Describe 4 ways T cell receptors are similar to immunoglobulins.
(1) TCRs have a similar overall structure, containing an antigen binding site, 2 variable domains and 2 constant domains.
(2) Both are generated via somatic recombination of sets of gene segments.
(3)The variable region of the TCR conatains 3 complimentarity-determining regions encoded by Valpha and Vbeta (6 in total), analogous to the the Vh and Hl domains.
(4) Diversity is generated by the same means (combination of gene segments, junctional diversity due to P and N nucleotides and combination of 2 different chains)
Describe 5 ways in which TCRs are different than immunoglobulins.
1) TCRs have only 1 antigen binding site, while Igs have at least 2.
2) TCRs are never secreted
3) TCRs are generated in the thymus (instead of bone marrow)
4) TCRs have no effector function, and it does not undergo isotype switching,
5) TCRs do not undergo somatic hypermutation
Compare the organization of T-cell receptor alpha and beta genes with the organization of immunoglobulin
heavy-chain and light-chain genes.
The alpha locus is like the Ig light chain locus in that both contain V and J gene segments and no D segments. The TCR beta locus resembles the Ig heavy-chain locus in that both contain V, D, and J gene segments.
T-cell receptors do not undergo isotype switching. Why is this?
T-cell receptors are not made in a secreted form and their constant regions do not contribute to T-cell effector function. Therefore there is no need for isotype switching in T-cells.
The role of CD3 proteins and Delta chain on the surface of the cell is to
a. transduce signals to the interior of the T-cell
b. bind to antigen associated with MHC molecules
c. bind to MHC molecules
d. bind CD4 or CD8 molecules
e. facilitate antigen processing of antigens that bind to the surface of T cells
a (they also help the TCR bind to the cell membrane)
Which of the following accurately completes this statement? The function of _______ T cells is to make contact with _______ and __________.
a. CD8; virus-infected cells; kill virus infected cells
b. CD8; B cells; stimulate B cells to differentiate into plasma cells
c. CD4; macrophages; enhance microbicidal powers of macrophages
d. CD4; B cells stimulate B cells to differentiate into plasma cells
e. All of the above
a, c, d
The immunological consequence of severe combined immunodeficiency dz (SCID) caused by a genetic defect in either RAG gene is
1. Describe the structure of an MHC I molecule, identifying the different polypeptide chains and domains.
2. Which domains or parts of domains participate in the following: antigen binding; binding the T-cell receptor.
3. Which domains are the most polymorphic?
the complete MHC I molecule is a heterodimer made up of one alpha chain and a smaller chain called Beta2 microglobulin. The alpha chain consists of 3 extracellular domains (alpha1,2,3), a transmembrane region and a cytoplasmic region. The polymorphic class I molecules are HLA-A, HLA-B, and HLA-C. The antigen binding site is encoded by the alpha1 and alpha2 domains. The alpha3 domains binds the coreceptor of CD8.
Describe the structure of an MHC II molecule, identifying the different polypeptide chains and domains. Which domains participate in antigen binding and binding the TCR? and Which domains are the most polymorphic?
MHC II are heterodimers made up of alpha and beta chains. The overall molecule is similar to MHC I, but the three polymorphic regions in MHC II are HLA-DP, HLA-DQ, and HLA-DR. Antigen binds to the peptide-binding groove formed by the alpha1 and beta1 domains. The beta2 domain binds the TCR CD4.
Amino Acid variation among MHC class II allotypes that present antigens to CD4 T-cells is concentrated
What is (a) antigen processing, (b) antigen presentation? and (c) why are these processes required before T-cells can be activated?
a) antigen processing: the intracellular breakdown of pathogen-derived proteins into peptide fragments that are of the appropriate size and specificity required to bind MHC molecules.
b) antigen presentation: the assembly of peptides with MHC molecules and the display of these complexes on the surface of antigen-presenting cells.
c) Antigen processing and presentation must occur for T cells to be activated because T-cell receptors cannot bind to intact protein, only peptides, and TCRs do not bind antigen directly, but rather must recognize antigen bound to MHC molecules on the surface of antigen presenting cells.
