Generation of Diversity- B cells Flashcards
Regions of a light/ heavy chains
- the constant regions are not next to the variable region in germline
- variable region at the site nearest to the constant consisted of a small gene segment called the J region
- the J region did not change its relative position in the genome in IgM and D producing B cells compared to the rest of the body but it did change in all other Ig producing B-cells- this implied a rearrangement of the DNA
- in heavy chain additional region, D region or diversity gene segment
- the remainder of the Variable region is coded for the V gene segment
- all at different chromosomal locations
Reasons where there must be gene rearrangment
- thousands of bases of DNA information to code for those amino acids
- billions of bases of information
- only 3 billion bases in the human genome
Gene rearrangement leads to many variable regions
- light chains have V and J regions that are randomly chosen
- heavy chains have V,D, and J regions that are also randomly chosen
- recombination is caused by RAG proteins and recombination signal sequences
- in addition you can combine light and heavy chains
- regions of true randomness exist as well
Number of combinations
- by random combination of these elements within each cluster, a large number of different V regions can be created (for example Kappa light chains have 40 V segments x 5 J segments making 200 possible Kappa light chains)
- heavy chains have 51 possible Vs x 27 possible Ds x 6 possible Js or 8262 possible heavy chains)
- as each combination is independent one can have over 200 light chains x 8262 heavy chains making up variable regions with kappa light chains
- over a million possible combinations
DNA splicing
- RAG 1 and RAG 2
- each immunoglobin gene segment has signal sequences: signal sequences determine which segments can be joined to each other
- joining is imprecise and also there is a special mechanism to increase diversity at the CDR3 site
- joining of V and J in light chains can involve 4 different nucleotide codons (group of 3 bases) to create an in frame gene sequence coding for variable light chain
- joining of D to J and V-D in Heavy Chain gene formation can involve multiple reading frames, as most Ds can be read in all three reading frames
Novel genetic sequences from junctional diversity
- P nucleotides are found in all joining junctions
- the hairpins were opened with RAG and RSSs and the palindromic P nucleotides are generated
- N nucleotides are added by TdT which add random nucleotides to V-D and D-J
- N regions are not found in light chains
- N and P nucleotides add a bit of randomness to splice site
Productive rearrangement/ unproductive rearrangement
-in the early pro-B cell there is H-chain gene rearrangement D-J rearrangements on both chromosomes
-in the late pro-B cell there is H chain gene rearrangement- try V-DJ rearrangement on first chromosome if it works move on if it doesn’t try second chromosome if it still doesn’t apoptosis ( you test with a surrogate light chain)
-in Pre-B cell there is L-chain gene rearrangement
first you rearrange K gene on first chromosome and if that doesnt work move on to the next chromosome if that doesn’t work do first lambda gene if that doesn’t work second lambda gene
Selective splicing of primary RNA transcripts to switch from IgM to IgD
- IgD constant region are expressed in mature/naive B cells but there is no class switching involving gene rearrangements
- IgD is formed by selective splicing of an RNA that is transcribed from both IgM and IgD constant regions
Class switching
- when class switching occurs except to IgD, a variable region is joined to a new constant region
- this is DNA splicing
- during class switching, the variable region undergoes an unusually high rate of mutation- those mutations that yield amino acids that improve binding to antigen are selected for
- the enzyme is AID, looping out for switch region recombination
- can’t go back to being IgM
- Order: IgM, IgD, IgG3, IgG1, IgA1, IgG2, IgG4, IgE, IgA2
Mutations take place during immune response
- somatic hypermutation
- takes place in the whole V domain
- single base changes that are random
- does not take place in the constant region
The T-cell Receptor
- the mechanism by which the T cell receptor is generated is similar to the mechanism of generation of immunoglobulin
- most mechanism of formation of the T cell receptor are the same as the B cell except the somatic hypermutation
Possible mutations
- Lack Rag: no T cells or B cells- combined immunodeficiency
- lack TdT- less diverse and less able to respond
