immuno final exam practice questions

The mechanism that permits immunoglobulins to be synthesized in either a membrane-bound or secreted form is

(1) allelic exclusion (2) codominant expression (3) class switching (4) the one-turn/two-turn joining rule (5) differential RNA processing

Kappa and lambda light-chain genes

(1) are located on the same chromosome (2) associate with only one type of heavy chain (3) can be expressed by the same B cell (4) all of the above (5) none of the above

A B cell becomes immunocompetent

(1) following productive rearrangement of variableregion heavy-chain gene segments in germ-line DNA (2) following productive rearrangement of variableregion heavy-chain and light-chain gene segments in germ-line DNA (3) following class switching (4) during affinity maturation (5) following binding of TH cytokines to their receptors on the B cell

Generation of combinatorial diversity among immunoglobulins involves

(1) mRNA splicing (2) DNA rearrangement (3) recombination signal sequences (4) one-turn/two-turn joining rule (5) switch sites

Describe three mechanisms by which antigen can enter the lymph node and make contact with the B-cell receptor

(a) Small, soluble antigens can be directly acquired from the lymphatic circulation by follicular B cells, without the intervention of any other cells. These antigens enter the lymph node via the afferent lymph and pass into the subcapsular sinus (SCS) region. Some small antigens may diffuse between the SCS macrophages that line the sinus to reach the B cells in the follicles. (b) Other small antigens leave the sinus through a conduit network. Follicle B cells can access antigen through gaps in the layer of cells that form the walls of the conduits. (c) Larger antigens are bound by complement receptors on the surfaces of SCS macrophages. Antigen-specific B cells within the follicles can acquire the antigens directly from the macrophages and become activated.

What is AICD? What is its function and how does it carry out its control of effector cell populations?

AICD is activation induced cell death. AICD is a way to reduce effector cell populations after antigen clearance. It reduces the pool of activated T cells after the antigen is cleared, and it helps to remove stray autoreactive T cells that are stimulated by self antigens. The Fas ligand binds to Fas. Both are present at cell surfaces as homotrimers. The binding of FasL causes a conformational change in Fas. This then binds death domain containing adaptor proteins. The adaptor proteins then recruit and activate caspase 8. This cleaves caspase 3. The activated caspase 3 cleaves I-CAD, the inhibitor of CAD, which is released to enter the nucleus and cleaves DNA

Explain the difference between the terms antigen-presenting cell and target cell, as they are commonly used in immunology.

APC = cells that can display antigenic peptides associated with class II MHC molecules and can deliver a costimularity signal to CD4+ TH cells. Target cell = any cell that displays peptides associated with class I MHC molecules to CD8+ TC cells.

Why doesn't apoptosis trigger inflammation the way necrosis does?

Apoptosis is programmed cell death, and the cell death occurs without triggering inflammation. Inflammation is not triggered because apoptosis is a controlled process of dismantling cells. Apoptosis is energy dependent, and the cells dismantle their contents without disrupting the membranes. The neighboring cells are induced to engulf the apoptotic cells before they can release inflammatory material. With necrosis, the cells die due to injury to toxicity. With the necrosis, the cells swell and burst. The contents of the cells are released. This release of contents can potentially trigger a damaging inflammatory response

Compare functions of B-1 and Marginal Zone B cells to conventional B-2 cells. What is the main purpose of these non-conventional B cells in immunity?

B-1 and Marginal Zone B cells have the ability to occupy different anatomical niches. For example, this is seen with the pleural and peritoneal cavities in mice. These also undergo apoptosis unless there is an interaction with self-antigens that occurs. B-1 and Marginal Zone B cells have limited V region diversity. The B-1 and Marginal Zone B cells are seen with IgM. These also have the ability to self renew. The conventional B-2 cells are present within the secondary lymphoid organs. The B-2 cells can go through apoptosis when there is an antigen challenge presented. B-2 cells possess a highly diverse V region. These are seen with IgG. B-2 cells are formed from the bone marrow. The main purpose is that the non-conventional B cells in immunity have the ability to generate antibodies against antigens that are seen within numerous types of bacteria. These cells also have the ability to provide many antibodies without having assistance or help from T cells

Upregulation of _________________ on dendritic cells, macrophages and B lymphocytes with activation provides the right signals to activated T cells.

CD80/86

Knockout of this T cell antigen in mice leads to uncontrolled proliferation causing lymphoadenopathy (enlarged lymph nodes), splenomegaly (enlarged spleen) , autoimmunity and death within 3-4 weeks

CTLA-4

What is class switching? What cytokines drives switching to IgG, IgE or IgA?

Class switch recombination is the generation of antibody genes for heavy chain isotypes other than u or delta by DNA recombination. With this, a B cell's production of a specific immunoglobulin changes. For Ig1 and IgE, IL-4 is involved. For IgA and IgG2b, TGF-beta is involved. For IgA synthesis, IL-5 is involved. For IgG3 and IgG2a, IFN-gamma is involved.

In general terms, describe the process of cross-presentation. Which cell types are most likely to be involved in cross-presentation, and what unique role does this process play in the activation of naïve CD8 T cells?

Cross-presentation is the ability of certain antigen-presenting cells to take up, process and present extracellular antigens with MHC class I molecules to CD8 T cells (cytotoxic T cells)

Trace the progress of early thymocytes from DN1 to DN4 including the markers that define each stage and the description of development for each stage. At what stage is the early thymocyte committed to the T cell lineage?