Describe in chronological order the steps of the endogenous antigen-processing pathway for intracellular (cytosolic) pathogens.
Proteins from pathogens growing in the cytosol are broken down into small peptide fragments by proteasomes. The peptides are transported into the lumen of the ER using TAP (heterodimer of TAP1 and TAP2). MHC I is already in the ER and binds to peptides that fit the peptide cleft. MHC I alpha-chains are bound to to the chaperone calnexin until B2-microglobulin binds, and then bind the chaperones calreticulin and apasin until the peptide binds. Tapsin binds to TAP1, positioning the MHC I molecule near the peptide source. MHC class I molecules bound to peptide dissociate from the chaperone molecules and progress to the Golgi apparatus for completion of glycosylation and transport to the cell surface in membrane-bound vesicles.
Structure of a T cell
Either alpha/beta or gamma/delta heterodimers
Beta/delta: VDJ transcribed first, temporarily binds fake alpha/gamma to stabilize on the surface
CD3 complex homodimer made from one of 3 chains - gamma, delta or epsilon.
Antigen binding distorts T cell structure
Distortion activates CD3 intracellular signalling
CD4: binds MHC class II (2 x 4 = 8) -- effected by HIV.
CD8: binds MHC class I (1 x 8 = 8) -- can exist as hetero or homodimer.
Location of T cells in Thymus
Cortex: contains immature T cells
Medulla: contains mature T cells
T cell maturation
TCR gene rearrangement: Gamma/delta first, then alpha/beta (CD3 is stimulated spontaneously as the complex comes together, helps it anchor on cell membrane)
Double-positive: both CD4 and CD8 (random decision process depending on who binds first)
Positive selection: T-cells avoid programmed death by binding MHC
Negative selection: T-cells killed when they bind too tightly to self MHC.
Does the following describe class I or class II MHC? HLA-A,B,C: expressed codominantly (6 different class __ chains expressed on almost every cell).
Composed of MHC encoded alpha unit and non-MHC encoded B2-microglobulin. 3 alpha regions: first 2 polymorphic and 3rd conservative for B2-microglobulin and CD8 binding. Peptide cleft: fits 9 AA peptide generated by proteasome, first 2 regions are very polymorphic among a population, Tertiary complex req. Alpha unit, B2-microglobulin and peptide to be expressed, TAP: pumps cytoplasmic proteins into ER where MHC class __ is wating. Peptide loading occurs in ER
MHC class I (endogenous)
Does the following describe a class I or class II MHC?
APC displays exogenous proteins DR,DP,DQ Presented by macrophages (phagocytic), B cells (efficient, low antigen levels req. for response) and dendritic cells (migratory).
Heterodimer of alpha and beta chain. Peptide cleft: fits 9-15 AA peptide, normally displays recycled self antigen. Invariant chain(Ii): plugs up MHC class __ until fused w/ peptide containing vesicle HLA-DM removes invariant chain (the CLIP portion in the peptide cleft)
Peptide loading occurs in specialized vesicular compartments.
MHC class II (exogenous)
TCR antigen presentation
TCRs only recognize peptide antigens (Ig recognizes anything)
Antigen must be unfolded
MHC restriction: T-cells only recognize self MHC molecules (why skin grafts don't are rejected)
Both class I and II are most often presenting self-antigen in their clefts.
Endocytosis of an extracellular foreign protein antigen is presented by which class of MHC?
MHC class II (leading to CD4 response)
Endogenous synthesis of a foreign protein antigen is presented by which class of MHC and elicits which type of T-cell response?
MHC class I => CD8
All nucleated cells express MHC class I, but which express MHC class II?
Vascular endothelial cells
Why do heterozygotes have an advantage in peptide selection?
