Lecture 30 - B cells. Flashcards
How do ~20,000–25,000 protein-coding genes result in a wide variety of antibodies and TCRs?
Through primary diversity, which is achieved by somatic recombination, creating combinatorial and junctional diversity in antibodies and TCRs.
What are the two types of diversity that contribute to primary diversity in antibodies and TCRs?
Combinatorial diversity and junctional diversity.
What is somatic recombination in the context of antibody production?
Somatic recombination is the process by which gene segments are rearranged to create different antibodies. It is similar to shuffling a deck of cards and dealing out different hands.
What is the analogy used to explain somatic recombination?
Shuffling a deck of cards and dealing out different hands.
What types of gene segments are involved in light chain and heavy chain recombination?
- Light chain: Variable (V) and Joining (J) segments.
- Heavy chain: Variable (V), Diversity (D), Joining (J), and Constant (C) segments.
Which gene segments are involved in the recombination of the light chain and heavy chain?
- Light chain: V and J segments.
- Heavy chain: V, D, J, and C segments.
What is unique about the D segment in antibody recombination?
The D segment is only present in the heavy chain.
What is CDR1 and CDR2, and where are they encoded?
CDR1 and CDR2 are regions of the antibody’s variable region and are encoded in the V segments of both light and heavy chains.
Where is CDR3 encoded and why is it important?
CDR3 is encoded in the joining of the V-J segments (light chain) and V-D-J segments (heavy chain). It is the most variable CDR and contributes to the diversity of antibodies and TCRs.
What is the structure of the variable region in both light and heavy chains?
The variable region consists of recombined V, D (only in heavy chain), and J gene segments.
How does V(D)J recombination contribute to antibody diversity?
V(D)J recombination randomly selects one segment from each V, D, and J gene pool to create a unique variable region in antibodies.
What is the role of the proteins involved in somatic recombination?
The recombination process is tightly regulated by machinery involving many proteins that also participate in DNA repair functions.
What is the process of choosing V, D, and J gene segments during B cell development?
During B cell development, V(D)J recombination occurs, selecting one segment from each of the V, D, and J gene pools to form the variable region of the antibody.
How many different loci are involved in light chain production, and what are they?
There are two different loci in the light chain: the κ (kappa) chain locus and the λ (lambda) chain locus.
What happens to the κ and λ loci in light chain production?
Only one chain (either κ or λ) will be expressed, and the other chain will be silenced.
What does each light chain locus contain?
Each locus contains many different Variable (V) and Joining (J) regions.
How many loci are involved in heavy chain production?
There is one locus involved in heavy chain production.
What gene segments are involved in the heavy chain recombination?
The heavy chain locus contains many different V (Variable), D (Diversity), and J (Joining) regions.
How do constant regions in the heavy chain relate to different antibody isotypes?
Different constant regions represent the different isotypes (e.g., IgM, IgD, IgG, etc.) of antibodies.
What is the source of the V, D, and J regions in heavy chain recombination?
The V, D, and J regions are inherited from the stem cell (precursor B cell).
What is the process called by which recombinase proteins join gene segments in antibody production?
The process is called somatic recombination.
What happens to the B cell receptor (BCR) in a naïve B cell?
The BCR of a naïve B cell is formed during somatic recombination, and this process is irreversible.
How many V regions are available to choose from for the heavy chain?
There are 46 V regions to choose from (e.g., VH1, VH2, VH3, etc.).
How many D regions are available to choose from for the heavy chain?
There are 23 D regions to choose from (e.g., DH1, DH2, DH3, etc.).
How many J regions are available to choose from for the heavy chain?
There are 6 J regions to choose from (e.g., JH1, JH2, JH3, etc.).
How would you create your own VH region?
To create a VH region, choose one V, one D, and one J region, and combine them as a single word: V#D#J# (e.g., VH1DH2JH3).
What is combinatorial diversity in antibody production?
Combinatorial diversity is the diversity resulting from the different combinations of V, D, and J gene segments.
What are recombination signal sequences (RSSs)
RSSs are sequences that flank each antibody gene segment and direct recombination by guiding the pairing of segments.
What is the structure of a recombination signal sequence (RSS)?
An RSS consists of a conserved nonamer (9 bp) and heptamer (7 bp), with either a 12-bp or 23-bp spacer sequence between them.
What is the “12/23 Rule” in recombination?
The “12/23 Rule” dictates that a 12-bp RSS must pair with a 23-bp RSS for recombination to occur.
How does recombination occur in the light chain and heavy chain?
- Light chain: V-J recombination.
- Heavy chain: D-J recombination followed by V-DJ recombination.
What happens to the segments that are not selected during recombination?
The segments between the selected ones are excised and form a loop in the DNA, which is then deleted.
How does DNA looping occur during recombination?
RSSs bring the regions together, creating a loop in the DNA, with the excised non-selected segments forming a signal joint.
What proteins are involved in the recombination process, and what is their function?
The RAG-1 and RAG-2 proteins are necessary for recombination, responsible for recognizing and cutting DNA at the immunoglobulin-encoding region and the RSS.
What happens to the DNA after recombination?
The DNA forms covalently closed hairpin ends at the excised loop, and the loop is deleted from the chromosome. The coding region of the selected V and J regions remains and forms the coding joint.
What is a signal joint?
The signal joint is the excised DNA loop that is deleted after recombination, no longer present on the chromosome.
What is the role of RAG proteins in recombination?
RAG-1 and RAG-2 are responsible for cutting the DNA at the immunoglobulin-encoding region and the RSS to facilitate recombination.
What is junctional diversity in antibody production?
Junctional diversity is the variation that occurs when nucleotides are added or removed at the junctions between V & DJ, D & J, or V & J gene segments during recombination.
What happens to the signal joint after recombination?
The signal joint is ligated together and discarded.
What happens at the coding ends during recombination?
Repair proteins bind to the hairpin, and Artemis, an endonuclease, opens the DNA hairpins.
How does Artemis contribute to recombination?
Artemis opens the DNA hairpins and can cleave them in three different ways, facilitating junctional diversity.
What are palindromic (P) nucleotides, and how do they contribute to recombination?
P nucleotides are added at overhangs during hairpin cleavage. They are complementary nucleotides that allow DNA repair enzymes to fill in the opposite strand.
How does exonuclease activity contribute to junctional diversity?
Exonuclease activity removes nucleotides on each side of the coding joint, further contributing to junctional diversity.
What is the role of Terminal Deoxynucleotidyl Transferase (TdT) in recombination?
TdT adds up to 20 nontemplate-encoded nucleotides (N nucleotides) to the cleaved strands, primarily in the heavy chain, contributing to the variability of CDRs (Complementarity Determining Regions).
How do repair enzymes contribute to recombination?
Repair enzymes trim off nonmatching nucleotides, fill in single-stranded gaps, and ligate the new DNA.
What are the main mechanisms that generate B cell receptor (BCR) diversity in naïve B cells?
The mechanisms include:
1. Multiple gene segment combinations
2. Heavy chain/light chain combinatorial diversity
3. P nucleotide addition
4. Exonuclease trimming
5. Nontemplated N nucleotide addition by TdT
What is the role of TdT in the heavy chain recombination?
TdT adds random N nucleotides between joints in the heavy chain, contributing to junctional diversity.
What is the impact of exonuclease activity on junctions in recombination?
Exonuclease activity can remove nucleotides at the junctions between gene segments, leading to further diversity in the final product.
What is the general outcome of the V(D)J recombination process?
The process results in a diverse range of BCRs due to the combinatorial diversity, junctional diversity, and random addition or removal of nucleotides at the junctions.
