Cell and Molecular Biology of the Immune System Flashcards
What is specific/adaptive immunity?
-Induced by exposure to an infection
-Shows a high degree of specificity
-Exhibits ‘memory’
Effective immune response must…
-Not overreact to benign or self
-Direct different effector mechanisms against different pathogens
-Recognise and respond to any invading organism
Features of specific immunity
-Mediated by lymphocytes (B/T cells)
-Clonally distributed receptors
-Large repertoire - low freq. of cells specific for any antigen
-Response takes time to develop, memory cells produced
2 phases of an immune response
- Innate response
- Adaptive response
What is BCR?
-B cell receptor - antibody
-Membrane form of immunoglobulin binds ‘free’ antigen
-Subsequently secreted when B cell is activated now known as antibody
What is TCR
-T cell receptor
-a chains and B chain
-membrane form only
-doesn’t bind free antibody, recognises peptide fragment of antigen bound to MHC expressed by APC
Antibody structure
-Basic 4 chain structure - 2 heavy chains and 2 light chains held together by covalent and non-covalent bonds.
-2 types of L chains (lambda and kappa)
-Each chain has a variable and constant region
-Bivalent
How many classes of antibody are there
5 - IgM, D, A, G and E
Called isotypes - determined by heavy chain
What is the V region encoded by in H chain and TCRB?
3 gene segments (V, D and J - V is biggest)
What is the V region encoded by in L chain and TCRa?
2 gene segments (V and J)
How do gene segments come together when functional immunoglobulins are produced?
After DNA breaks, a single V and a
single J gene segment are joined
together to encode the V region of
the light chain. This process is random - there are two loci containing L chain genes: lambda and kappa).
Similarly, a single random V, D
and a J gene segment are joined
together in a single B cell
to encode the V region of the
heavy chain
Occur in the same B cell to end up with an antibody with randomly generated chain regions.
What order are Igs rearranged during B cell development?
-H chain gene segments rearrange (D-J, then V-DJ)(greater variability in H chain as V, D and J segments)
-Light chain gene segments rearrange - kappa segments (V-J) first
-If this is unsuccessful, then lambda segments (V-J) rearrange
What is the recombination process?
-Ig gene segment rearrangement is guided by special sequences flanking each of the V, D and J gene segments = recombination signal sequences
-Rearrangement involves a complex of enzymes = V(D)J recombinase
-(recombination activating gene)
-RAG-1 and RAG-2 genes encode lymphoid-specific components of the recombinase
-Mutations in RAG genes result in immunodeficiency
What is allelic exclusion?
-In each B cell, only 1 rearranged H chains gene from one chromosome is expressed.
-Similarly, only one rearranged L chain from one chromosome is expressed by each individual
-Light chain isotope exclusion - each B cell expresses either a rearranged kappa or lambda light chain - never both.
-These mechanisms ensure that each individual B cell produces one randomly generated antibody.
Mechanisms for generation of antibody diversity
- Multiple gene segments for each chain.
- Combinatorial diversity - different V,D, J segments recombine to produce different sequences
- Combinations of heavy and light chains
- Junctional diversity increases diversity further - imprecise joining, N regions
- Somatic hypermutation (SHM)
What is SHM?
-Performed by the enzyme, activation-induced deaminase (AID)
-AID acts on DNA to deaminate cytosine to uracil
-Uracil is then recognised by error-prone DNA repair pathways leading to mutations
What are the antibody classes respective heavy chains?
IgM - mu
IgD - delta
IgG - gamma
IgA - alpha
IgE - epsilon
Membrane (BCR) vs secreted antibody
-Following Ag recognition as each B cell differentiates, it will start to secrete its unique BCR as an antibody
-The secreted form made by each B cell has an alternative constant region that lacks a transmembrane region. As the original re-arranged VDJ regions are not altered, the secreted antibody has the same antigen specificity as the membrane BCR.
-The membrane and secreted forms are produced by alternative RNA processing
Describe heavy chain constant regions
-Encoded by a different C region gene segment (i.e. Cm, Cd, Cg, Ce , Ca)
-4 gamma chain gene segments correspond to the four IgG subclasses; similarly two a chain gene segments
-At the the heavy chain locus, the Cm segment is physically closest to the V, D and J gene segments and so IgM is the first antibody expressed by each developing B cell
What happens after B cells have assembled a random BCR?
