Immunogenetics - Bowden (Completed) Flashcards
What part of the TCR complex is part of signal transduction? 7
The 2 ζ chains (3 ITAMs each)
What is unique about gene arrangement events in B cells? What do we call this? 11
Process called Clonal selection
Gene arrangement events occur in the ABSENCE of Ag
What is Allelic Exclusion? 11 (Add0
Monospecific, each arrangement codes for a particular receptr (add)
What is combinatorial Diversification? 13
Involves multiple germ line genes (one from mom, dad)
V-J or V-D-J recombination
What is Junctional Diversity? 13
Adding nucleotides during process of D-J or V to DJ joining
Light chain is to B cell as a __________ is to a T cell? 14
α chain
From left to right what is the order of VDJ on a gene? 15
V –> D –> J –> C
How many constant regions does a VDJ gene contain? What do they pertain to? 15
9 constant regions - one for each Ig region
What’s the order of joining of the V, D, J, and C regions of a germline gene? 18
D-J join –> V-DJ joining –> VDJ-C joining
What segment isn’t present in the light germline DNA of the BCR? 18
There is no D segment in the light chain
What process causes the excising of various V,D,J,C regions to end with a single segment of each? 19
Somatic Recombination
What is responsible for ensuring genes are joined in the correct order? 20
Recombination signal sequences (RSS)
What is the role of V(D)J Recombinase? 21
Forms the new regions between the various V,D, and J regions after they have been cut
What produces recombination activating genes (RAG-1 and RAG-2)? 21
Produced by lymphocytes
What is the role of terminal deoxyribonucleotidyl transferase (TdT)? 23
Adds nucleotides to asymmetrically cleaved hairpins - called P nucleotides
How do N nucleotides differ from P nucleotides? 23
P nucleotides - nucleotides added to asymmetric hairpins by TdT
N nucleotides - nucleotides added in a non-templated (random manner)
What does the addition of N and P nucleotides during junctional diversity do? 23
It further diversifies the third hypervariable region
What kind of errors are often seen during junctional diversity? What region does this occur in? 24
Frameshift mutations
Occur in the hypervariable region
For a single specific Ig how are the heavy and light chains manufactured? 27
The heavy and light chain are produced completely separate from one another for added variability
When does the second combinatorial diversity occur? 30
After both receptor chains have been expressed on the cell surface of the B or T cell
What is somatic hypermutation? 13
Point-mutations that occur in fully assembled chains (V-D-J and V-J)
Aka magic
Where would you find a T cell stem cell, pro-lymphocyte, pre-lymphocyte? 33
Within the generative organ of the bone marrow
In B cells which gets produced first, the heavy or the light chain? 36
Heavy then light
When does the CD19 marker become associated with a B cell? 37
At the Pro-B cell stage
What are the CD markers for B stem cells? 37
CD43+
What are the CD markers for Pro-B cells? 37
CD43+
CD19+
CD10+
What are the CD markers for Pre-B Cells? 37
CD43+ (B220lo)
When is the heavy chain added to a B cell? What else is present? 37
At the Pre-B stage, a surrogate light chain is also present
When is the light chain added to the B cell? 37
At the Immature B cell stage
What is the role of the surrogate light chain? 37
To hold the heavy chain in place as a place-marker until the light chain can be produced
Once it’s determined that the heavy chain of the Pre-B cell is functional what is the next step? 37
Proliferation of that cell
the heavy chain works so a bunch of copies are made to hopefully add a bunch of various light chains to gain diversity
When is the B cell checked for reactivity against self-Ags (negative selection)? 37
During the immature B cell stage after the light chain has been added to the BCR
What stops B cell heavy chain recombination? 38
Production of a successful, working Heavy chain
What does alternative splicing of mRNA have to do with Heavy chain expression? 40
Pre-mRNA can be processed in variety of ways
Initially pre-mRNA can be brought to and made contiguous with the μ or δ constant chain resulting in IgM or IgD
What is the role of Bone marrow stromal cells with B cells during their development? 41
Stromal cells express adhesion molecules and secrete cytokines to further development
What stages of stromal cell support and B cell maturation involve VLA-4 and VCAM-1? 41
(Maturing B cell) VLA-4 — VCAM-1 (stromal cell)
- ) Stem cell/lymphoid progenitor
- ) Early pro-B cell
- ) Late pro-B cell
What is the role of RAG 1 & 2? 21
Cleaves hair pin loops and ligases breaks back together
What region defines the medulla of the Thymus? 44
The inner cortical region
What region defines the thymic cortex? 44
The outer cortical region
What T lymphocytes are present in the thymic cortex? 44
Double negative T cells (CD3-4-8-)
Double positive T cells (CD3+4+8+)
What is the only route by which progenitor cells enter the thymus and mature lymphocytes leave the thymus? 45
Via the blood (No HEV)
What stage of T lymphocyte leaves the bone marrow? 46
T lymphocyte stem cell heads to the Thymus before entering the pro-T stage
When does the developing T cell achieve a CD marker of CD25+
At the pro-T stage within the cortex of the thymus
At what stage is the first chain added to the T cell? What is the chain that is added first? 46
At the Pre-T cell stage the β-chain is added (α-chain surrogate)
What markers are present on a T stem cell, Pro-T cell, and Pre-T cell? 46
T stem cell –> c-kit+, CD44+
Pro-T cell –> c-kit+, CD25+, CD44+
Pre-T cell –> c-kit+, CD25+
During the maturation process how are CD4+ and/or CD8+ added to the developing T cell? 48
Developing T cells are first double-negative with no cluster differentiation
Next, the T cells are double-positive with both CD4+ and CD8+
Finally, either CD4+ or CD8+ is removed via proteolysis (what does this depend on?)