DN1 is defined by the marker c-kit++CD44+CD25-. During DN1, the thymocytes enter the thymus and are still capable of giving rise to multiple cell types. They express only the C-KIT and CD44. Within the thymic environment they become resident in the cortex. DN2 is defined by the marker c-kit++CD44+CD25+. During DN2, the thymocytes proliferate and express CD25. In the late stages, they commit to the T-cell lineage. THe expression of both C-KIT and CD44 is reduced. During the transition from DN2 to DN3, the rearrangement of TCR-gamma, TCR-delta, and TCR-beta chains continues. This is when the first decision in T-cell development occurs of whether to join the TCRgamma-delta or the TCRalpha-beta T-cell lineage. DN3 is defined by the marker c-kit+CD44−CD25+ The thymocytes that successfully rearranged their beta chain are committed to the TCR alpha-beta T-cell lineage lose expression of CD25. The proliferation is stopped, and they enter their final phase of their DN stage of development. DN4 is defined by the marker c-kitlow/−CD44−CD25. These mature directly into CD4+CD8+ DP thymocytes. The early thymocyte becomes fully committed to the T-cell lineage in the late DN2 stage

During which stage of T cell development in the thymus do cells become fully committed to the T cell lineage?

DN2

What is the purpose of Positive and Negative Selection in the thymus? Outline/diagram how Positive & Negative Selection and Neglect occur in the cortex of the thymus. What is the relationship between TCR affinity and this selection?

DP thymocytes undergo thymic selection based on affinity of TCR for MHC/peptide in the thymic cortex. Positive selection selects thymocytes that have receptors capable of binding self-MHC molecules, and this results in MHC restriction. The cells that can bind to MHC shift from DP to SP. The cells are selected to the CD4+ subset if the TCR can bind to an MHC class II molecule and bind to a CD4 molecule. If the TCR binds to an MHC class I molecule, the cell is selected to the CD8+ subset. Negative selection selects against thymocytes that have high-affinity receptors for self-MHC/peptide complexes, and this results in self-tolerance. Most cells fail positive selection and fail to receive the needed survival signals. Negative selection rids autoreactive clones and is responsible for central tolerance. Through clonal deletion, the induction of apoptosis in cells with too strong anti-self signaling or binding occurs. Negative selection is optimally mediated by APC and interactions that activate mature T cells. There are three possible outcomes that occur based on affinity of TCR when T cells encounter these self-peptide/MHCs. One is that the TCRs bind, and the cells die by neglect. The second is that the TCRs bind too strongly, and negative selection/deletion occurs. The third is that the TCRs bind just right, and positive selection to the single-positive stage occurs.

How do caspases trigger cell death? What is the difference between the extrinsic and intrinsic apoptotic pathways?

Different stimuli initiate apoptosis, but all activate caspases. Caspases trigger cell death through initiator caspases and effector caspases. The initiation is activated by a death stimulus. The effector caspases are activated by initiator caspases. The effector caspases cleave critical targets that are needed for cell survival, like cytoskeletal proteins. The effector caspases activate other enzymes that dismantle the cell. This results in an ordered disassembly of intracellular molecules as cell contents are packaged into vesicles, and thus the apoptotic bodies can be engulfed. The intrinsic pathway is mediated by mitochondrial molecules. On the other hand, extrinsic pathways use membrane-bound or soluble ligands to stimulate caspase activation.

Which of the following is NOT involved in the intrinsic apoptotic pathway of TCR-mediated negative selection in the thymus?

FADD

Which master transcriptional regulator is associated with TReg cells?

FOXP3

In the kinetic model for lineage commitment of T cells, weaker positive selecting TCR signals result in CD4+ lineage commitment, while stronger positive selecting TCR signals result in CD8+ lineage commitment.

False

Give three examples of nonprofessional APCs. When are these cells most likely to function in antigen presentation?

Fibroblasts-skin Flial cells-brain Pancreatic beta cells

Graph the concentration and isotype of serum Ab following primary and secondary immunization with Ag. Describe the functional differences between primary and secondary B cells that participate in the primary and secondary responses

For the lag period after antigen administration, the naive B cell takes about 4-7 days, the memory B cells take about 1-3 days. The time of peak response for naive B cell takes 7-10 days, whereas memory B cells take 3-5 days. With magnitude of peak antibody response, it varies with naive B cells, but with memory B cells it is generally 10 to 1000 times higher than primary response. With the type of antibody produced, IgM predominates in early response for naive B cells, whereas IgG predominates with memory B cells and IgA with mucosal tissues in memory B cells. With antigens, the naive B cell is thymus independent and thymus dependent, whereas memory B cells are primarily thymus dependent. With antibody affinity, naive B cells have low affinity, and memory B cells have high antibody affinity. With naive B cells, the life span of cells is short-lived whereas with memory B cells it is long-lived. Both naive B cells and memory B cells have recirculation.

Briefly describe the similarities and differences among cytokines, growth factors, and hormones.

Growth factors, cytokines, chemokines and hormones share many similarities and act as essential biochemical messengers for housekeeping functions, as well as, having crucial roles in pathological and stem-cell related growth and regulation. Each performs both homeostatic and stimulatory functions. Cytokines, growth factors (GF), and hormones are all chemical messengers that mediate intercellular communication. The regulation of cellular and nuclear functions by cytokines, growth factors, and peptide or protein hormones is initiated through the activation of cell surface receptors (Rc). All receptors have two main components: 1) a ligand-binding domain that ensures ligand specificity and 2) an effector domain that initiates the generation of the biological response upon ligand binding. The activated receptor may then interact with other cellular components to complete the signal transduction process. Many growth factors bind to receptors that are linked through G-proteins to membrane-bound phospholipase C (PLC). Cytokines are a broad category of small proteins about 5-20kDa, which are synthesized and secreted by immune cells or some non-immune cells. Immune cells include B lymphocytes, T lymphocytes, mast cells, etc and non- immune cells contains endothelial cells, fibroblasts, various stromal cells and so on. Cytokines act through combining related receptors. the combination can regulate cell growth, cell differentiation and modulate immune response. Hormones are regulatory biochemicals and produced in all multicellular organisms by glands. Hormones are not only proteins, which can also be steroid, amino acid derivatives and fatty acid derivatives. They can change the homeostasis, reproduction, development, and/or behavior. Hormonal signaling involves biosynthesis, storage and secretion, transport, recognition, relay and amplification and degradation.