Because they are more likely to have 6 different class I chains expressed on MHC I molecules. (Homozygotes my express as few as 3, so they bind to fewer foreign antigens)
Notch1 (if a cell has Notch => T-cell, if not => B-cell)
Alpha:beta and gamma:delta cells develop from a common _________ ___________ T-cell precursor.
What is the function of AIRE?
AIRE is an autoimmune regulator that activates transcription in the thymus of every self peptide, so T-cells go through negative selection of EVERY self antigen.
What would happen if B2-microglobulin could not bind the the MHC class I alpha-chain?
The MHC class I would be retained in the ER and would not be transported to the cell surface. It would remain bound to calnexin and not fold into the conformation needed to bind to peptide.
What would happen to MHC class I if the TAP transporter were missing or non-functional?
Peptides would not be transported into the lumen of the ER. So MHC I would not have any peptides to present on the cell surface => MHC class I deficiency. (bare lymphocyte syndrome is characterized by a defective TAP)
What removes CLIP from MHC class II?
Where does an MHC class II bind with a peptide to present on the cell surface?
MHC II molecules that are being transferred from the ER to the cell surface intersect with phagolysosomes, which contain degraded extracellular pathogens. These peptides are loaded into the antigen-binding groove after CLIP is removed by HLA-DM. The MHC-peptide complex then progresses to the cell surface to present the antigen.
Other than protecting the petpide-binding groove in MHC class II, what is the function of the invariant chain?
Transporting MHC II to the phagolysosome.
How do MHC variation due to multi-gene family and allelic polymorphism influence the antigen that a person's T-cells can recognize?
TCRs bind to peptide antigens in the form of peptide:MHC complexes. To bind specifically the TCR must fit both the peptide and the part of the MHC molecule surrounding it in the peptide-binding groove.
Because each individual produces a number of different MHC molecules from their MHC class I and class II multigene family, the TCR repertoire is not restricted to recognizing peptides that bind to just one MHC molecule. Instead, the TCRs repertoire can recognize peptides with different peptide-binding motifs during an immune response., increasing the likelihood of antigen recognition and T-cell activation.
The polymorphism in MHC molecules is localized in the regions affecting T-cell reetpor and peptide binding. Thus, a TCR that recognizes a given peptide bound to variant 'a' of a particularMHC molecule is likely not to recognize the same peptide bound to variant 'b' of the same MHC molecule. Polymorphism also means that MHC molecules of one person will bind a different set of peptides from those in another person. Taken together, these outcomes mean that because MHC polymorphism, each individual recognizes a somewhat different range of peptide antigens using a different repertoire of TCRs.
Place the following phases of a B-cell's life history in the correct chronological order.
a. negative selection
b. attacking infection
c. finding infection
d. searching for infection
e. repertoire assembly
f. positive selection
e, a, f, d, c, b
Place the following stages of B-cell development in the correct chronological order.
a. early pro-B
b. large pre-B cell
c. immature B cell
d. stem cell
e. late pre-B cell
f. small pre-B cell
d, a, e, b, f, c
What is the importance of bone marrow stroma for B-cell development?
bone marrow stroma provide the necessary environment for B-cell development by expressing secreted products and membrane-bound adhesion molecules. Cytokines such as IL-7 have an important role in later stages of B-cell development, stimulating the growth and cell division of late pro-B and pre-B cells.
What would be the result of anit-IL-7 antibodies in the bone marrow?
developing B-cells would be arrested at the late pro-B or pre-B cell stage and would not be able to progress normally to the immature B-cell stage.
What are the 2 main checkpoints of B-cell development in the bone marrow?
1) when the heavy-chain binds the surrogate light-chain (VpreBgamma5).
