Adaptive Immunity Flashcards
Adaptive immune responses are initially slower to develop than innate responses. What can it do that innate can’t?
Shows specificity and memory
Where do B and T lymphocytes acquire antigen receptors?
B lymphocytes acquire them in the bone marrow
T lymphocytes acquire them in the thymus
What happens to antibody when stimulated by presence of antigen?
Cells release antibodies (receptors) which bind the toxin so it can be eliminated
Difference between integral membrane proteins on B lymphocytes and soluble proteins secreted by plasma cells?
Integral membrane proteins on B lymphocytes – Antigen receptors
Soluble proteins secreted by plasma cells – Antigen eliminators
Explain the 4 chain structure of antibody?
Arms (N termini) recognise antigen and bind
Tail (C termini) initiates elimination of antigen
Arms and tail joined by disulphide bonds, as well as a hinge allowing them to move relative to eachother
2 arms each consisting of 2 domain light chain and 2 domains of heavy chain
Tail consisting of 2 domains from both heavy chains; These connect to arm heavy chains
What did cleavage of antibodies with papain uncover? (hint - 2 fragments)
One of the fragments bound to antigen – Fragment Antigen Binding (Fab)
- When the antibody is cleaved by pepsin, the F(ab’)2 fragment can bind antigen divalently, like it would in the intact molecule
The other fragment was found to be crystallised in solution – Fragment Crystallisable (Fc)
- Interacts with elements of the innate immune system (antigen elimination)
What are the 5 immunoglobulin classes that differ in the amino acid sequence of the heavy chain? (hint - one has unknown traits)
Give traits for each
IgG – γ
- Main classes in serum and tissues
- Important in secondary responses
IgM – μ
- Important in primary responses
IgA – α
- IN serum and secretions
- Protects musical surfaces
IgD – δ
- ?
IgE – Epsilon
- Present at very low levels in serum
- Involved in protection against parasites and allergy
What are the 2 light chain types?
Are they class restricted?
Kappa
Lambda
These are not class restricted; Can have IgG-Kappa or IgG-Lambda antibodies
What do the variable and constant regions do? Different specificities?
Where are they?
V region - Bind antigen; Differ between antibodies with different specificities
- Domains at N terminus
C region - Same for antibodies of a given H chain class or L chain type
- 6 domains
Describe the homologous nature of antibodies? (hint - domain names)
Hypothesised to form a series of globular domains, each stabilised by an intra-chain disulphide bond
- V-H, V-L
- CH1, CH2, CH3; On each chain
- CL1; On each chain
What is the immunoglobulin fold?
Specific folding pattern of antibody domains
C domain has 7 beta strands
V domain has 9 beta strands
What is the immunoglobulin gene superfamily involved in?
Recognition, binding, adhesion
What are the hypervariable regions? (hint - CDRs)
Loops on the end of V domains in Fab
3 complementary determining regions (CDRs) on the end of both light and heavy chains
- CDR1-3
What interactions are involved in antibody-antigen binding?
Strength? Specificity and affinity?
Non-covalent e.g. electrostatic, H-bonds
Individually weak but if many form simultaneously due to high complementary between epitope and antibody, the interaction is specific and high affinity
What immunoglobulins do B cells use as a receptor and how do they initiate signalling? (hint - ITAM)
IgM and/or IgD used as B-cell receptors
These recognise and bind antigen but can’t generate a signal
Membrane immunoglobulins are associated with 2 other proteins, Igα and Igβ
These contain ITAM (Immunoreceptor Tyrosine Activation Motif) in their tail to initiate signalling when antigen binds
What are the main determinants of antibody diversity? (hint - CDRs)
Where does most variability come from?
Variations in the sequence and length of CDRs are the main determinants
CDR3 tends to be most variable
Heavy chain contributes more to antigen binding and is more variable than the light chain
What are the 3 big things that drive antibody diversity?
What do they give rise to? (hint - V regions)
Multiple genes
Somatic Mutation
Somatic Recombination
Somatic recombination and mutation gives rise to a limited number of inherited gene segments, which make up the V regions
How many sets of immunoglobulin genes are there and what/where are they?
How is variability achieved? (hint - V and C region genes)
Heavy (H) chains - Chromosome 14
Kappa (κ) chains - Chromosome 2
Lambda (λ) chains - Chromosome 22
Each locus has multiple V region genes and one, or a few, C region genes
Somatic recombination of V region genes generates variation
How many DNA segments encode light chain V regions?
What segments?
2 segments of DNA
V and J (joining) regions
How many DNA segments encode heavy chain?
What segments?