Outline the steps in lymphocyte differentiation from HSCs to CLPs. Include expression of cell-surface markers and the cell types that each stage can differentiate into.

HSCs that are bound for a B-cell fate, the following transcription factors are involved in the earliest stages of the B-lineage development: Ikaros, Purie box factor 1 (PU.1), and E2A. Ikaros recruit the chromatin remodeling complexes to certain regions in the DNA. Ikaros also ensures the accessibility of the genes that are needed for the B-cell development. Low levels of PU.1 favors lymphoid differentiation, and cells with higher levels of PU.1 are bound for a myeloid fate. Gfi 1 downregulates the expression of PU.1 to the levels that are needed for the progression down the B-cell pathway. The E2A expression contributes to the maintenance of the HSC pool by participating in the regulation of cell cycle control in the population. HSCs express the cKIT (CD117) which is the cell-surface molecule and is the receptor for stem cell factor (SCF). SCF-c-Kit interaction is important for the development of MPPs. HSCs also express the stem cell associated antigen-1 (Sca-1). With the CLP stage, the progenitor still retains the potential to mature along the NK, conventional DC, or T-cell lineages. The signals that are received through IL-7R promotes the cell survival and enhances the production of EBF-1 and other transcription factors that are needed for later steps in the B-cell differentiation pathway. The IL-7R-mediated JAK-STAT pathway induces the up-regulaiton of the anti-apoptotic molecule Mcl1. This signalling also results in the upregulation of the C-myc and N-myc genes. These signal the proliferation of the later pro-B-cell stage. CLPs are c-Kit low, Sca-1 low, and IL-7R +. The myeloid potential is lost. The chromatin containing the immunoglobulin locus becomes increasingly accessible. The developing lymphocyte approaches the point at which it is committed to the B-cell lineage.

Which of the following polarizing cytokines would favor differentiation to TH2 cells?

IL-4

According to the clonal selection theory, all the immunoglobulin molecules on a single B cell have the same antigenic specificity. Explain why the presence of both IgM and IgD on the same B cell does not violate the unispecificity implied by clonal selection

IgM and IgD differ in their constant-region domains whereas antigen specificity is determined by the variable-region domains. Molecules of IgM and IgD that have different C domains but identical VH and HL domains are found on a given B cell; thus, the cell is unispecific although it bears two isotypes.

What is lineage commitment? What models have been proposed to explain this commitment?

Lineage commitment refers to how as thymocytes are being screened by their TCR affinity for self-antigens, they are also being guided in their lineage decisions. The positively selected DP thymocytes decide to be CD8+ cytotoxic T cell lineage or CD4+ helper T cell lineage. The lineage commitment requires changes to the genome organization and gene expression. This involves silencing of one coreceptor gene (CD4 or CD8), and the expression of genes associated with lineage function. The following models have been proposed to explain this commitment: the instructive model, the stochastic model, and the kinetic signaling model.

Immature B cells bearing potentially autoimmune receptors can be managed in three ways to minimize the probability of disease. Describe these three strategies, noting whether they are shared by T cell progenitors

One strategy is with clonal deletion. Clonal deletion is an apoptotic process. It removes B cells and T cells that express receptors of self before they become mature lymphocytes. Another strategy is with central tolerance. Central tolerance involves how B cells that express receptors of self within the bone marrow are lost. A third strategy is with light chain receptor editing. This entails that B cells that are autoreactive and reactivate RAG genes. These such B cells become anergic and do not respond to any antigens

What are Recent Thymic Emigrants (RTES)? Why are researchers interested in these cells?

Recent thymic emigrants (RTES) are not functionally mature as naive T cells in the periphery. These do not proliferate or secrete cytokines as vigorously in response to TCR stimulation. Researchers are interested in these cells because RTEs are seen as an important source of mature T cells in individuals who are lymphonenic. For example those who are undergoing chemotherapy, are aged, or are newborns

Define the following terms:

Self-MHC restriction - Both CD4+ and CD8+ T cells can recognize antigen only when it is presented by a self-MHC molecule b. Antigen processing - required to generate peptides that interact specifically with MHC molecules; sequence of events that lead to degradation of a protein antigen into peptides c. Endogenous antigen - produced within the host cell itself d. Exogenous antigen - produced outside of the host cell and enters the cell by endocytosis or phagocytosis

Growth of hematopoietic stem cells (HSCs) on stromal cells that express NOTCH ligand will drive the development of these multipotent cells to become

T cells

What is the difference between T-Dependent and T-Independent Ags? What B cell types mediate each response? Describe the different types of antigen signals for each antigen.

T-dependent responses are generated when protein antigens are recognized and require the participation of CD4+ helper T cells. This class of B-cell response is known as T-dependent response. It is mediated by B-2 B cells binding to the TD antigens. The T-dependent response requires two different signals. The first signal is generated when a multivalent antigen binds and cross-links membrane immunoglobulin receptors (mIg). The second signal is provided by an activated T cell. This activated T cell binds to the B cell through its antigen receptor and through separate interactions between CD40 on the B cell and CD40L (CD154) on the activated TH cell. The bound T cell then delivers cytokines and other signals to its partner B cell to complete the activation process. T-independent responses are directed toward multivalent or highly polymerized antigens. This does not require T-cell help. The antigens that serve such responses are known as T-independent antigens (TI antigens). One class of the TI antigens are seen with the lipopolysaccharide moiety of Gram-negative bacteria and how it interacts with the B cell via both mIg and innate immune receptors. These TI-1 antigens are mitogenic, meaning it induces proliferation, for most B cells at high concentrations. This is a result of their ability to bind to pattern recognition receptors on the surface of the B cell. But, when there are lower concentrations, they only activate B cells that bind antigen with their Ig receptors. TI-2 antigen are the other class of TI antigens. These are highly repetitive antigens that are not inherently mitogenic, but have the ability to cross-link a large fraction of the Ig receptors on the surface of a B cell allowing them to deliver an activation signal in the absence of T-cell helps. Many TI-2 antigens are specifically and covalently bound by the complement component C3d. Most T-independent responses are mediated by B-1 and marginal zone B-cell types.

