L12 – Humoral Immune Responses Flashcards
List the 5 Ab isotypes? Which are membrane bound? Which are secretory?
IgG, A, M, E, D
A = secretory and mucosal defense M,D = membrane bound
Compare secondary humoral response time, Ab titre and affinity, Ab class to primary.
Secondary:
- Shorter lag phase with exponential production of Ab from Ag-specific memory B cells
- Mainly IgG, little IgM
- Much higher Ab affinity to Ag
Describe the primary humoral response? (cell type, Ig type, time)
- long lag phase (~7-10 days), persistence depends on nature of Ag
- antigen-specific naïve B cells undergo clonal selection and differentiation into plasma cells and memory cells
- IgM first**, then IgG
Describe the B-cell receptor composition.
- Ig heavy and light chains
* Ig α/β chain for intracellular signal transduction
Summarize the intracellular signalling events to activate transcription after BCR activation by Ag? ***
- Antigens (Ag) crosslink B cell receptors (Ig) cluster on B cell surface
- Phosphorylate immunoreceptor tyrosine-based activation motifs (ITAM)**
- Recruit and phosphorylate tyrosine kinases** (especially LYN, SYK)»_space; initiate intracellular signaling
- Activate multiple pathways at the same time:
(i) Burton’s tyrosine kinase (BTK)
(ii) Phospholipase Cγ (PLCγ) , increase intracellular [Ca2+]
(iii) Phosphatidyl inositol-3-kinases (PI3K)/AKT - Activate transcription factors
List the gene transcriptions that occur after BCR signalling? *****
Activate genes for:
1) Cell proliferation (e.g. c-fos, cmyc)
2) Receptors for cytokines ** (e.g. IL4R, IL-6R, TGFBR, IFNGR)
3) Surface molecules e.g. MHC class II ** for Th cells
4) Specific enzymes: e.g. Activation induced deaminase (AID) – for somatic hypermutation in Ig
Describe how B cells present antigens after BCR activation/ crosslinking?
BCR crosslinking
> > Endocytosis BCR- Ag complex and break down
> > Use MHC - II from activated gene transcription to load Ag
> > Present MHC-II with Ag on surface to interact with Th
Classify Ag by their downstream reaction after binding to BCR?
Thymus dependent Ag»_space; Ag require contact help from T helper cell for optimal Ab response
Thymus- independent Ag»_space; no interaction with Th
What are the 2 ways in which T cells can induce B cell proliferation and class switching?
Cell-cell contact through CD40 and CD40L binding between Th and B cells
T cells secrete cytokines for Ig class switching and survival
Describe the cell-cell contact between Th and B cells and its purpose?
critical for Ig isotype class-switching to IgA, IgG, IgE
Signal 1: antigen binding to BCR increases the expression of MHC-II, CD40 (both constitutively expressed)»_space; IgM pentamer made immediately
Signal 2: MHC-II loaded with Ag bind to TCR on Th»_space;
upregulates CD40L on activated Th
Signal 3: CD40L on Th bind to CD40 on B cell»_space; induce Ig class switching
What disease is caused by CD40L deficiency in Th cells?
X-linked hyper IgM syndrome: no CD40/CD40L interaction = no Ig class switching
who also have a defect in macrophage function, immunodeficiency, more infection susceptibility
Describe the interaction between Thymus-independent Ag and B cells? (think TI-2 and TI-1 and their respective functions)
- antibody response without the need of Th cell contact
1. TI-1 Ag binds to BCR and other receptors (i.e. Toll- like receptors) »_space; Polyclonal B cell activation regardless of antigen specificity**
2. TI-2 Ag causes extensive Ig crosslinking on B cell»_space; Plasma cell with weaker Ab response (IgM mainly), No memory cells
Compare the source of TI-1 and TI-2 Ag?
TI-1 antigens (e.g. lipopolysaccharides (LPS)*** on Gramnegative bacterial cell wall components) = mitogens
TI-2 antigens (carbohydrates/polysaccharide*** with repeating subunits, e.g. pneumococcal polysaccharides, Salmonella polymerized flagellin)
Location of B cell activation and differentiation?
Germinal centers of lymphoid follicles of secondary lymphoid organs
List 2 processes that occurs in the dark zone of germinal centers in lymphoid follicles?
- B cell proliferation (clonal expansion)»_space; become centroblasts
- Somatic hypermutation in Ig gene»_space; change affinity to Ag
Describe the processes that occur in Class switch recombination (CSR)** in immunoglobulin gene to induce Ig class switching? (think about the enzymes involved)
Before switching, alternative splicing makes IgM, IgD
involves several enzymes:
1) Activation-induced deaminase (AID)*****: cytosine»_space; uracil
2) UNG: excises uracil from DNA
3) APE: nicks phosphodiester backbone in DNA
> > free ends of the DNA are rejoined by NHEJ
@ C SEGMENT GENES!!! NOT HYPERVARIABLE REGIONS IN SOMATIC HYPERMUTATION
Which Ig isotypes are made before and after class switching? Which cells make these isotypes?
