10. Lymphocyte development and Antigen receptor Gene rearrangement Flashcards
what do lymphocytes start as
stem cells
where do B cells begin
Bone marrow
what happens after stem cells
pro lymphocyte
made after pro-lymphocyte
pre lymphocyte
what does an pre-lymphocyte become
immature lymphocyte
where does an immature lymphocyte become a mature lymphocyte and what is different
peripheral lymphoid organ or tissue and become antigen dependent
what are the series of events that occur for B and T cell production
commitment, proliferation, rearrangement of Ag receptor genes, selection events, Differentiation
what give rise to B and T cells
pluripotent stem cells
what is the role of hematopoietic stem cells (HSC)
give rise to common lymphoid progenitor (CLP)
what do CLPs produce
B cells, T cells, and NK cells
what type of B cells can a Pro B cell from a CLP become
follicular, marginal zone, B1
what type of T cells can a pro t cell become
aB T and yDelta T
what TF make a pro B cell from CLP
EBF, E2A, Pax5
what are the steps for a Pro T to be made from CLP
T cell precursor from CLP then Notch 1 and GATA 3 make a Pro T
what stimulates the proliferation of commited B and T cells
cytokines
what is proliferation so important
need a large pool of progenitor cells to generate high diversity of mature lymphocytes
why is Ag receptor arrangement important
provides survival signals for the cell
what cytokine provides the proliferation of T human cell progenitors and where
IL 7 produced from stromal cells in the thymus
what cytokine leads to the proliferation of NK cells
IL 15
what cytokines lead to B cell development
unknown
what is the ultimate determining factor for B and T cell commitment
signals from Ag receptors
what do signals from cell surface receptors activate
TF that induce Gene expression and rearrangement of Ag receptor genes
what allows for gene expression and rearrangement in developing B cells
Ig Heavy chain locus opens and becomes accessible to proteins
what in Developing aB T cells allows for TCR gene expression and rearrangement
TCR gene locus opens
what type of proteins are Notch
cell surface molecules
how do Notch proteins work
proteolytically cleaved when interact with ligands of neighboring cells
what does the cleavage of Notch proteins lead to
intracellular portion migrates to nucleus to modulate expression of target genes
which cells does GATA 3 effect the gene expression of
aB T cells
what is the type of recombination that occurs for pre TCR genes
V (D) J
what genes are expressed for B cell development
Rag1 and 2 proteins, surrogate light chains, Iga and IgB signaling proteins
what do Rag 1 and 2 proteins do
regulate BCR rearrangement
what is the surrogate light chain for in B cell development
pre-B cell receptor
what are the epigenetic mechanism used for B and T cell development
DNA methylation, silence Non-coding RNAs, modify histone tells of nucleosomes
what is methylated to silence genes
cytosine residues
is modification of histone tails activate or inactivate genes
both
what is the goal of epigentics
make genes available or unavailable in chromatin
euchromatin
loosely packed chromatin for TC
silenced genes have chromatin in what state
heterochromatin
what types of modifications are done to histone tails
acetylation, methylation, ubiquitination
how are chromatin remodelled
protein remodeling complexes
how does RNA play a role in silencing gene expression
non-coding RNAs can silence it
main type of non-coding RNA
microRNA (miRNA)
how do miRNA work
control on a post transcriptional level by impairing translation or promote degradation of mRNA
size of miRNA
22 nts
which genes encode Ig
u H chain, k chain, upside down y chain
what are the alleles for heavy and light chains
VLCL and VHCH
how many alleles of heavy and light chains are expressed on B cells and what does this mean
only one of each by allelic exclusion
what does allelic exclusion do
governs the expression of TCR and B cells
what is the ratio of maternal to paternal allotype expression
equal for so many B and T cells
what are the germlines in stem cells
ig and TCR
how are variable regions of T and B cells determined
rearrangement of DNA
what do the V and C stand for in alleles of stem cells
variable and Constant
what is DNA chromosomal rearrangement the major mechanism for
epitope-specific diversity of BCR and TCR
3 mechanism of DNA rearrangement
Somatic recombination, mRNA splicing, junctional diversity
what do the 3 mechanisms of DNA rearrangement involve
deletion and reannealing of gene segments
what are the segments on BCR and TCR
variable, Diversity and Joining segments
what do VDJ stand for
variable, diversity and segments
what parts of BCR have diversity segment
Heavy chain
what part of TCR aB has diversity segment
Beta
which part of BCR does not have repertoire with junctional diversity
lambda
what determines which chromosome: maternal or paternal, is used
first for gene rearrangement
4 gene segments of heavy chain
V,D,J and Constant
what chromosome houses Heavy chain genes
14
how are all B cells different
randomly deletes all but one copy of VDJ segments so all are different
RAG=
random activating genes
what do RAG 1 and 2 do
encode enzymes for VDJ recombination of BCR and TCR
what is RAG 1 and 2 expression restricted to
B and T lymphocytes during developmental stages
Pre B cells are made up of what chains
u heavy and surrogate light chains
how are DNA segments present in a mature B cell
single copy of each segment
1st deletion for B cell diversity
D and J chosen with DNA between them deleted
2nd step in B cell deletion
V is chosen and DNA between it and DJ deleted
3rd step in deletion of B cell segments
C is chosen then DNA between it and VDJ is deleted
what is the chance of having a productive rearrangement
10%
what is a productive rearrangement
B cell segments without a stop codon
how are productive rearrangements confirmed
test with TC and TL