Describe the phenotypic and functional differences between T1 and T2 immature B cells.

T1 immature B cells have the following markers: CD21-, CD23-, CD24+, CD93+, mlgMhi, and mlgD-/lo. T1 immature B cells percolate through the T-cell zone. The self-reactive T1 B cells can be eliminated via apoptosis because of the strong antigen signaling. The diacylglycerol concentrations increase. T2 immature B cells have higher levels of mlgD, CD21, and BAFF-R expression. These recirculate among blood, lymph nodes, and spleen, and can enter B-cell follicles and marginal zone. T2 is resistant to antigen apoptosis. The diacylglycerol concentration increases.

What are T3 B cells? What is one possible function of these cells that could help to prevent autoimmune reactions?

T3 B cells are primarily self-reactive and anergic. The T3 B cells are characterized to be CD93+ mIgMlow CD23. CD23 is a low-affinity receptor for some classes of immunoglobulin. Recently it has been indicated that the T3 population might be representative of B cells that have been rendered anergic by contact with soluble self antigen, but have not yet been eliminated from the B-cell repertoire. As a result of how they are anergic, the B cells can respond to receptor stimulation without the help or assistance of T-cells. As the excess self-antigens are absorbed, this could prevent autoimmune diseases.

TH CELL ACTIVATION GENES

TH CELL ACTIVATION GENES THcell activation is initiated by interaction of the TCR-CD3 complex with a processed antigenic peptide bound to a class II MHC molecule on the surface of an antigen-presenting cell. This interaction and the resulting activating signals also involve various accessory membrane molecules on the TH cell and the antigen-presenting cell. Interaction of a TH cell with antigen initiates a cascade of biochemical events that induces the resting TH cell to enter the cell cycle, proliferating and differentiating into memory cells or effector cells. Many of the gene products that appear upon interaction with antigen can be grouped into one of three categories depending on how early they can be detected after antigen recognition: ■ Immediate genes, expressed within half an hour of antigen recognition, encode a number of transcription factors, including c-Fos, c-Myc, c-Jun, NFAT, and NF-B ■ Early genes, expressed within 1-2 h of antigen recognition, encode IL-2, IL-2R (IL-2 receptor), IL-3, IL-6, IFN-, and numerous other proteins ■ Late genes, expressed more than 2 days after antigen recognition, encode various adhesion molecules These profound changes are the result of signal-transduction pathways that are activated by the encounter between the TCR and MHC-peptide complexes

Name one distinguishing feature of TI-1 and TI-2 antigens.

TI-1 antigens are mitogenic and induce activation through both the BCR and innate immune receptors. TI-2 antigens bind tightly to the complement components C3d and C3dg, and so are bound by both the BCR and the complement receptor CD21 (CR2).

At what age does the thymus reach its maximal size?

Teenage years (puberty)

Describe in brief the overview of B cell activation from naïve B cell in the T cell zone to proliferation, SHM and CSR in the germinal centers to differentiation of cells that circulate in the periphery.

The B cells that are engaged with the T-dependent activation first encounter the antigen-specific T cells that are outside of the B-cell follicles. Some of the activated B cells differentiate into antibody producing plasma cells within the primary foci that are present outside the follicles. These cells then migrate to the medullary cords of the lymph node or the bone marrow and continue to secrete antigen-specific antibodies. The other antigen activated B cells enter the follicle and then divide and differentiate. When the follicles fill with proliferating B cells, a germinal center develops. This is where follicular dendritic cells reside. The cell density here is quite high. Immunoglobulin genes undergo class switching within the germinal center. The u constant regions are replaced by constant regions of other isotypes. The variable regions is subject to somatic hypermutation. The mutated variable regions are subject to antigen mediated selection within the germinal centers. The low-affinity and autoimmune receptor-bearing B cells die. The B cells with enhanced receptors leave the germinal centers for the periphery.

Describe the general structure and probable function of the B-cell-coreceptor complex.

The B-cell coreceptor is a complex of three cell membrane molecules: TAPA-1 (CD81), CR2 (CD21), and CD19. Binding of the CR2 component to complement-derived C3d that has coated antigen captured by mIg results in the phosphorylation of CD19. The Srcfamily tyrosine kinase Lyn binds to phosphorylated CD19. The resulting activated Lyn and Fyn can trigger the signal-transduction pathways: In one pathway, Syk activates PLCgamma2 by tyrosine phosphorylation. PLCgamma2 then hydrolyzes PIP2, a membrane phospholipid, to produce the second messengers DAG and IP3. DAG and Ca2+ released by the action of IP3 collaboratively activate the PKC, which induces additional signal-transduction pathways. The activated receptor complex also generates signals that activate the Ras pathway. Activated Ras initiates a cascade of phosphorylations that culminates in the activation of transcription factors that up-regulate the expression of many genes

Describe the Clonal Selection hypothesis. How does clonal selection generate a primary and secondary response?