- IgM or IgD are transcribed before switching by ALTERNATIVE SPLICING in naïve B cells
- IgM switched to other classes e.g. IgG, IgA or IgE by C GENE REARRANGEMENT in activated B cells and plasma cells
Which gene segment is involved in somatic hypermutation of Ig gene?
Hotspots of SHM = complementarity determining regions (CDRs)/ hypervariable region of V domains of H and L chains
= the antigen binding site
List the 4 processes that occur in the light zone of the germinal centers in secondary lymphoid organs?
1) Centroblasts»_space; centrocytes
2) Ig affinity maturation
3) Ig class switching
4) Differentiation of plasma cells and memory cells
Describe the maturation process that selects B cells with the correct Ig against the specific Ag?
Affinity maturation:
- Somatic hypermutation rearrange HV gene segment without changing overall specificity
- B cells making Ig with the highest Ag specificity are selected, the rest die
List the 2 cells responsible for B cell development and maturation
T cells (follicular T helper cells)
Follicular Dendritic cells
Describe how Follicular T helper cells help B cells maturation?
Cell-cell contact via CD40-CD40L»_space; Ig class switching
Make cytokines e.g. IL-4/IL-5/IL-13»_space; Critical of B cell survival and Ig class switching
Describe how Follicular dendritic cells help B cells maturation?
- Compete with B cells for antigen presentation
- Outcompete B cells with low affinity Ig
> > only B cells with high affinity Ig survive by expression of survival protein BCL-2»_space; Plasma and memory cells
Describe how antigen re-exposure causes increased humoral response and antigen specificity. (think about mutations in Fab)
- IgM made in primary response has little mutation in Fab, no effect on Ag binding
- IgG in late primary response has increased mutation in hypervariable region of Fab > increase binding affinity (low Kd)
- IgG in secondary/ tertiary response has increased somatic hypermutation in Fab»_space; much higher binding affinity
Isotype class switching affects Ig affinity or specificity. T or F?
False
Does not affect Ig affinity or specificity but biological effector activities varies
How do T cell cytokines contribute to Ig class switching?
Ig class switching involves CYTOKINE-DEPENDENT TRANSCRIPTION in the rearranged constant region
e.g. IL-4 promotes IgE, TGFβ promotes IgA, interferon-gamma promotes IgG1 and IgG3
Describe the processes in somatic hypermutation in Fab? (Not Class Switch Recombination)
1) Involves activation-induced (cytidine) deaminase (AID): during cell replication: cytosine»_space; uracil (= error in DNA)
2) Uracil removed
3) Replaced by another nucleotide by **error-prone DNA polymerase*
> > introduce high rate of random mutations in DNA
Location of memory B cell formation and which Ag stimulates diff. of B cells?
Germinal centres, light zones of lymphoid follicles
T-dependent antigens
Describe the isotypes of Ab made from memory B cells?
Mostly isotype switched (e.g. IgG+ , IgE+ or IgA+)
Higher affinity for the inducing antigens
What immune cells offer long term protection or immune responses against specific antigens
Memory B cells
Compare the C segment genes that make the different isotypes of Ig?
IgG (Cγ) : IgG1, IgG2, IgG3 and IgG4 IgM (Cμ) IgD (Cδ) IgA1 (Cα) IgE (Cε)
Which Ig is the Major Ab for secondary responses?
IgG
4 Functions of IgG?
- Major Ig for secondary response
- Neonatal immunity
- Complement activation
- Opsonization and sensitization of NK cells via FcγR
Which Ig is the major one for primary responses?
IgM
Which type of Ag induces the most IgM.
TI-2 antigens (crosslinking, carbohydrate based)
Binding affinity, diffusion capacity and function of IgM?
- Lowest binding affinity
- Cannot diffuse across vascular wall
- Potent agonist of complement system
Proposed functions of IgD?
- maintain the quiescent state of autoreactive B cells
- secreted IgD appears to enhance mucosal homeostasis
2 functions of IgE? how is it transported in blood?
Found on surface of mast cells, basophils and eosinophils (express high affinity Fc receptor for IgE, FcεRI)
Helminthic parasite
hypersensitivity reactions
Isoforms of IgA and function?
monomers and dimers connected by a J-chain
Secretory dimer»_space; trans-epithelial transport»_space; mucosal immunity
e.g. saliva, milk, colostrums, urogenital and tracheobronchial secretions