what happens once productive is confirmed
recombination of other segments stops
how are Ig M or IgD molecules made
Tc driven by promoter to produce u or delta heavy chain by alternative splicing
where are IgM and IgD first made
cell surface of naive B cells
what produces secondary isotypes
class-switch recombination
how do CSRs work
exchanges constant region of Heavy chain with dow stream constant region genes
what enzyme is required for Ig Class switching
Activation-induced cytidine deaminase
What does AID do
creates DNA breaks at switch (S) regions
where are S regions located
before Constant regions
what happens after AID cuts
DNA repair
what happens if VH rearrangement is productive
B cells proliferate and take care of VL
what are they type of light chains in BCR
k and lambda
which VL do not have D segments
k and lambda
how does recombination work for VL
same as VH with productive rearrangement test
each B cells has what type of chains
one VH and one VL
where is k chain located
2nd chromosome
where is lambda chain located
22nd chromosome
how many C regions will a heavy chain have
3
what are the segments in TCR a and B chains
V J and C and D
how many of each segment is in a chain of TCR
V has 70-80 copies, J has 61 copies
how many copies of segments in B chain of TCR are their
V has 52, J has 6 the 2 D the 7 J, 2 C
steps in TCR gene rearrangement
same with same enzyme as BCR
how many different TCR are their
3 x 10^6
how many different BCRs are their
10^6
what further increases the diversity of TCR and BCR besides VDJ recombination
Junctional diversity
what is the junctional diversity of TCRs and BCRs
10^16 and 10^11
where is junctional diversity generated
points between joining genes
what causes junctional diversity
loss of nucleotides by exonucleases and addition of N and P nucleotides
how are P nucleotides made
asymmetric opening of hairpin loops
what do self- complementarity of P nucleotides lead to
palindromic appearance
what do P nucleotides produce
openings of short single stranded extensions that can be incorporated into junctions and removed via exonuclease activity
How is RAG and P nucleotides related
Rag Cleaves Hairpin loops and adds P nucleotides
what adds N nucleotides and where
terminal Deoxynucleotidyl Transferase (TdT) adds N to P nucleotides and other strand
what type of diversity does VDJ provide
combinatorial diversity
what is the same in all B and T cells
surrogate light chain
what do all B and T cells have
weak self recognition
when are B and T cells chosen for negative selection
strong self antigen recognition
what is the 1st checkpoint in T and B cell development
first polypeptide chain of Ag receptor completed so the H of BCR and the B of TCR
2nd checkpoint in B and T cell development
production of 2nd polypeptide chain is complete so a in TCR and k or lambda in BCR
why are these checkpoints used
ensure that Ag receptor gene rearrangement has correctly occurred for mature cells
what does selection eliminate
harmful self-reactive lymphocytes
what makes up a pre ag receptor
only one polypeptide chain
polypeptide chain in pre BCR
u Heavy
pre TCR has what chain
TCR B chain
where can apoptosis of BCR and TCR occur
1st and 2nd checkpoint
percentage of B and T cells with productive in frame rearragement of 1st polypeptide chain
30%
what happens to BCR and TCR with out of frame rearrangement
Pre not expressed and have apoptosis for no survival signal
what signals are provided by pre B and T cells
survival, proliferation and further development
if 2nd polypep of TCR and BCR has productive rearrangement, what occurs
pre to complete but still immature
positive selection
BCR and TCR with useful ag receptors
what do positive selected T cells have
CD 8 or CD4 who recognize MHC and not self-Ag
where does positive selection occur
thymus
why is negative selection needed
maintain central tolerance of self Ags
what does negative selection do
eliminate harmful T cells while altering harmful B cells
apoptosis of T cells by
clonal deletion
harmful B cell altered by
receptor editing with 2nd Ig rearrangment then if fails then clonal deletion
what do B1 cells come from
Fetal liver derived stem cells
what makes B 2 cells
Bone marrow precursors
why do B1 cells have limited BCR diversity
no TdT in fetal liver so no junctional diversity
where are most B1 cells found
self renewing in peritoneum and mucosal sites
what is secreted by B1 cells
IgM Abs
what are IgM for
react with Microbial polysacharides and lipids, oxidized lipids
what bacteria are most effected by IgM
Gram +
which antibodies are natural
IgM
what provides most of the IgM levels in early phases of infection
B1-B cells
what happens once immature B2 cells are made
relocated to the spleen
what do B2 cells become
marginal zone cells or follicular cells
where are follicular B cells found
recirculating lymphocytes
where are MZ cells found
spleen and lymph nodes
where in spleen do MZ cells go
splenic marginal zone, near marginal sinus
what do MZ cells respond to
Blood borne Ags
what is requires replenishment from bone marrow
F B cells
what do F B cells respond to
protein Ag
what are F B cells dependent on
T cells
what do F B cells easily undergo
isotype switching and affinity maturation
mature F cells become
long lived plasma or memory B cells
MZ are independent of
T cells
MZ respond and produce
to polysaccharide Ags and generate natural Abs
which organs house MZ cells
spleen and lymph nodes
which microbes to MZ cells react on and what do they produce
blood borne microbes and make short lived IgM plasma cells
what is mediated by MZ cells
T cell independent humoral immune response to circulating pathogens and some T cell dependent
Rearrangement of TCR B, y, sigma is initiated
simultaneously
what happens for a TCR to be of y sigma origin
rearranges before B
what percent of T cells are y sigma
10%
why do y sigma TCR have limited diversity
very few VDJ segments