2) When a complete B-cell receptor is expressed on the B-cell surface.
a. What is the fate of B-cells after the heavy-chain is formed? (checkpoint 1)
b. after the light-chain is formed? (checkpoint 2)
c. how does this lead to allelic exclusion?
a. if the VDJ rearrangement gave rise to a functional pre-B cell receptor the late pro-B cell will undergo clonal proliferation. If the rearrangement was non-functional the B-cell has one more chance to create a functional BCR or it undergoes apoptosis.
b. production of a functional light-chain results in assembly of a surface Ig and the survival and maturation of the B-cell. If it is non-functional the B-cell has 3 more chances to create a functional light-chain or it undergoes apoptosis.
c. allelic exclusion = ensuring that only 1 heavy-chain and 1 light-chain are expressed. If a cell passes checkpoint 1, heavy-chain rearrangement is stopped, and the surrogate light chain becomes unavailable. Checkpoint 2 signals the end of light-chain rearrangement.
a) What is the function of B1 cells?
b) and how do they differ from B2 cells?
a) B1 cells are best associated w/ the innate immune response because of their rapid response to antigen, their limited diversity and their polyspecificity.
b) B1 cells express CD5, have few N nucleotides at VDJ junctions and have a restricted range of antigen specificities. They produce IgM antibodies of low affinity and respond mainly to carbohydrate, rather than protein, epitopes. B1 cells are polyspecific for antigen (they bind several different antigens).
Which of the following are true of centrocytes?
a. somatic hypermutation has occurred.
b. they are large proliferating cells.
c. isotype switching is complete.
d. they produce secreted forms of immunoglobulins.
e. they lack MHC class II molecules on the cell surface.
What is the purpose of negative selection?
to ensure that auto-reactive B-cells are prohibited from emerging in the body.
Immunological tolerance in the B-cell repertoire is called _______ tolerance when it develops in primary lymphoid organs, and ______ tolerance when it is induced outside the bone marrow.
A plasma cell is characterized by which of the following?
a. it differentiates in the medulla of lymph nodes and teh bone marrow.
b. it dedicates 10-20% of total protein synthesis to antibody production.
c. levels of MHC class II molecules are elevated.
d. it undergoes extensive proliferation in germinal centers.
e. it produces secreted Ig instead of membrane-bound forms.
What is the T-cell analog to the B-cell VpreBgamm5 surrogate light chain?
preTalpha, which combines the T-cell receptor B-chain with the first 2 T-cell receptor chains to be expressed, to form the pre-TCR. pTalpha also binds CD3 to complete the complex that induces T-cell proliferation and the cessation of rearrangement of the Beta-chain if it is functional.
Double negative thymocytes initiate rearrangement at the ______ locus before all other T-cell receptor genes.
Beta and gamma
The function of negative selection of thymocytes in the thymus is to eliminate
During the early developmental stages of alph:beta T-cells in the thymus there are two key checkpoints that must be satisfied to permit the progression of T-cell development. Explain what occurs at each checkpoint.
1) after the rearrangement of the beta-chain locus the first checkpoint occurs, which tests for the ability of the beta-chain to associate with pTalpha and form the pre-TCR on the cell surface.
2) The 2nd checkpoint occurs after the rearrangement of the alpha-chain locus, which tests for the ability of the alpha-chain to associate w/ the beta-chain and form the TCR on the cell surface.
If a TCR on a double + thymocyte binds to a self-peptide:self-MHC class I complex with low affinity the result is
In T-cells, allelic exclusion of the alpha-chain is relatively ineffective, resulting in the production of some T-cells with 2 TCR's of differing antigen specificity on their cell surface.
a) Will both receptors have to pass + selection for the T-cell to survive?
b) Will both receptors have to pass negative selection for the cell to survive?
c) What if T-cells having dual specificity survive these selections and are exported to the periphery?
a) No, only 1 has to pass for the cell to move to the next stage.
b) Both have to pass negative selection, if just 1 fails the cell will undergo apoptosis.
c) a cell could that binds recognizes self antigen and a pathogenic antigen could be activated => an autoimmune response.
Why don't T-cells undergo somatic hypermutation?
Because of the req. of T-cells of dual recognition (peptide-MHC) somatic hypermutation would be likely to change the specificity of the T-cell.
