Lecture 10: Lymphocyte Development and Antigen Receptor Gene Rearrangement Flashcards
Hematopoietic stem cells give rise to ___ which give rise to
Common lymphoid progenitors CLPs which give rise to B cells, T cells and NK cells
What happens when Pre-Ag receptor is successfully rearranged
It provides survival signals that select the cell
What drives the proliferation of human T cell progenitors and where is it produced
IL-7 produced in the thymus
What other function does IL-7 have besides proliferation of T cells
When produced by stromal cells in bone arrow, it promotes B cell development
What drives development of NK cells and where
IL-15 in the thymus
First step in developing B cells
The Ig heavy chain locus opens up and becomes accessible to proteins that will mediate Ig gene rearrangement and expression
First step in developing a/b T cells
The TCR beta gene locus opens up and becomes accessible for TCR gene rearrangement and expression
Which transcription factors commit cells to the T cell lineage
Notch-1 and GATA-3
Function of Notch protein
Notch is cleaved and intracellular portion travels to nucleus to modulate expression of target genes
GATA3 also helps induce this expression
Which proteins regulate TCR/BCR rearrangement
Rag-1 and Rag-2
Which genes undergo V(D)J recombination
Those which encode the components of pre-TCR
Which TFs induce expression of genes for B cell development
EBF, E2A and Pax-5
DNA methylation on cytosine results in
Gene silencing
Allelic exclusion refers to
The restriction that only one of the light chain and heavy chain alleles (maternal or paternal) is expressed in a single B cells
Variable regions of the chains in T and B cells are determined by
Rearrangement of the DNA
What is the major mechanism of epitope-specific diversity of BCR/TCRs
DNA chromosomal rearrangement
The process of rearrangement includes
Deletions of DNA/RNA nucleotides and reannealing gene segments.
This is done by Rag1 and Rag2 recombination enzymes and is called V(D)J recombination
RAG-mediated DNA breaks are repaired exclusively by
Non homologous end joining
Three mechanisms of rearranging gene segments
Somatic recombination
mRNA splicing
Junctional diversity
How does each B cell generate unique combinations of V(D)J segments
In a single B cell all copies but one each of VDJ are randomly deleted
Recombination starts with which chain in B and T cells, respectively, and then moves to which chain
Starts with heavy chain in B cells
Starts with Beta chain in T cells
If functional, rearrangement of light chain (B cells) or alpha-chain (T cells) occurs
Achieving BCR diversity heavy chain step one
First, D and J are chosen and DNA between them is deleted
Achieving BCR diversity heavy chain step two
Second, a V segment is chosen and DNA between V and DJ is deleted
Achieving BCR diversity heavy chain step three
Third, a C is chosen and DNA between VDJ and C is deleted
What is the chance % of producing a productive rearrangement (w/o stop codons in sequence)
10%
H-chain chromosome
K-chain chromosome
Lambda-chain chromosome
H-14
K-2
L-22
THE VDJ rearrangement segment facilitates the synthesis of _________, and is controlled by ______
Facilitates synthesis of a Mu or delta heavy chain, controlled by alternative mRNA splicing
This then associates with lightchain forming IgM or IgD molecule
Non-IgM/IgD heavy chains are produced by ______, which is a process that
Class-switch recombination (CSR), a process that exchanges the constant region of the heavy chain
Class switch recombination (CSR) requires what enzyme that is only expressed at what time
AID- only expressed in activated B cells
AID method of operation
AID introduces uracil residues into DNA of S (switch) regions, inducing generation of DNA breaks at these regions followed by repair
Junctional diversity
TdT and/or RAG adds or removes nucleotides to the exposed ends of the V, (D) or J genes before they are reunited
Junctional diversity is helpful how
It increases the diversity of TCRs and BCRs
Junctional diversity is created where
At the points between joining genes
Junctional diversity results from
The loss of nucleotides through action of exonucleases and the addition of N and P nucleotides
RAG and TdT function in junctional diversity