-They leave the bone marrow, and their V region genes may undergo SHM in specialized areas of secondary lymph organs following the binding of Ag. and the activation of the B cell
-This leads to further BCR diversity being generated
-These activated B cells start to secrete their BCR (as an antibody)
-Class-switch recombination may then occur, as a result of the cytokines that may be present in this environment
Generation of diversity of TCR
Similar to that seen in BCR:
-multiple V,D,J gene segments
-combinatorial diversity
-junctional diversity
Unlike BCR, TCR is never secreted and no SHM occurs in TCR genes
Why are there such high levels of MHC polymorphism?
-Allows the binding of many peptides that can be presented to T cells - provides a clear evolutionary advantage to the population as can respond to almost unlimited no. of different pathogens
However,
-Increases risk of immune-mediated disease
-Reduces pool of available donor organs
Summarise MHC Class 1 and 2
-Highly polymorphic (diverse at population level)
-Individual has a limited number of different forms
-Influence which peptides can be presented
Ag processing and presentation by MHC class 1 molecules
1) Ag. (e.g. viral protein) synthesised in cytoplasm
2) protein cleaved to peptides by proteasome
3) peptides transported to endoplasmic reticulum by TAP transporter
4) peptides bind to MHC class I molecules
5) MHC-I/peptide complex then transported to cell surface
Ag processing and presentation by MHC class 2 molecules
1) Ag. (e.g. bacteria) endocytosed into intracellular vesicles inside the cell
2) protein cleaved to peptides by acid proteases in vesicles
3) vesicles fuse with vesicles containing MHC class II molecules
4) peptides bind MHC class II molecules
5) MHC-II/peptide complex then transported inside vesicles to cell surface
What do MHC class 2 molecules bind to?
Invariant chain in ER - prevents peptides binding in the groove
What is the class 1 pathway?
TAP and LMP
What is the class 2 pathway?
HLA-DM
What are APCs
Antigen Presenting Cells
-Take up and present extracellular Ag to activate helper CD4+ T cells
-Often macrophages, B cells, dendritic cells
What does the CLIP protein do?
Temporarily occupy the peptide-binding site of MHC class II molecules until it is replaced by a more suitable peptide
What is the role of the invariant chain?
- Chaperone for MHC class 2 molecules
- Stabilization of MHC class II structure
- Targeting MHC class II molecules to endosomal compartments
- Generation of CLIP
- Facilitation of peptide loading onto MHC class II molecules
B cell development
- Develop from haematopoietic stem cells in bone marrow that express PAX5 transcription factor
-Continuous process
-Involves re-arrangement and expression of Ig genes
-Expression of lymphocyte, and then B cell-specific, markers e.g. CD45, then CD19
-Removal of self-reactive cells
Describe transition of pre-B cells to immature B cells
Pre B cells
-H chain genes rearrange first (mu chain), moves to cell surface with Iga and IgB and expressed with surrogate light chain -> pre-B cell receptor
-Then light chains rearrange, and displace V preB -> IgM BCR
Immature B cells
Organisms that may cause disease
Bacteria
Viruses
Fungi
Parasites - worms, protozoa
Why are different effector mechanisms needed for different infections
Dependent on:
-Type of pathogen
-Localisation
-Challenge
-Stage of infection
Characteristics of innate defence mechanisms
-Rapid
-Act early
-Non-specific
-Ineffective against many pathogens
Types of T helper cells and what they are active against.
TH1 - active against intracellular pathogens, activate macrophages and stimulate cytotoxic T cells
TH2 - active against extracellular pathogens, support antibody production
TH17 - active against extracellular bacteria and fungi, important in attracting inflammatory cells such as neutrophils, induced early in infection
Name some gram positive bacteria
Staphylococcus aureus, Streptococcus spp.
Thick layer of peptidoglycan not present in gram negative cell wall.
Name some gram negative bacteria
Gram negative e.g. Campylobacter, Salmonella, Shigella
How can components of bacterial cell walls induce innate responses?