Would a MHC class II deficiency (a) affect the development of CD4, CD8 or both types of T-cells?
b) How would this impact B-cells?
a) Just CD4, CD8 would still be selected for when dual + T-cells bound to MHC class I.
b) To produce antibody B-cells req. the cytokines released from CD4 T-cells. So lacking CD4 cells would lead to low immunoglobulin levels and the inability of B-cells to differentiate into plasma cells.
As we age our thymus shrinks, or atrophies by a process called involution. Yet T-cell immunity is still functional in old age.
a) How do T-cell #s in the periphery remain constant in the absence of continual replenishment from the thymus?
b) how does this differ from the maintenance of the B-cell repertoire?
a)T-cells in the periphery self-renew by cell division (and are long lived) after thymic atrophy.
b) B-cells are short lived and replenish from immature precursors derived from the bone marrow.
How does the processing of T-cells in the thymus differ from processing in the periphery? what are the advantages/disadvantages?
AIRE is present in the thymus which causes genes normally expressed in other tissues to be expressed in the thymus, generating a more comprehensive repertoire of self-peptides, increasing the # of potentially autoreactive T-cells that are killed during negative selection.
a) What is the role of regulatory CD4 T-cells (Treg)?
b) How is it different from other CD4 T-cells?
a) Treg suppresses the proliferation of naive autoreactive CD4 T-cells by secreting inhibitory cytokines. This inhibitory action req. that both the Treg and other CD4 T-cells interact w/ the same antigen-presenting cell.
b) Treg expresses CD25 and FoxP3 (transcription repressor protein) on the cell surface.
True or False:
In adults the mature T-cell repertoire is self renewing and does not require a thymus for provision of new T-cells.
People with defective AIRE (autoimmune regulatory) gene exhibit
People lacking a thymus have _________.
Why would an organ transplanted from any donor other than an identical twin almost certainly be rejected in the absence of any other treatment?
acute rejection is due to immune responses to the HLA class I and II molecules of the graft that are different from those of the recipient and are therefore viewed as foreign. If the HLA class I and II molecules are the same, the differences in the peptides that are bound by the MHA molecules will trigger rejection through an allorecognition response.
_______ describes polymorphic antigens that vary between individuals.
How does chronic rejection differ from acute rejection?
Acute rejection occurs only a few days after transplantation and is mediated by CD4 and CD8 T-cell adaptive immune responses to the foreign peptides bound the HLA molecules. Chronic rejection occurs months to years after transplantation and is mediated by anti-HLA class I and II alloantibodies produced via the indirect pathway of recognition. CD4 T cells are activated by dendritic cells presenting donor-derived HLA class II allotypes of the recipient. These activated CD4 T-cells activate B-cells, which are presenting donor-derived allogeneic HLA peptides. This cognate interaction results in the production of anti-HLA class I and class II antibodies.
Identify 3 classes of drugs that are used to suppress acute transplant rejection, provide examples of each and name the toxic side-effects of each class.
1) corticosteroids: can lead to fluid retention, weight gain, diabetes, bone demineralization and thinning of skin. (prednisone)
2) cytotoxic drugs: nonspecifically prevent DNA replication in all mitotic cells (ie cancer drugs), liver and bladder damage. (cyclophosphamide, methotrexate)
3) T-cell activation inhibitors: nephrotoxicity and suppression of B-cell activation. (cyclospoirn A, rapamycin)
Graft-versus host disease is a consequence of what?
mature T lymphocytes from the donor mounting an immune response against tissue of the recipient.
What limits the effectiveness of mouse antibodies in tissue transplantation?
MoAbs (mouse antibodies) aer antigenic in species and stimulate anti-MoAb responses. Anti-CD3 MoAbs are often administered to suppress T-cell activity in recipients, but repeated doses leads to the formation of a anti-MoAb immune response that diminishes and eventually completely stops the activity of the anti-CD3 MoAbs.
Why does a bone marrow donor need to be HLA matched to the recipient?
The recipient's MHC molecules mediate + selection of thymocytes in the thymus that interact w/ donor-derived MHC molecules in the periphery.