Antigen Recognition & Lymphocyte Development Flashcards
What are lymphocyte receptors?
Protein receptors displayed on the cell surface
What must happen for a lymphocyte receptor to work?
Receptor must:
1) bind to a ligand (main receptor)
2) signal transduction (accessory proteins)
What is the lymphocyte receptor complex?
Together:
1) the main receptor (which binds the ligand)
2) accessory proteins (take care of signal transduction)
What is the end result of receptor engagement (of the receptor complex)?
Activation: altered gene transcription/activation/effector function
What does BCR recognize? TCR?
BCR: recognizes 3D structures; macromolecules in their native conformation/as they exist in nature
TCR: recognizes only linear peptides that are complexed with MHC (8-30 aa in length)
What does it mean to say that T cells are MHC restricted?
They can only activate when they bind to an antigen presented by MHC, need BOTH
If no MHC, no activation of T cells
B cells are NOT MHC restricted
Name and describe the two receptor domains
1) variable domain: responsible for binding to antigen. This domain varies from one receptor to another, furthest from cell membrane
2) constant domain: structural, responsible for effector functions, same from one receptor to another, closest to cell membrane
Expression of co-receptors of TCR and BCR dictates what?
Dictates the developmental stage the cell is in
List TCR co-receptors
CD3
CD4
CD8
List BCR co-receptors
Iga Igb CD19 CD21 CD81
Describe the structure of the TCR
2 membrane bound chains:
1 alpha
1 beta
Each has a variable and a constant domain
Note: n terminus is at top and is portion that actually contacts antigen= hyper variable chains or complementary determining regions and this is what determines antigen binding in T cell receptor (short portions of variable loops in variable chains)
What part of TCR actually contacts antigen and what is it called? What does this region determine?
N terminus = hyper variable region= complementary determining regions
Determines antigen binding in T cell receptors
Small portions of aa’s of the variable loops (found on the loops at the tips of the variable region)
Overall B cell maturation process? Difference of T cell?
1) common progenitor in bone marrow– commits to b or t
2) B stays in bone marrow
3) matures in bone marrow
4) mature naive B cell leaves primary lymph and cycles between secondary lymph and blood, looking for antigen
5) majority never find their antigen– die and recycle
T cell: matures in thymus once committed to T, then same fate as B cell
How many complementary determining regions are there that interact with antigen? (same for antibodies)
3 CDRs in each Va and VB domains= 6 total. All six must match with the MHC 6
What is the most important interaction in immunology?
TCR MHC + antigen b/c as T cell goes, so goes while adaptive response
What must happen for T cell to activate?
Recognize both MHC and antigen. If not nothing happens and it moves on to next receptor
What must happen to start signal transduction in a T cell? (What is included in the TCR CD3 complex?
TCR CD3 complex includes: a and B chains of main TCR (CD8 or CD4) complexed with signaling proteins, which are collectively called CD3. Together with the zeta chains, CD3 and zeta chains initiate signal transduction.
If bind the right combo of MHC and antigen, have signal transduction
B cell receptor structure?
Y shape, 2 membrane bound arms, 4 chains
2 heavy (identical to eAch other), men bound 2 light (identical to each other), not men bound, stuck to heavy by S-S bonds
Tips= FAB regions (fragment antigen binding), can cleave into subunits, bind antigens (antigen binding fragments), each made of entire light chain (variable) and heavy variable domain
Base of y= Fc region (crystallize in sol) set effector functions, only heavy chain constant, set effector function of secreted antibodies
How many classes of antibodies are there?
5
Each FAB region has 6 CDRs
Each antibody can bind two of same antigen
What is the hinge region of BCR?
Region that can pivot, flex, flatten in order to bind two of the same antigen at same time
BCR light or heavy chain variable?
Both heavy and light have variable regions
What is included in the BCR complex?
Membrane bound antibody associated with signaling molecules: Iga and IgB
CD81, CD21, CD19= signal transduction
Explain the functional domains of an antibody? (Same as BCR)
2 functional domains:
1) FAB fragment (antibody binding fragment): light chain and heavy chain variable regions, and first constant region=tip of Y
2) Fc fragment (crystalline fragment): heavy chain constant= base of Y, determines antibody class and effector functions
BCR vs TCR effector functions?