3 segments of DNA
V, J and D (diversity) regions
How and when does rearrangement of light and heavy chains genes occur?
Somatic recombination with V gene being spliced to a J gene; Intervening DNA is excised
New V gene is transcribed with intervening sequences removed by RNA sequencing
Occur during B cell differentiation, leading to permanent changes in the DNA
What gene segments does CDR3 correspond to and why?
VDJ
CDR3 is on heavy chain and these are the heavy chain gene segments
Somatic recombination involves lymphocyte specific recombinases and conserved recognition signal sequences (RSSs). What are RSSs and where are they found?
Conserved sequence that consists of heptamer (7bp) + nonamer (9bp); These are separated by 12 or 23 random nucleotides
Found directly adjacent to the coding sequence of V, D or J gene segments
What do RSSs do?
What is the 12-23 base pair rule?
Guide rearrangements of the V, D and J segments
12-23 base pair rule – Gene segment with a 12bp spacer only joins with a gene segment with a 23bp spacer; Ensures correct V-D-J joining
What enzymes are in the V (D) J recombinase complex? (3 key enzymes)
Normal DNA cleavage/repair enzymes
RAG1-RAG2 protein complex (encoded by Recombination Activation Genes)
Specialised endonucleases e.g. Terminal deoxynucleotide transferase (TdT)
What does the RAG1-RAG2 complex do during somatic recombination?
Recognises and aligns the RSSs adjacent to the gene segments to be joined
Complex has endonuclease activity and cleaves the DNA
Cleaved DNA is repaired to form the coding joint (V and J segments now next to one another) and the signal joint (intervening DNA excised into loop)
How is diversity in immune repertoire achieved via antigen independent mechanisms? (3 things) (hint - -combination)
Multiple copies of each V region gene segments
- [Vn x Jn]
- [Vn x Dn x Jn]
Heavy and Light chain combination
- [Vκ x Jκ] + [Vλ x Jλ] x [VH x DH x J]
Recombination is imprecise which leads to junctional diversity
- Nucleotides may be lost or added; Variable addition of nucleotides at junctions massively contribute to CDR3 diversity
How is diversity in immune repertoire achieved via antigen dependent mechanisms? (1 thing)
Somatic mutation of V regions following antigen activation
- Point mutations
- Base changes tend to be clustered in CDRs
Steps 1-3 in process of somatic recombination
End result? (hint - signal)
- RAG-1-RAG-2 complex recognises and aligns the RSSs adjacent to the gene segments to be joined
- Two ssDNA breaks are made close to the RSSs
- Free 3’-OH attacks phosphodiester bond on other strand of DNA to create a hairpin at the segments to be joined and a flush ds break at RSS boundary
(The blunt ends formed at Stage 3 are ligated to form the “signal joint” and this DNA is typically excised)
Steps 4-8 in process of somatic recombination (just formed hairpins) (hint - imprecise)
- DNA hairpins are cleaved at random, symmetrically
- Or asymmetrically
- For V-D-J joining of the H chain, nucleotides can be added by terminal deoxynucleotide transferase (TdT)
- Unpaired overhangs are filled in by DNA polymerase or may be excised by an exonuclease
- DNA ligase joins the nicked and repaired hairpins to form the “coding joint”
What is somatic hypermutation?
How does it occur? (hint - AID)
Where are mutations clustered in mature B cells?
Point mutations introduced in rearranged V regions
Activation-induced cytidine deaminase (AID), an enzyme expressed by B cells in lymphoid tissue responding to antigen
Clustered in CDRs
Somatic hypermutation can add diversity, but what is its main function?
Affinity maturation – Higher affinity receptors selected as immune response proceeds – “Survival of the Fittest”
What is class switching?
What are its effects? (hint - flexible response)
Same recombined V region associates with different C region genes, changing antibody heavy chain e.g. IgM –> IgG, IgA etc.
Antigen specificity retained, different localisation/effector functions induced
- Flexible response to pathogens
How can class switching occur?
- Reversible?
How is the mechanism different to V-D-J joining? (hint - AID)
DNA recombination between switch regions
- Intervening DNA lost
- Irreversible
Different mechanism to V-D-J joining; Initiated by AID acting at switch regions
What is mechanism of AID activation for class switching and somatic hypermutation?
AID deaminates cytidine to form uracil
Activity triggers DNA repair pathways
Repair pathways in B cells are error-prone, leading to different mutational outcomes:
- Mismatch repair, base excision repair – Somatic hypermutation
- Single strand nicks –> Double strand nicks (common in G-rich tandem repeat switch regions) – Class switching
Why must AID be tightly regulated?