The Clonal Selection Hypothesis suggested that the receptor molecule on the lymphocyte surface and the antibody products secreted by that cell had identical antigen-binding specificities. The hypothesis also indicated that the stimulation of a single B cell would result in the generation of a clone of cells having the identical receptor specificity as the original cell. These clones would both migrate to and serve function within the secondary lymphoid organs. The daughter cells within each clone would be able to secrete large amounts of antibody to neutralize the pathogen. The progeny cells would remain viable within the organisms and available to neutralize a secondary infection by the same pathogen. Essentially the main predictions of events are as follows. The immature B lymphocytes have immunoglobulin (Ig) receptors on their cell surfaces. All receptors on a single B cell have identical specificity for antigen. Next, when the antigen is stimulated, the B cell will mature and migrate to the lymphoid organs and replicate there. The clonal descendants will have the same receptor as the parental B cell. The clonal descendents will also secrete antibodies with an identical specificity for antigen. Towards the end of the immune response, more B cells that have the receptors for the stimulating antigen will remain within the host that were present before the antigenic challenge. The memory B cells will be able to have an enhanced secondary response. The B cells with receptors for self antigens are deleted during embryonic development.

Although the majority of T cells in the body express ab TCRs, up to 5% of T cells express gd TCRs. Explain the difference in antigen recognition between these two cell types and their function.

The choice of a cell to become a gamma-delta TCR or alpha-beta TCR depends on when and how fast the genes that code for each of the four receptor chains successfully rearrange. In order for a cell to become an alpha-beta T cell, the cell has to generate a TCRB chain, and this depends on a single in-frame VDJ rearrangement event. In order for a cell to become a gamma-delta T cell, a thymocyte has to generate two functional proteins that depend on two separate in-frame rearrangements. It is more probable for alpha-beta TCR expression.

What is the role of NOTCH in the development of T cells from hematopoietic stem cells (HSCs)?

The commitment to the T-cell lineage is dependent on Notch, which is a receptor. Notch is associated with embryonic cell development. Notch binding can commit cells to the T lineage even if the thymus is not present. The T-cell precursors encounter the Notch ligands, after arriving in the thymus through the blood vessels at the corticomedullary boundary. The growth of hematopoietic stem cells on stromal cells that express Notch ligand drives the development of the multipotent stem cells to the T-cell lineage. Notch regulates the decision of a lymphoid precursor to become a T versus B lymphocyte. When an active version of Notch1 is overpressed in hematopoietic cells, T cells rather than B cells develop in the bone marrow.

IgG, which contains heavy chains, developed much more recently during evolution than IgM, which contains mi heavy chains. Describe two advantages and two disadvantages that IgG has in comparison with IgM.

The disadvantage of IgG compared with IgM are its lowest capacity to (1) agglutinate antigens and (2) activate the complement system, both of which are due to lower valency of IgG. While IgM is an indicator of a current infection, an IgG indicates a recent or past exposure to the illness. IgM is a temporary antibody that disappears within two or three weeks. It is then replaced by IgG which lasts for life and provides lasting immunity to the person.

What is the "1st checkpoint" in B cell development? When does it occur and what is the purpose of this checkpoint?

The first checkpoint in B cell development entails how the B-cell development is stopped and the cell is lost to apoptosis, if the pre-B-cell receptor cannot be displayed on the cell surface because of non-productive VHDJH gene rearrangements. The progress for the checkpoint is dependent on a signaling event through the pre-B-cell receptor. It is thought that this is mediated through interactions between arginine-rich regions within the non-immunoglobulin portion of the lambda 5 component of the surrogate light chain or in the CDR3 regions of some heavy chains with negatively charged molecules on the surface of the stromal cells.

What mechanisms generate the three hypervariable regions (complementarity-determining regions) of immunoglobulin heavy and light chains? Why is the third hypervariable region (CDR3) more variable than the other two (CDR1 and CDR2)?

The heavy chain and light chain subunits of antibodies both contain three regions called complementarity determining regions (CDR1, CDR2, and CDR 3 ). CDRs are responsible for the high specificity of the fragment antigen binding (Fab) of antibodies. CDRs are hypervariable among antibodies, allowing for an extensive range of antigens with which different antibodies can interact. During B-Cell Maturation, the gene for CDR3 of the heavy chain is formed when a random VH segment, and a random DH segment, and a random JH segment are joined together. A similar process occurs with only VL and JL segments to form CDR3 of the light chain. To further randomize CDR3, DNA breaks are introduced at the junction sites, leading to nucleotide addition or subtraction before the breaks are sealed. Three different mechanisms cause random nucleotide alteration at the junction sites (P-addition, N-addition and junctional flexibility). Another mechanism for Fab variability is somatic hypermutation. Somatic hypermutation occurs after a B-cell encounters its specific antigen and involves a greatly increased chance of mutation in all CDRs.

How do we delete thymocytes reactive to tissue-specific antigens in the medulla of the thymus?

The thymocytes can be deleted through Automine Regulator (AIRE) proteins. These proteins induce the expression of tissue-specific proteins in the thymic epithelial cells. Thus, new T cells can be screened against the antigens in a safe manner within the thymus. AIRE is expressed in the nucleus where it upregulates the expression of tissue specific antigens. The recognition of antigens with high affinity results in the deletion of self-reactive T cells to prevent the release of these cells into the periphery to provoke autoimmunity

Explain why we don't delete all cells we positively select? What are the current models to explain the thymic selection paradox?

This can be explained by the affinity hypothesis. The strength of the signal received is important. All of the TCRs are of one type that can recognize one peptide. The MHC class I molecules on the thymic epithelial cells have no affinity, low affinity, or high affinity for their peptide. The degree of selection for or against the CD8 SP T cells is determined. When double positive thymocytes receive low affinity signal, it is positively selected. When it receives high affinity signal, it is autoreactive and negatively selected. When there is no signal, death by neglect occurs. This can be also explained by the altered peptide model. The cortical epithelium induces positive selection. It makes peptides that are distinct from peptides made by other cells, including the thymic cells that induce negative selection. The thymocytes that are selected on the unique peptide/MHC complexes would not be negatively selected when they browse the medulla and other negatively selecting cells.