TCR does NOT have effector functions b/c never secreted
BCR receptor functions determined by constant regions of secreted Ig, heavy chains class switch and change from membrane to secretory
Do antibodies get better at binding over time?
YES:)
Antigen binding region TCR vs. BCR
Ig made of 3 CDRs in each Vh and VL.
TCR made of 3 CDRs in each Va and VB
Signaling functions BCR (Ig) vs. TCR
BCR: IgA and IgB proteins
TCR: CD3 and zeta proteins
T/F we have billions of lymphocytes in our bodies and each T/B cell recognizes something completely different from all the other T/b cell
TRUE
What is the overall source of receptor diversity?
Genes that encode receptor proteins are segments (not complete)
Multiple segments for each portion of variable domain are randomly rearranged into a linear complete sequence, which is source of receptor diversity
Constant domain has no gene segment rearrangement
Combinatorial diversity vs. junction all diversity
Combinatorial= just putting the segments together (VDJ recombinase (products of), RAG-1 & 2)
Junction all diversity= random additions and deletions of nucleotides at junction site of different segments (TDT enzyme)
Variations possible in receptors
Variations possible in receptors
10^11 antibody receptors possible
10^16 T cell receptors
Corresponding chains and VDJ for B and T cells
VDJ stand for?
Constant vs variable domain segment rearrangement?
B= heavy (VDJ) and light (VJ) T= beta (VDJ) and alpha (VJ)
Heavy made first
Light made second
V= variable D= diversity J= joining
Constant has not gene segment rearrangement. Variable does
What are RAG genes? And RAG recombinase? TdT?
Rearrangement genes
RAG recombinase is protein that does the recombining: looping and cutting enzyme
TDT: terminal deoxynucleotidal transferase= enzyme that adds random nucleotides, leads to Junctional diversity
Explain the process of receptor rearrangement
1) D-J (via RAG recombinase) rearrangement, random nucleotides, ligand links
2) VDJ rearrangement (pick V)
3) transcription
4) alternate splicing
5) test with fake light chains
6) light chain: only VJ, rearrange and test
Each cell does this independently and randomly
Explain receptor editing/receptor rescue
As long as cell doesn’t use the most 5’ V and most 3’ J, turns rearrangement on again if the chains chosen previously are self reactive or defective
If it does happen, there are no rearrangements left b/c the middle stuff is cut out
When does alternate splicing occur?
After transcription
What is the flanking sequence linkages?
23s and 12s link, enzymes grab 23 and 12 then know where to put them together for the same orientation so that V’s and J’s don’t get inverted
23s and 12s = flanking sequences, they are used to recognize ends of segments, that gets folded, that’s where enzymes cut (RAG)
Junctional diversity
Overhangs are used to to tack on random nucleotides, happens when D and J are put together
Different with T cell receptors V’s B cell making process?
Different locus, same VDJ rearrangement
Beta first, alpha second
Beta: DJ, VDJ, transcription, alternate splicing, test BETA, if works: alpha chain rearrangement (V and J only), surface test, do it till it works or run out of options
Difference: gap between D and J sets, technically 2 shots depending on what choose (most 5’ V and 3’J, everything else gone, otherwise second shot)
Simplified form VDJ
Heavy (b cell)= beta (t): DJ, then VDJ
Light (b)= alpha (t): VJ only, not D
What happens if RAG gene not working?
Immunodeficiency because no b or T cells (bubble boy syndrome) b/c cannot recombine (no adaptive, cannot respond to microbe attacks)
Rearrangement summary
Heavy (BCR) beta (TCR) first: each has V,D, & J
- DJ, V-DJ at DNA level
- VDJ linked to closest C by alternative splicing at RNA level
Light (BCR) and alpha (TCR) second: only have V and J
- only occurs after successful heavy and beta rearrangements
- V-J at DNA level
- VJ linked to C by alternative splicing at RNA level
*codes through VDJC, then alternative splicing
Explain how allelic exclusion factors in to forming the receptors
Allergic exclusion: only 1 (mom or dad) receptor is expressed
Whichever chromosome site finished first is one tested first (slower one is stopped), rearrange other one if that doesn’t work
Prevents dual expression & ensures each lymphocyte only expressed 1 antigen specific receptor
As opposed to MHC which is codon infant expression
Critical concept about lymphocyte receptors?
