a primary infection?
5 days
what happens to plasmablasts?
- USUALLY stay in lymph node and die by apoptosis within 5-10 days
- migrate to bone marrow and become plasma cells and continue antibody production
what are the 2 outcomes of germinal center?
- plasma cells: secrete large quantities of high affinity antibodies
- memory B cells: important for memory response
how do memory B cells maintain their memory response?
they maintain the capacity to produce high affinity antibodies
what happens in germinal centers?
- b cells recieves signals 1 and 2 again
- b cells differentiate
- somatic hypermutation, affinity maturation, class switching
what’s another name for germinal center
secondary lymphoid follicle
how long does it take for germinal center to peak in size?
7-12 days
name 3 ways how antibodies can act?
- neutralization
- opsonization
- complement activation
what’s an antibody?
BCR: Y shaped glycoprotein
what is a regular antibody structure?
2 identical disulfide-linked heavy (H) chains and 2 identical light (L) chains
where is the variable region on an antibody? what does it form?
on both the heavy and light chain, on the N terminus.
forms the 2 identical antigen-binding sites
what does Vl, Vh, Cl, Ch stand for?
- VL : Variable region in light chain
- VH : Variable region in heavy chain
- CL : Constant region in light chain
- CH : Constant region in heavy chain
what can result from variable region of Ab binding to antigen?
neutralization of the antigen
what is the role of antibody’s constant region (Fc)?
- involved in complement activation
- can bind Fc receptors on phagocytes, eosinophils, mast cells
what are Fab fragments?
Fragment Antigen Binding: 2 fragments on an antibody that bind the antigen.
part of constant heavy AND light chain.
what is Fc fragment?
one fragment on an antibody on the constant region of one of the heavy chain.
is recognized by antibody receptors.
what are the heavy and light chains held together with?
disulfide covalent bonds
what type of 3D structure if found in antibody?
Beta strand
what are where are CDRs?
Complementarity Determining Region: antigen-binding site in direct contact with Ag made of 3 hypervariable loops.
found at the antibody extremities.
what type of bonds are involved in
ab-ag binding?
non-covalent bonding:
* Hydrogen bonds
* van der Waals
* Hydrophobic
* Ionic
where on the antigen is the epitope?
can be anywhere
what varies in length on the antibody to confer specificity?
CDR
what differs between the different classes of antibodies?
- length/sequence of the constant region of heavy chain
- number of Ig-like domain
describe IgM
- pentameric: 5 antibodies linked via disulphide bonds
- heavy chain = one variable region and 4 constant regions
- expressed on mature B cells BEFORE activation
what antibody is the most effective initiator of complement cascade
IgM
describe IgD
- heavy chain = one variable region, 3 constant regions
- part of the first wave of secreted antibodies (right after IgM)
describe IgG
- heavy chain = one variable region and 3 constant regions
- most abundant in plasma
- has 4 subclasses
when is IgG produced?
following differentiation in Germinal Center
describe IgE
- heavy chain = 1 variable region and 4 constant regions
- produced in response to Helminth infections
describe IgA
- heavy chain = one variable region and 3 constant regions
- monomer in plasma, dimer in mucous
- dimer linked via J chain
- 2 subclasses
give 2 examples of antibody treatment
- antibody against TNF-a against Rheumatoid arthritis
- antibody against IgG Fc region
where are the heavy and light chain encoded?
on different chromosomes
how many CDRs are there in 1 Ig?
12 (3 per chain)
each chain as which region genes?
- Light chain: Variable (V) and Joining (J) constant (C) region gene
- Heavy chain: Variable (V), diversity (D), joining (J), and constant (C) region gene segments
where are CDR1, 2, and 3 encoded?
CDR1 and 2 are encoded in the V segments of light and heavy chains.
CDR3 is encoded in the joining of V-J segments of light chain; and in the V, D, J gene segments of the heavy chain
what CDR is the most variable?
CDR3
in what order are the different gene segments found? where?
light chain variable region: C, V, J
heavy chain variable region: C, V, D, J
what chain has 2 different loci (only one is expressed)? name them
light chain.
k chain or lambda chain
what do antibody’s different constant regions represent?
the different isotypes (IgM, IgD, IgG)
is somatic recombination reversible?
no it’s irreversible
what is combinatorial diversity?
Diversity that is the result of different combinations of V, D, and J regions
what are RSSs and their role?
Recombination Signal Sequences: they flank each antibody gene segment and get recognized by recombinase enzymes.
They direct the pairing of segments
what are RSSs structure?
nonamer + heptamer sequence in which either a 12 or 23 bp long spacer lies
what’s the 12/23 rule?
a 12-bp RSS must pair with a 23-bp RSS for recombination to occur. this allows for only 1 segment of each to be chosen
in what order does the heavy chain build itself?
D-J, then V-DJ
how long is the spacer of RSS for VH (variable region of heavy chain)
23 bp
what is RAG? what are they for?
Recombination Activating Gene: RAG-1 and -2 are necessary for recombination. they recognize and cut DNA at the RSS and COVALENTLY close the DNA hairpin ends.
what is the signal joint?
loop of DNA that gets excised and deleted
what is the coding joint?