AID mutation causes immunodeficiency – Hyper-IgM Syndrome Type 2
How are T lymphocyte receptor different to B cell?
Acquired in the thymus
Receptors are only expressed on membranes, not as soluble proteins
What are the 2 major subpopulations of T cells?
Give CD_ +ve for both
What do they do?
T helper cells (CD4 +ve) - Augment immune responses
T cytotoxic cells (CD8 +ve) - Specifically kill infected host cells
How are T lymphocyte receptors (TCRs) similar to immunoglobulins?
Which parts of which chains are most variable?
Extracellular domains of the TCR are homologous to variable and constant regions of immunoglobulins
- Each V region contains 3 CDRs
CDR3 regions of α and β chains are the most variable
What chromosomes are T cell receptor α and β chains on?
CD3 subunits are a part of the TCR complex. What do they contain?
α - Chromosome 14
β - Chromosome 7
Contains ITAMs (Immunoreceptor Tyrosine Activation Motifs) in their cytoplasmic regions
How does somatic recombination occur in TCR V region?
Same way as in B cells
How is diversity achieved in TCR genes? (3 ways) (hint - similar to B cell)
What do V regions of TCRs not undergo?
- Multiple copies of V region gene segment [Vn x Jn/Vn x Dn x Jn]
- α and β chain combination [Vα x Jα] x [Vβ x Dβ x Jβ] = ~ 6 x 106
- Junctional diversity
- Concentrated in the CDR3s of TCR α and β chains
- TdT active throughout T cell receptor gene rearrangement
V regions of TCRs do not undergo somatic mutation; Why?
Why is B cell immunity important in defence against extracellular pathogens? (hint - native)
B cells recognise free “native” antigens
How do T cells recognise intracellular antigens? (hint - processing)
Major Histocompatibility proteins (MHC) which presents processed antigen at cell surface for T cell recognition
What context can T lymphocytes ONLY recognise antigen in?
Why is this?
Context of self-MHC molecules
TCR recognises complex of peptide + self-MHC
- CDR1 and CDR2 bind self-MHC
- CDR3 binds peptide (antigen) that is bound to MHC
What are the 2 classes of MHC proteins involved in antigen recognition?
What cells express them?
MHC Class I
- Expressed by all nucleated cells
- Present peptides derived from ENDOgenous proteins to CD8 +ve T cells
MHC Class II
- Expressed by certain leukocytes e/g/ dendritic cells, macrophages, B cells
- Presents peptides derived from EXOgenous proteins to CD4 +ve T cells
What are the traits of the 2 domain types in MHCs, membrane-proximal and -distal?
Proximal - Ig like
Distal - Bind peptide via peptide binding cleft
Membrane distal domains in MHC are polymorphic, are inherited and don’t rearrange. What does this mean in general and for diversity and specificity?
Polymorphic so take many forms; Very broad specificity, binding a wide range of peptides
Lower diversity than B and T cell receptors
Size of peptides bound by MHCI and MHCII?
MHCI - Binds peptides 8-10 a.a. long
MHCII - Binds peptides 13-18 a.a. long
Process of antigen presentation by MHCI (4 steps) (hint - TAP)
Endogenous meaning?
Endogenous - Antigen is made within host cell that is infected
Intracellular antigen transported to proteasome to break down misfolded proteins
Then transported to ER by TAP (Transporter Associated with antigen Presentation)
Peptides loaded onto MHCI in ER
MHCI-peptide transported to cell surface for recognition by cytotoxic T cell
Process of antigen presentation by MHCII
Exogenous meaning?
Exogenous - Antigen is taken up from outside the host cell
Antigen taken up by phagocytosis/endocytosis
Acidification in vesicles promotes unfolding and proteolysis
Peptides associate with MHCII in the endocytic compartment
MHCII-peptide transported to cell surface for recognition by helper T cell
What is cross-presentation?
What does this allow and when is it important? (hint - avoid infection)
Some dendritic cells present exogenous antigen associated with MHCI to cytotoxic T cells
Allows antigen presentation to cytotoxic T cells without the dendritic cells themselves being infected
Important in cytotoxic T cell responses to many tumours
Where does most allelic variation occur in MHC genes?
Predominantly in the peptide-binding regions
Consequences of MHC polymorphism?
Graft rejection
Ensures wide recognition of foreign peptides but variability of MHC molecules is small compared to that of TCR
T cell responses determined by an individual’s MHC type
Association with certain autoimmune disease
CD4 and CD8 both contain Ig like domains, allowing them to act as what for the TCR complex? (hint - co-)
What 2 things require this function?