THYMINE SELECTION

Thymocytes undergo two selection processes in the thymus: (Both processes are necessary to generate mature T cells that are self-MHC restricted and self-tolerant.) ■ Positive selection for thymocytes bearing receptors capable of binding self-MHC molecules, which results in MHC restriction. Cells that fail positive selection are eliminated within the thymus by apoptosis. It ensures MHC Restriction & takes place in the cortical region of the thymus and involves the interaction of immature thymocytes with cortical epithelial cells. ■ Negative selection that eliminates thymocytes bearing high-affinity receptors for self-MHC molecules alone or self- antigen presented by self-MHC, which results in self-tolerance . It ensures Self-Tolerance. Thymocytes with highaffinity receptors undergo negative selection by an interaction with thymic stromal cells. During negative selection, dendritic cells and macrophages bearing class I and class II MHC molecules interact with thymocytes bearing highaffinity receptors for self-antigen plus self-MHC molecules or for self-MHC molecules alone. Cells that experience negative selection are observed to undergo death by apoptosis Early evidence for the role of the thymus in selection of the Tcell repertoire came from chimeric mouse experiments by R. M. Zinkernagel and his colleagues.

In 2007 it was discovered that ___________________ produce unique peptides that confer moderate affinity to the T cell receptor (TCR) on immature T cells as part of positive selection of CD8+ cells. The interaction of this altered peptide with the TCR elicits a survival signal in the immature T cell and promotes differentiation to a mature killer cell.

Thymoproteosome

What characterizes Treg populations? What mechanisms do Tregs use to inhibit the proliferation of other T cell populations?

Treg cells are regulatory T cells. These cells are a CD4+ subset that helps to quench adaptive immunity. This subset is characterized by the expression of FoxP3 transcription factor. Treg cells negatively regulate immune responses. They can develop in the thymus and appear to represent an alternative fate for autoreactive T-cells. These cells function to do the following: deplete the local area of stimulating cytokines, produce inhibiting cytokines, inhibit APC activity, and directly kill T cells. Induced Treg cells arise during the activation of T cells in the presence of TGF-beta. The TGF-beta induces FoxP3 master regulator, shifting activating cells into this subset. iTreg cells secrete IL-10 and TGF-beta to downregulate inflammation by inhibiting APCS and suppress other T-cell subsets.

Both T and B cells must undergo both positive and negative selection. However, B cells and T cells have some notable differences in selection. Briefly describe these differences in selection between T and B cells.

With B cells, the initiation of the development begins in the bone marrow. It undergoes positive selection. The MHC presented peptides are seen with negative selection. There is light/TCR-alpha chain allelic exclusion. With T cells, the initiation of the development begins in the thymus. It undergoes positive or negative selection. There are no MHC presented peptides with negative selection. There are multiple TCR-alpha chains. Initially, both the B cell lineages and T cell lineages start within the fetus and neonate. The gamma-delta T cells and the B-1 B cells then pursue into their individual niches. The differences arise within the ELP stage and the CLP stage. During these stages, the T-cell progenitors migrate out of the bone marrow to the thymus. The B-cell progenitors stay in the bone marrow.

Investigations suggest that [x] plays a key costimulatory role during the initiation of activation of T cells, while [y] plays a key role in maintaining the activity of already differentiated effector and memory cells

[x] → CD28 [y] → Inducible costimulator (ICOS)

Indicate whether each of the following statements is true or false. If you think a statement is false, explain why. (MHC)

a. A monoclonal antibody specific for BETA2-microglobulin can be used to detect both class I MHC K and D molecules on the surface of cells. TRUE b. Antigen-presenting cells express both class I and class II MHC molecules on their membranes. TRUE c. Class III MHC genes encode membrane-bound proteins. d. In outbred populations, an individual is more likely to be histocompatible with one of its parents than with its siblings. False. They are soluble proteins that do not function in antigen presentation. They include several complement components, TNF-alpha, and Lymphotoxin alpha. e. Class II MHC molecules typically bind to longer peptides than do class I molecules. TRUE f. All cells express class I MHC molecules. False. Most nucleated cells express this molecule, bu neurons, placental cells, and sperm cells at certain stages of differentiation appear to lack class molecules. g. The majority of the peptides displayed by class I and class II MHC molecules on cells are derived from self- proteins. TRUE

Indicate all of the following which would lead to T-cell anergy?

a. A naive T-cell interaction with a dendritic cell in the presence of CTLA-4 Ig. c. A naive T cell stimulated with antibodies that bind only the TCR

Which of the following conditions would lead to T-cell anergy?

a. A naïve T-cell interaction with a dendritic cell in the presence of CTLA-4 Ig. b. A naïve T cell stimulated with antibodies that bind both the TCR and CD28. c. A naïve T cell stimulated with antibodies that bind only the TCR. d. A naïve T cell stimulated with antibodies that bind only CD28

Activation and differentiation of B cells in response to thymus-dependent (TD) antigens requires TH cells, whereas the B-cell response to thymus-independent (TI) antigens does not.

a. Discuss the differences in the structure of TD, TI-1, and TI-2 antigens and the characteristics of the humoral responses induced by them. The B-cell response to thymus-dependent (TD) antigens requires direct contact with TH cells, not simply exposure to TH-derived cytokines. Antigens that can activate B cells in the absence of this kind of direct participation by TH cells are known as thymus-independent (TI) antigens. TI antigens are divided into types 1 and 2, and they activate B cells by different mechanisms. b. Binding of which classes of antigen to mIg provides an effective competence signal for B-cell activation? TI-2 antigens activate B cells by extensively crosslinking the mIg receptor. However, TI-2 antigens differ from TI-1 antigens in three important respects. First, they are not B-cell mitogens and so do not act as polyclonal activators. Second, TI-1 antigens will activate both mature and immature B cells, but TI-2 antigens activate mature B cells and inactivate immature B cells. Third, although the B-cell response to TI-2 antigens does not require direct involvement of TH cells, cytokines derived from TH cells are required for efficient B-cell proliferation and for class switching to isotypes other than IgM.