Coding region of selected V and J regions that remains
what is junctional diversity?
During recombination, nucleotides may be added or removed at the junctions between V & D, and D & J (or the V & J for the light chain)
what is artemis?
an endonuclease that opens the DNA hairpin once signal joint has been excised
what can REMOVE nucleotides on each side of the coding joint?
exonuclease
what kind of nucleotides can be ADDED at overhangs ends?
palindromic nucleotides: TCGA in 5’ -> 3’
what can add up to 20 N-nucleotides to the cleaved strands primarily in the heavy chain?
TdT
what does N-nucleotides addition by TdT cause?
variation in length of CDRs
in what order do the enzymes work to generate BCR diversity?
RAG1//2, Artemis, Exonuclease, TdT
what is TCR made of?
one alpha chain, one beta chain, each with a variable and a constant region
what segments are found on each TCR chain?
alpha: V and J segments
beta: V, D, J segments
where does TCR somatic recombination take place?
in the thymus
what is the difference between BCR and TCR recombination?
BCR: D segment is surrounded by 2 RSS of 12bp spacing
TCR: D segment is surrounded by a 5’ 12 bp RSS and a 3’ 23 bp RSS
how many CDR does TCR have?
6 (3 on each chain)
what CDR is the most important for diversity in TCR? where is it encoded?
CDR3: encoded in D and J segments of TCR chains
where are CDR1 and 2 encoded in TCR?
they are encoded in the V segment of alpha AND beta chains
what is allelic exclusion for?
ensure that each B cell synthesizes only one heavy and one light chain, even if we receive 2 of each from parents
how does allelic exclusion happen?
one chromosome gets methylated and silenced
what antibodies are expressed by mature but naive B cells?
IgM, IgD
how does the production of Ig subtypes happen?
Alternative RNA splicing
what can alternative splicing also decide?
if an mRNA is secreted or transmembrane
where does secondary diversification happen?
in germinal center in lymph node follicles
what does somatic hypermutation cause?
increases affinity for a BCR’s antigen, without changing specificity
what is class switching?
a process that replaces one heavy chain constant region with one of a different isotype
where do somatic hypermutations happen in the BCR?
in the variable region, mostly in CDR loops
how to BCR know what “class” to switch to? (what isotype)
from TFH cytokines released in the germinal center
can naive B cells undergo somatic hypermutation and class switching?
no, only activated B cells
what structure in BCRs guide class switching?
switch regions upstream of C genes
what extra diversity is found in BCRs that isn’t in TCRs?
- H and L chain combination
- secondary diversification (somatic hypermutation and class switching)
what is found in germinal center light zone?
TFH (retain antigen in the light zone)
can B cells who enter germinal center produce antibodies?
yes they can produce IgM/IgD
where does secondary diversification happen?
germinal center
what mostly happens in the light zone of germinal center?
plasma and memory cell differentiation
what happens in the dark zone of germinal center?
somatic hypermutation
what are FDCs?
Follicular Dendritic Cells: DCs in germinal center that condense Ag
where are FDCs?
in light zone of germinal centers! because that’s where differentiation happens
how does affinity maturation work?
higher affinity B cells bind antigen in germinal center light zone and recieve signal 1 again.
lower affinity B cells don’t and die by apoptosis.
what are the outcomes of germinal center?
- plasma cells: secrete IgG, IgA, or IgE antibodies of high affinity
- Memory B cells:
what is AID?
Activation-Induced cytidine Deaminase: responsible for somatic mutations
how does AID work?
it deaminates cytidine residues in ssDNA: cytidine -> uridine. Uridine then gets removed. produces individual point mutations in the variable chain
are all AID changes productive?
no
when does class switching occur?
in the germinal center after the second exposure to antigen (2nd signal 2)
what is required for class switching?
complete signal 2 (Ag + CD40)
between what regions does recombination occur in BCR?
between switch regions
in class switch recombination: When B cell receives cytokine signal, transcription is activated _______ of constant region
upstream
after transcription begins, DNA is now accessible to what enzyme?
AID
how is immunological memory easier to detect/monitor?
Abs can be measured in serum
Memory T cells reside in tissues
where do most IgM (sometime IgD) producing cells come from in primary response?
primary focus
what do memory B cells express higher levels of than naive B cells?
MHC class II and B7 co-stimulatory ligand
why do memory B cells express more MHC class II and B7 molecules?
- makes it easier to present antigen to Tfh
- increases antibody production
where do memory B cells reside?
circulation, spleen, lymph nodes
what do memory T cells have a large expression of?
CD28: don’t require strong co-stimulatory signal or cytokines
what distinguishes Tcm (central memory t cells) vs Tem (effector memory t cells)
location (Tcm in secondary lymphoid tissue) (Tem in tertiary/peripheral tissue)
does passive immunization generate memory response?
no. no memory cells are made out of it because we inject preformed antibodies
what is the original antigenic sin?
when presented to a know infection, memory cells are enlisted instead of activating naive cells that target new unique epitopes
what are adjuvants?
molecules in vaccines that enhance the immune response