Allows them to act as co-receptors
Required to stabilise interactions and facilitate signalling
What does engagement of CD4/CD8 co-receptors with TCR complex enhance? (hint - ITAMs)
Enhances phosphorylation of the ITAMs, promoting T cell activation
What is Thymic selection? (hint - +ve and -ve selection)
Process to select T cells which correctly identify self-MHC and MHC + peptide
+ve selection - TCR doesn’t bind self-MHC so it is rejected
-ve selection - TCR doesn’t bind self-MHC + peptide so it is strongly rejected
IgM traits
- Structure
- Location
- Hinge?
- Affinity and avidity (strength of the antibody-antigen interaction)
Pentamer (5 antibody monomers + J chain)
Usually serum-restricted
No defined hinge region
Low affinity but high avidity
IgM monomer traits
- Valency (number of binding sites)
- Complement
- Primary response
- Hinge?
- High valency (deca- /pentavalent) –> Good agglutinator of particulate antigen
- Can activate complement very efficiently
- Important in primary antibody responses
Has functional hinge
IgA secretory form
- Structure
- Valency
- Complement
- Monomer binding
IgA dimer + J chain + Secretory component
High valency
Doesn’t activate complement
Monomer binds Fc receptors on phagocytes
How does secretory IgA transport bacteria that has penetrated mucosa back into lumen?
What is IgA’s passive role?
IgA secreted in submucosa and binds pathogen
Transport system then takes complex through epithelia into lumen
IgA has passive role in preventing adhesion/infection
Where is IgG D presented, along with IgM?
Where is it produced and what does it interact with?
Present as antigen receptor on many B lymphocytes, together with IgM
Produced by B cells/plasma cells in upper respiratory tract
Interacts with receptors on basophils, inducing antimicrobial, inflammatory and B cell stimulatory factors
IgE traits
- Hinge?
- FcR
- Affinity
- Allergy and parasites
No defined hinge
Binds with high affinity to FcR on mast cells and basophils
Important in allergy and defence against large extracellular parasites
What are the biological roles of immunoglobulins? (7 roles)
Label pathogens for elimination/destruction
Specific binding/multivalency:
- Antibodies are at least divalent; Avidity vs affinity
Immobilise pathogens – IgM
Agglutinate particles e.g. bacteria – IgM, IgA
Form “immune complexes” with soluble antigen
Block binding of pathogens to host cells – IgG, IgA
- E.g. antibodies to bacterial adhesins or viral receptors
Neutralise toxins (e.g. tetanus, diphtheria, cholera) – IgG, IgA
What are the Fc Effector functions? (3 functions)
What do these mechanisms depend on?
Invoke destruction of labelled pathogens
Activate complement
Bind Fc receptors on leukocyte surfaces
Depend on:
- Site and type of infection
Stage of the immune response; Primary or secondary
What does C1 consist of?
What is produced from C1 when complement is activated?
How does classical complement activation occur? (hint - IgG)
C1 = C1q + C1r + C1s
C1r is a serine protease that cleaves and activates C1s, another serine protease
C1q must interact with 2 Fc regions for activation to occur; Cross-linking
Which antibody is a more potent complement activator than IgG and why?
IgM as C1q can bound to one IgM with its pentameric structure as opposed to 2 for IgG
How do IgG and IgA act as opsonins?
Bind bacteria with Fab regions
Fc regions bind Fc receptors (FcRs) on phagocytes, inducing phagocytosis
What do FcRs do for pathogens too big to phagocytose?
Which antibodies are involved?
Release lysosomal contents
IgG and IgA
What does IgG binding to FcRs on NK cells facilitate? (hint - ADCC)
How?
Antibody-dependent cell-mediated cytotoxicity (ADCC)
Binding of IgG-coated targets cells to FcRs on NK cells
Cross-linking of FcRs causes degranulation of enzymes and perforin to kill pathogen
What is mediated and what happens when IgE binds FcRs on mast cells and basophils?
What form of antigen is needed and why? (hint - multi-)
Mediate allergy/defence against large parasites
Mast cells secrete inflammatory mediators and cytokines
Multivalent antigen to cross-link antibody
B cell response to certain antigen doesn’t require T cell help. What is this called and how does it work?
Thymus independent antigens
Multiple cross-linking of B cell receptor by antigen induces a rapid response and production of IgM antibodies
B cell response to certain antigen requires T cell help. What is this called and how does it work?
Thymus dependent antigens
Require T cells for differentiation of B cells into plasma cells
Long-lived memory B cells may also be generated
Responses can involve somatic hypermutation and class switching