Although the five immunoglobulin isotypes share many common structural features, the differences in their structures affect their biological activities.

a. Draw a schematic diagram of a typical IgG molecule and label each of the following parts: H chains, L chains, interchain disulfide bonds, intrachain disulfide bonds, hinge, Fab, Fc, and all the domains. Indicate which domains are involved in antigen binding. Variable Region- At the end of the F(ab)'2 region is the part that contacts the antigen. This region is called the "variable" domain because when one compares the amino acid sequence of two different antibodies (which bind two different antigens), this region is "variable". The rest of the antibody molecule is relatively constant and are designated as "constant region". Intrachain disulphide - The light chain is divided into two regions and the heavy chain is divided into four regions. Each of these domains is approximately 110 amino acids long and they have an intrachain disulfide bond in the middle 60 amino acids. This structure is seen in members of the immunoglobulin gene superfamily. Interchain disulphide- Besides the intrachain disulfide bonds, there are also interchain disulfide bonds that link the two heavy chains together (this is in the hinge region) and also, interchain disulfide bonds that keep the heavy and light chains together in the Cl (constant light region) region. Fc-region - The fragment crystallizable region (Fc region) is the tail region of an antibody that interacts with cell surface receptors called Fc receptors and some proteins of the complement system. This property allows antibodies to activate the immune system. In IgG, IgA and IgD antibody isotypes, the Fc region is composed of two identical protein fragments, derived from the second and third constant domains of the antibody's two heavy chains Hinge region - provides flexibility to permit binding to different antigenic arrangements

Indicate whether each of the following statements concerning B-cell maturation is true or false. If you think a statement is false, explain why

a. Heavy chain VH-DH-JH rearrangement begins in the pre-B-cell stage. b. Immature B cells express membrane IgM and IgD. c. The enzyme terminal deoxyribonucleotidyl transferase (TdT) is active in the pre-B-cell stage. d. The surrogate light chain is expressed by pre-B cells. e. Self-reactive B cells can be rescued from negative selection by the expression of a different light chain. f. In order to develop into immature B cells, pre-B cells must interact directly with bone-marrow stromal cells. g. Most of the B cells generated every day never leave the bone marrow as mature B cells

Which of the following does not participate in the formation of antigen-antibody complexes?

a. Hydrophobic bonds b. Covalent bonds c. Electrostatic interactions d. Hydrogen bonds e. Van der Waals forces

Indicate whether each of the following statements is true or false. If you think a statement is false, explain why.

a. Indirect immunofluorescence is a more sensitive technique than direct immunofluorescence. TRUE b. Most antigens induce a polyclonal response. TRUE c. A papain digest of anti-SRBC antibodies can agglutinate sheep red blood cells (SRBCs). FALSE d. A pepsin digest of anti-SRBC antibodies can agglutinate SRBCs. TRUE e. Indirect immunofluorescence can be performed using a Fab fragment as the primary, nonlabeled antibody. TRUE f. For precipitation to occur, both antigen and antibody must be multivalent. TRUE g. Analysis of a cell population by flow cytometry can simultaneously provide information on both the size distribution and antigen profile of cell populations containing several different cell types. TRUE h. ELISA tests using chemiluminescence are more sensitive than chromogenic ones and precipitation tests are more sensitive than agglutination tests. FALSE Agglutination is more sensitive. i. Western blotting and immunoprecipitation assays are useful quantitative assays for measuring the levels of proteins in cells or tissues. j. Assume antibody A and antibody B both react with an epitope C. Furthermore, assume that antibody A has a Ka 5 times greater than of antibody B. The strength of the monovalent reaction of antibody A with epitope C will always be greater than the avidity of antibody B for an antigen with multiple copies of epitope C.

Indicate whether each of the following statements about the spleen is true or false. If you think a statement is false, explain why.

a. It filters antigens out of the blood. (T) b. The marginal zone is rich in T cells, and the periarteriolar lymphoid sheath (PALS) is rich in B cells. (F( MARGINAL ZONE IS RICH IN B CELLS, AND PALS IS RICH IN T CELLS)) c. It contains germinal centers. (T) d. It functions to remove old and defective red blood cells. (T) e. Lymphatic vessels draining the tissue spaces enter the spleen. (F (THE SPLEEN IS NOT SUPPLIED WITH AFFERENT LYMPHATICS)) f. Lymph node but not spleen function is affected by a knockout of the Ikaros gene. (F (IN ADDN TO BEING ESSENTIAL FOR THE FORMATION OF NK CELLS, IKAROS IS ALSO ESSENTIAL FOR THE FORMATION OF T AND B CELLS. THEREFORE, ITS KNOCKOUT WOULD PREVENT LYMPH NODES AS SERVING AS SITES FOR THE GENERATION OF ADAPTIVE IMMUNE RESPONSES))

Indicate whether each of the following statements is true or false. If you think a statement is false, explain why

a. Monoclonal antibody specific for CD4 will coprecipitate the T-cell receptor along with CD4. FALSE b. Subtractive hybridization can be used to enrich for mRNA that is present in one cell type but absent in another cell type within the same species. c. Clonotypic monoclonal antibody was used to isolate the T-cell receptor. TRUE d. The T cell uses the same set of V, D, and J gene segments as the B cell but uses different C gene segments. FALSE, IT USES DIFFERENT V AND C GENE SEGMENTS e. The alpha-beta TCR is bivalent and has two antigen-binding sites. FALSE f. Each ALPHA-BETA T cell expresses only one alpha-chain and one beta-chain allele. g. Mechanisms for generation of diversity of T-cell receptors are identical to those used by immunoglobulins. FALSE, bcs they are generally similar to those that generate antibody diversity, BUT somatic mutation does not occur in TCR genes, as it does in immunoglobulin genes. h. The Ig-Alfa/Ig-Beta heterodimer and CD3 serve analogous functions in the B-cell receptor and T-cell receptor, respectively.