What determines whether a developing T cell will be CD4+ or CD8+? 49
It’s based off what HLA class they interact with first
Where is the TCR β gene locus found? 50
On chromosome 7
What is a desired affinity for self-Ag for developing T and B cells? 52
Weak antigen recognition
Differentiate positive vs negative selection. 52
Positive selection - Weak Ag recognition by a B or T cell = keep the cell
Negative Selection - strong Ag recognition by a B or T cell = degrade the cell
What happens to a B cell that binds soluble self-Ags? 54
B cell is induced to become anergic
What happens if a B cell, at first, binds self-Ag? 55
Occurs in anergic B cells that have been shut down due to self-Ag recognition
B cell undergoes receptor editing of the light chain
There are two genes λ κ
If κ is expressed and is self-reactive you can shut it down and try using the λ gene
If the λ gene works and is producing light chain that is not self-reactive tolerance is established and the new λ chain will eventually out-compete the κ chain
How many genes are their for the light chain of a B cell?
2 –> κ and λ
Why is IgM the first Ig produced by B cells? 56
It’s the first on the gene and therefore first expressed
What’s happening during T cell positive selection? 58
Self-restriction establishment
Recognition of self-MHC
What is the cluster differentiation of Tregs? 62
CD4
CD3
CD25 (receptor for IL-2)
What is the function of Treg cells? 62
To inhibit self-reactive T cells
What is the transcription factor that allows for Treg development? 62
FoxP3
What is important to understand about clonal selection? 64
Happens within lymphocytes during development INDEPENDENT of Ag
What do B cells undergo that T cells do not undergo? 65
Somatic Hypermutation
What is a weak affinity required instead of no affinity for self-Ags for B and T cells?
Needed to stimulate the HLA response and so that they recognize what a self-cell
Besides ensuring correct order of VDJ heavy segments, what does recombination signal sequences do? 20
Provides recognition sites for enzymes that cut and rejoin DNA
What are the two light chain genes? 20
λ and κ
What do errors in Junctional diversity often cause? 24
Frameshift mutations can triple the diversity of VDJ recombination (within the variable region)
What is the second type of combinatorial diversity? 30
The rearrangement of:
Heavy & light chains - B cell
α/β or γ/δ - T cells
Completely randomized to allow for greater diversity
Differentiate the markers for an immature B cell vs a mature B cell. 37
Immature B cell:
CD43-
IgM-lo
Mature B cell:
IgM-hi
What stages of stromal cell support and B cell maturation involve Kit and SCF? 41
(Maturing B cell) Kit — SCF (stromal cell)
- ) Early pro-B cell
- ) Late pro-B cell
What stages of stromal cell support and B cell maturation involve CAMs? 41
Stem cell/lymphoid progenitor cell
Pre-B cell stage
What T lymphocytes are present in the thymic medulla? 44
Single positive:
Mature CD4+8- or CD8+4-
(Macrophages also)
Where is the T cell α chain locus found? 50
On chromosome 14
What is the total potential repertoire w/ junctional diversity? 50
10^(7) B and T cell clones
What’s happening during T cell negative selection? 58
Central self tolerance establishment
Only T cells that don’t become activated by self-ag survive
How many different possible Abs can be produced?
10^(14)
Where are the two heavy chains for B cells encoded?
On chromosome 14
What is the make-up of the light chain of a BCR?
either 2 κ or 2 λ make up the two light chains, but NEVER mixed
What chromosome is the λ chain found on?
Chromosome 22
What chromosome is the κ subunit found on?
Chromosome 2
How many V, J, and C segments does the κ subunit of the light chain have?
3-35 V regions
5 J regions
1 C region
What is the condition of a Pre T cell when it enters the thymus?
Double negative = no receptors
What chromosome is the β chain of a TCR encoded from?
Chromosome 7
Which cells is V(D)J recombinase expressed in?
Within immature B and T cells
What does RAG 1 & 2 bind to?
RSS
12bp and 23bp spacers
What is the mechanism of action for RAG 1 & 2 (walk-through)?
Both RAGs bind RSS (spacers) at the end of regions of interest (V,D,J regions)
The RAGs then come together forming a loop and cleave out an undesirable segment/region
Single loops can be combined with double loops, but never 2 single or 2 double
What is somatic hypermutation?
Happens in B cells only after they have bound Ag and begins proliferation
During proliferation there are point mutations on the heavy and light chain resulting in affinity maturation
What is the principle determinants for graft acceptance or rejection?
HLA proteins (Class I & II)
Differentiate cleft binding for Class I vs Class II?
Class I: <10 aa
Class II: 13-25aa
How many types of Class I HLA are there?
6 types