RAG cleaves hairpin loops and adds P nucleotides (P nucleotides form from asymmetric opening of hairpin)
TdT adds N nucleotides
Checkpoint #1 of the selection process
After the production of the first polypeptide chain of the two-chain Ag receptor is completed
Checkpoint #2 of the selection process
Follows the production of the second polypeptide chain of the Ag receptor
Pre-Ag receptor structure
Pre-Ag receptors contain only one polypeptide chain present in mature Ag receptor and a surrogate receptor chain
Pre-BCRs contain
Ig(Mu) heavy chain
Pre-TCRs contain
TCR Beta chain
What is the first Ag receptor gene to be completely rearranged in B cells
Ig heavy chain
Developing B cells that successfully rearrange their Ig heavy chain genes then express
The Mu heavy chain protein and assemble the pre-BCR
Developing T cells that make productive TCR B chain gene rearrangement synthesize
The TCR B chain protein and assemble the pre-TCR
What percent of B/T cells make a productive in-frame rearrangement
30%
Assembled pre-BCRs/TCRs provide signals for
Survival, proliferation and further development of early B/T cell lineages
Negative/positive selection is part of what checkpoint
Checkpoint 2
If cells make productive rearrangement of second chain, they express
Complete Ag receptor while they are still immature. These cells will be positively selected
What is important for maintaining the central tolerance to many self Ags
Negative selection
When does negative selection occur
Shortly after Ag receptors are first expressed on developing B/T cells
Negative selection effect on T cells vs B cells
It will eliminate harmful T cells but ALTER harmful b cells whose Ag receptors bind strongly to self Ags in thymus/bone marrow
Clonal deletion
Process of eliminating harmful T cells
Receptor editing
Second attempt at Ig gene rearrangement, if this fails, B cells will enter clonal deletion
What chain will be altered first in a B cell that binds self Ags. What chain will be altered second?
Kappa Light chain will be rearranged first
Lambda light chain is rearranged second, if needed
B cells developing from fetal liver-derived stem cells differentiate into
B-1 lineage
B cells developing from bone marrow precursors give rise to
B-2 lineage
B-1 cell receptor diversity
Limited BCR diversity because TdT is not expressed in the fetal liver
Without TdT- no junctional diversity
B-1 cells are found where and secrete what
Found in peritoneum and mucosal sites
Spontaneously secrete IgM that react with polysaccharides/lipids/oxidized lipids
B-1 B cell involvement in early phases of infection
Contribute most of the serum IgM during early phases
Immature B-2 B cells relocate where after what
After rearrangement of BCR chain genes, they relocate to spleen
B-2 cells can differentiate into
Marginal zone MZ B cells or Mature follicular FO B-2 cells
FO B-2 cells are
Recirculating lymphocytes
MZ B-2 cells are abundant where but also found where
Abundant in spleen (in marginal sinus), also found in lymph nodes
MZ B cells respond to
Blood-borne pathogens
Responses of MZ B cells are independent or dependent on T cell help
Independent of T cell help (usually) but can mediate SOME T cell dependent immune responses
They are self renewing
FO B-2 cells may develop into plasma/memory cells depending upon
Mature FO B-2 cells depend upon T cell activation to mature into plasma/memory cells
True or False - MZ B cells undergo immunoglobulin isotype switching and affinity maturation
Probably false? FO B-2 cells are the ones that do this because they respond to protein Ags in T cell dependent manner
MZ B cell diversity
Similar to B-1 cells, they have limited diversity which respond to polysaccharide Ags and generate natural Abs
MZ B cells respond very rapidly to ____ and differentiate into what
Respond rapidly to blood borne microbes and differentiate into short-lived IgM secreting plasma cells
What causes the limited diversity of expressed gamma/delta TCRs
Only a few of the available V, D and J segments are used in mature yd T cells
What determines T cell being yd or ab
If it first succeeds in rearranging its TCR y or delta loci before it makes a productive TCR beta rearrangement, it is selected to the yd T lineage
-happens about 10% of time