Antigen specific clones arise before encounter with antigen!
Diversity of lymphocyte receptors is linked to lymphocyte development
Overview of lymphocyte development
Pro B/T: rearranging heavy/beta chain
Pre-B/T: testing heavy w/ fake light to test for correct shape (pseudo-chains)
If works: light and alpha rearrangements
Receptor on surface: whole thing tested
+- selection
Pos: can receptor recognize MHC? If no it’s worthless T cell
Neg: does it recognize self antigen? Want no
Receptor editing: test, bring back in, rearrange until all possibilities exhausted
T cell development
Start as double negative (CD4-, CD8-)
Double +
Positive selection: testing binding for MHC and peptide, shed one coreceptor
Single + or failure
5%survive rest have something wrong
During dev: any antigen recognized while maturing is assumed to be bad*** be only antigen present in bone marrow or thymus is self antigen!!!!
Positive vs negative selection in T cells
Pos: receptor’s ability to recognize MHC
Neg: ability to recognize antigen (do not want cuz that means self reactive bc in bone marrow or thymus)
What is AIRE
A transcription factor, autoimmune regulator that allows for thymic cortical epi cells to express non-thymic specific antigens (liver, testicular, etc)
What would happen if there was a defect in HLA-DM?
HLA-DM gets MHC class II to let go of clip peptide, so without it you can only screen against clip peptide, so you will get CD8s and NO CD4s (bc don’t have MHC class II)
T cell dev summary
Committed to T cell–> migrate to thymus
Pro-T: receptor rearrangement, several survival checkpoints
-/-, +/+, + (sequence of co-receptor expression)
Co-receptor is tied to MHC interaction
Which T cells are committed when they leave the thymus?
Only regulatory CD4 T cells are committed when they leave the thymus: they escape out of negative selection, suppress the immune response
T cells and their purpose? CD8 CD4 T1 CD4 T2 CD4 T17 CD4 regulatory
CD8: target and killed infected cells
CD4 T1: pro-inflammatory, activate macrophages and help for B cell antibody production
CD4 T2: help for B cell antibody production, esp. IgE
CD4 T17: enhance neutrophil response
CD4 regulatory: suppress immune response
B cell development MHC restricted?
No, only T
B cell development
In bone marrow: (lymphocytes committed to B lineage dev in BM)
Stem,pro (rearranging heavy, many checkpoints), pre, immature b (IgM) (has rearranged both, pass neg selection–> to periphery)
Periphery: mature b (mature= IgM and IgD)
Class set by heavy chain constant
Becomes mature while in periphery/final maturation step there
Only IgM is immature, IgD as well = mature (dual expression)
Positive V’s negative selection in b cells
Positive: can the receptor transmit a signal? (Functioning receptor?) (simpler than T cell)
Negative: high affinity binding in primary lymph tissue:
bind strong = die,
bind weak= inactivated, ignorance/anergy
No binding= good B cell!!
*can it recognize antigen in bone marrow, if yes it’s binding self which is BAD!
What happens with weak B cell binding in neg selection?
Driven towards ignorance or anergy
Ignorance: ignores its antigen: if binds it–> won’t activate then won’t express right adhesions to get to lymph node/ follicle then can’t interact with T cell
Anergy: permanent state of ignorance
B cell pop dynamics: what are the three types of B cells?
B-2= follicular B cells
B-1
Marginal zone = resting mature B cells
Traits of B-2 cells
Follicular B cells
- most common type
- found in lymph node
- standard B cells activating and making antibodies
B2 cell characteristics
- formed early in life
- long lived relative to folliculars (b1)
- express CD5
- found in peritoneal and pleural cavity fluid, recognize non normal flora (if intestinal perforation, etc)
- IgM at low levels
- some antibodies always made (such as blood antibodies) even though never activated (against other species)
- selected by ubiquitous self and foreign antigen
Marginal zone B cell characteristics
- in marginal sinus of spleen
- resting mature B cells
- quick response to common environmental and self antigens in the blood stream, crank out IgM into blood stream
