Lecture 3 - B cell specificity Flashcards
What are the 3 properties of Ab?
Specific, induced, responses which are rarely generated against self antigens
Describe the side chain hypothesis proposed by Erlich and why it was ultimately incorrect
A single antigen binding to its specific receptor triggers Ab release while keeping other specificities off. Incorrect because it couldn’t explain how a single B cell could secrete only the Ab needed if every cell had the capacity to respond to every antigen and also the cell wouldn’t know not to generate anti0self Abs
Describe the clonal selection hypothesis proposed by Burnett and it’s 4 tenets
A given lymphocyte is monospecific, binding of antigen-specific receptor activates the cell (receptor engagement), triggering specific/unique Ab release (receptor=product), and each cell/daughter cell retains specificity over the course of an immune response (clonal integrity)
How many daughter cells can a responding lymphocyte give rise to?
5K to 50K
What do activated B cells eventually give rise to?
Ab secreting plasma cells and memory B cells
What do activated T cells eventually give rise to?
Cytokine producing cells that regulate other immune cells (helper T cells) and killer cells that lyse infected host cells (cytotoxic T cells)
Are responses by lymphocytes to foreign antigens monoclonal?
No, since most antigens possess many epitopes the Ab/immune response is usually polyclonal, different clones each with a unique VL-VH pair will respond to any given antigen
How does clonal selection explain Ag specific memory responses?
After the first exposure the lymphocyte repetoire includes a greater number of cells specific for the origina antigen so increased numbers of Ag-specific cells are available to respond against the antigen the second time and therefore more Ab secreting cells and more Ab is generated the second time
How does clonal selection explain the Ag specific Ab graph from the first lecture?
During primary response Ag specific cells generate large numbers of clonally related daughter cells then when Ag is introduced again the increased numbers of Ag-specicic cells available results in much more and faster Ab secretion
How are monoclonal Ab’s generated for use as drugs and diagnostic tests?
Spleen cells collected from mouse challenged with Ag are fused with myeloma cells (plasma cancer cells) forming B cell hybridomas then cells are selected for specific Ab from which you can harvest monoclonal Ab (can change mouse constant regions to human)
What did Karl Landsteiner’s immunization experiments with novel haptens tell us about the immune response?
Since as long as these haptens were attached to larger protein molecules the rabbits always generated hapten-specific Ab, the immune system is abole to respond to virtually any structure
Why do immunologists use inbred mice to investigate immune system development and function?
Because inbred mice are homozygous at every loci. Reduces confounding
How can you generate allele specific Ab’s to proteins encoded by Ig heavy chain genes?
By immunizing a strain of mouse with the IgH from a different strain (and vice versa)
What is the germ-line model of Ab diversity?
There are millions of Ig H genes and millions of Ig L genes with a unique H chain gene that corresponds to every unique H chain protein and the same for L chains. These millions of genes would be inherited via the germ line from parents to their offspring
What does clonal diversity predict about different clones of IgM + B cells?
Will have identical Ig heavy chain constant regions but different variable region sequences for H and L chains. Also a given B cell clone will be all kappa or all lambda
When would the process required for generating antigen receptors on B cells take place?
During pre-B cell/B cell pre-cursor phase during which the pre-B cell would turn on an Ig heavy chain gene and an Ig light chain gene. This process couldn’t be influenced by an anatigen because these receptors are needed to recognize the antigen.
How does the germ line model of Ab diversity fail?
Doesn’t explain how so many Ab with unique specificities could use identical constant regions (e.g. all Ab responses begin by secreting IgM Ab). Also can’t explain how Ab with identical specificity might switch constant regions (eg. Startinga s IgM but becoming IgG)
What is the somatic model of Ab diversity?
All cells have only one Ig H, one Ig kappa and one Ig lambda gene. These are all mutated intentionally in pre-B cells, spearking unique mutation patterns across Ig genes in each pre-B cell so that every precursor would have the potential for unique specificity. Wouldn’t be passed on genetically.
What did Tonegawa’s southern blot experiment show?
The genes enncoding Ig lamda light chains had moved in a lambda+ B cell (but not in a kappa+ B cell)
What is V(D)J recombination?
The variable region domain of an antibody molecule occurds due to the recombination and juxtaposition of 2 (for L chains) or 3 (for heavy chains) genetic elements/segments
How does recombination differ in heavy versus light chains?
The Ig H chain has an additional diversity segment. D-J rearrangements occur before V-DJ rearrangemnts. Also L chain gene recombination occurs after H chain recombination
What are two other additional sources of diversity aside from V(D)J?
Junctional diversity (because joining events aren’t precise causing overhands that must be filled in with appropriate nucleotide) and terminal deoxyribonucleotide transferase (TdT) (adds additional nucleotides to N regions between P and DJ regions.
How is transcriptional expression of Ig genes related to V(D)J recombination?
V(D)J recomb brings promoter sequences close to the enhancer which promotes transcription of the rearranged V gene
Which genes play a role in repair and rejoining during V(D)J recombination?
Recombinase activating gene 1 and 2 (RAG1+ RAG2)
How does RAG1/2 know where to cut?
It recognizes recombination signal sequences (RSS) which are conserved heptamers and nonamers separated by 12 or 23 base pairs which are located adjacent to V, J and D exons and only cuts there to avoid cutting of other genes
What needs to happen for a pro-B cell to become a pre-B cell?
Successful rearrangement of an Ig heavy chain allele so mutations that prevent Ig H chain recombination prevent pre-B cell generation
How does a pro-B cell know to proliferate and differentiate into pre-B cells?
Heavy chain protein forms a receptor by binding to a surrogate light chain. This pre-B cell receptor signals pro-B cells to proliferate and differentiate into pre-B cells
How does pre-BCR expression cause immunodeficiency (eg. Bruton’s disease)?
Results in loss of B cell development since pro-B cells don’t differentiate into pre-B cells
How does allelic exlusion work to achieve B cell monospecificity?
Basically only 1 Hchain and 1 L hain allele can give rise to a producitve rearrangement. Productive H chain recombination leads to pre-BCR assembly which also signals RAG1/2 to downregulate allowing for early pre-B cell proliferation without recombination. During late pre-B cell stage, RAG1/2 reactivated so that light chain recomb can occur. After a productive light chain recomb the resullting BCR again signals termination of RAG1/2 expression
How does VDJ recombination relate to immunodeficiency?
Mutations in any required mediators of VDJ recomb will result in coombined (B and T) immunodeficiency e.g. Omen’s syndrome missense in RAG genes
How does VDJ recombination relate to acute lymphoblastic leukemia?
Leukemia derived from preB or pre-T cells will express TdT
How does VDJ recombination relate to the detection of malignant B cells?
Since most cancer cells are monoclonal, the detection of a dominant Ig rearrangement (which would usually be random) is diagnostic e.g. too many kappa or too many lambda light chains
How does VDJ recombination relate to translocations and lymphoid malignancies?
VDJ recomb (and class switching) produce dsDNA breaks that contribute to translocations which can lead to overexpression of a cellular growth or survival promoting gene e.g. cMyc