-Bind to Toll-like receptors (TLR) on macrophages
-10 TLR genes in humans: receptors recognise distinct molecular patterns on microbes
-NOD-like receptors (nucleotide binding oligomerization domain (intracellular sensors) in cytoplasm
What can binding of PAMPS to TLR do?
-Promote inflammation
-Promote dendritic cell maturation
-Influence differentiation of T cells
-Activate B cells
What is the role of antibodies in bacterial infection?
-Opsonisation
-Complement activation
-Bind to and neutralise toxins
-Bind to surface structures to prevent mucosal adherence
How can gram negative bacteria be killed?
Complement lysis
Advantages of activated macrophages
Better at phagocytosis and killing
More efficient APCs
Stimulate inflammation
2 types of leprosy
- Tuberculoid leprosy - strong TH1 response, few live bacteria, slow progression, granuloma formation
- Lepromatous leprosy - strong TH2 and antibody response, large no. of bacteria in macrophages, disseminated infection, fatal
Difference between antibodies and T cell effector mechanisms
Antibodies - protect against extracellular pathogens
T cell effector mechanisms - protect against intracellular organisms
Innate defences against viruses
Interferons, natural killer cells
What are natural killer cells
-Type of innate lymphoid cells
-Large granular lymphocytes
-Recognise structures on viral infected cells
-Can recognise stressed cells in absence of Igs and MHC
-Kill by extracellular mechanism- perforin and granzyme
-Rapid
Therapeutic use of interferons
-rIFNα can be used to treat hepatitis B and C
-Some cancers
-Side effects- can be very severe
What are natural killer cells?
-Innate lymphoid cells
-Large granular lymphocytes
-Recognise structures on viral infected cells
-Recognise stressed cells in absence of Igs and MHC
-Kill by extracellular mechanism
-Fast
NK cell receptors
Needs to distinguish between infected and uninfected host cells otherwise would a disaster
- Activating receptors: recognise carbohydrate ligands, triggers killing
2). Inhibitory receptors: recognises MHC class I molecules (no binding, only TCRs can do this)
Killing by cytotoxic T cells
- Secretion of cytotoxic granules
- Fas ligand on T cell interacts with Fas on target -> death
Both induce apoptosis
How does HIV act?
-Attacks specific immune system
-Targets CD4 T cell, macrophages and dendritic cells
-Progressive development of AIDS leads to opportunistic infections
Evasion mechanisms
1) Concealment of antigens
2 Antigenic variation
3) Immunosuppression
4) Interference with effector mechanisms
How is antigenic variation caused?
- Large number of antigenic types
- Mutation
- Recombination
- Gene switching
What is immunosuppression
- Infection of immune cells
- Induction of regulatory T cells
What are regulatory T cells?
-Type of CD4 cell
-Regulate immune system, suppress differentiation and proliferation of TH1 and TH2 cells
-Maintain tolerance to self-antigens, help prevent autoimmune disease
Examples of interference with effector mechanisms
- Molecules interfering with antibody function
- Molecules interfering with complement
- Molecules binding cytokines
- Subvert responses by producing molecules with cytokine activity
- Inhibition of phagocytic killing
Passive immunity characteristics
Short lived (half life of IgG ~ 3 weeks)
Active immunity characteristics
-Exploits ‘immunological memory’
Requirements of an effective vaccine
Safe
High level of protection
Long-lasting protection
Right type of response
Low cost
Stable
Easy to administer
Minimal side-effects
Types of vaccines
-Inactivated (dead)
-Attenuated (live but disabled)
-Subunit - use isolated antigens
-Toxoid
-Conjugate
Pros and cons of live vaccines
Pros:
Single dose effective
May be given natural route
May induce local and systemic immunity
May induce right type of response
Cons:
Reversion to virulence
Possibility of contamination
Susceptible to inactivation
Causes disease in immunocompromised host
What is reverse vaccinology?
Whole genome screening to identify proteins that could be used as vaccines
What is an adjuvant?
A substance administered with an antigen to promote the immune response
Adjuvants enhance immune responses:
-by providing a “depot”
-by immunostimulatory properties
How do adjuvants act?
-Activate dendritic cells via TLRs or NLRs
-Cause release of endogenous danger signals
-Promote antigen uptake by dendritic cells
-Stimulate release of chemokines/cytokines
-Promote ‘cross-presentation’ of exogenous antigens by class I