Cells that can present antigen to TH cells have been classified into two groups—professional and nonprofessional APCs.

a. Name the three types of professional APCs. For each type indicate whether it expresses class II MHC molecules and a co-stimulatory signal constitutively or must be activated before doing so. - Dendritic Cells Class 2, costimulatory activity, activate naïve T cells - Macrophages- Must be activated (TLR signaling) before they express MHC class 2 or costimulatory membrane molecules such as CD80/86 - B cells- Always express MHC 2. Must be activated by PAMPS before expressing costimulatory molecules

Cells that can present antigen to TH cells have been classified into two groups—professional and nonprofessional APCs

a. Name the three types of professional APCs. For each type indicate whether it expresses class II MHC molecules and a co-stimulatory signal constitutively or must be activated before doing so. - Dendritic Cells Class 2, costimulatory activity, activate naïve T cells - Macrophages- Must be activated (TLR signaling) before they express MHC class 2 or costimulatory membrane molecules such as CD80/86 - B cells- Always express MHC 2. Must be activated by PAMPS before expressing costimulatory molecules

Molecules of the CD1 family were recently shown to present nonpeptide antigens.

a. What is a major source of nonpeptide antigens? Intracellular bacteria are a major source of nonpeptide antigens b. Why are CD1 molecules not classified as members of the MHC family even though they associate with Betta2- microglobulin? Members of the CD1 family associate with Beta2-microglobulin and have structural similarity to class I MHC molecules. They are not true MHC molecules because they are not encoded within the MHC and are on a different chromosome. c. What evidence suggests that the CD1 pathway is different from that utilized by classical class I MHC molecules? THE PATHWAY FOR AG PROCESSING TAKEN BY CD1 MOLECULES DIFFERS DROM THAT TAKEN BY MHC MOLECULES. A MAJOR DIFFERENCE IS THAT CD1 AG PROCESSING IS NOT INHIBITED IN CELLS THAT ARE DEFICIENT IN TAP, WHEREAS IN CLASS I MHC MOLECULES CANNOT PRESENT AG IN TAP DEFICIENT CELLS.

ANERGY

absence of the normal immune response to a particular antigen or allergen TH-cell recognition of an antigenic peptide-MHC complex sometimes results in a state of nonresponsiveness called clonal anergy, marked by the inability of cells to proliferate in response to a peptide-MHC complex. Whether clonal expansion or clonal anergy ensues is determined by the presence or absence of a co-stimulatory signal (signal 2), such as that produced by interaction of CD28 on TH cells with B7 on antigen-presenting cells. In the absence of a co-stimulatory signal, there is minimal production of cytokines, especially of IL-2. Anergy can also be induced by incubating TH cells with normal APCs in the presence of the Fab portion of anti-CD28, which blocks the interaction of CD28 with B7

Discuss the origin of the competence and progression signals required for activation and proliferation of B cells induced by (a) soluble protein antigens and (b) bacterial lipopolysaccharide (LPS).

activation of B cells by soluble protein antigens requires the involvement of TH cells. Binding of antigen to B-cell mIg does not itself induce an effective competence signal without additional interaction with membrane molecules on the TH cell. In addition, a cytokine-mediated progression is required for B-cell proliferation. Some bacterial cell-wall components, including lipopolysaccharide (LPS), function as type 1 thymus-independent (TI-1) antigens. Type 2 thymus-independent (TI-2) antigens are highly repetitious molecules such as polymeric proteins (e.g., bacterial flagellin) or bacterial cell-wall polysaccharides with repeating polysaccharide units. In the periphery, the antigen-induced activation and differentiation of mature B cells generates an antibody response. The antibody response to proteins and most other antigens requires TH cells. These are thymus- dependent or simply Tdependent (TD) responses. Responses to some antigens, such as certain bacterial cell-wall products (e.g., LPS) and polymeric molecules with repeating epitopes, do not require TH cells and are independent (TI) antigens. The vast majority of antigens are dependent.

Explain the difference between antibody affinity and antibody avidity. Which of these properties of an antibody better reflects its ability to contribute to the humoral immune response to invading bacteria?

affinity measures the strength of the noncovalent interactions between one antigen binding site and 1 epitope. Avidity measures the combined strength of multiple interactions between a multivalent antibody and antigen.

___________ is expressed in the nucleus of mTECs where its primary function is to upregulate the expression of tissue specific antigens (TSAs) to direct deletion of self-reactive T cells before they are released into the periphery

autoimmune regulator (AIRE)

Mechanisms of negative selection to ensure central tolerance of T cells include all of the following except

clonal selection

The majority of cells in the thymus

die from neglect in the cortex.

The frequency of somatic mutation in Ig genes is greatest during

generation of memory B cells

What are the two primary roles of the thymus?

is the site ofT-cell development and maturation. It is a flat, bilobed organ situated above the heart. Each lobe is surrounded by a capsule and is divided into lobules,which are separated from each other by strands of connective tissue called trabeculae.Each lobule is organized into two compartments: the outer compartment, or cortex, is densely packed with immature T cells, called thymocytes, whereas the inner compartment,or medulla,is sparsely populated with.thymocytes. FUNCTION of thymus: to generate and select a repertoire of T cells that will protect the body from infection. As thymocytes develop, an enormous diversity of T-cell receptors is generated by a random process that produces some T cells with receptors capable of recognizing antigen-MHC complexes.

Defne the following terms and give examples using the human HLA locus: polygeny, polymorphism, and codominant expression. How exactly does each contribute to ensuring that a diversity of antigens can be presented by each individual?

polygeny, different genes encode for different MHC molecules; refers to the presence of multiple genes for MHC I and MHC II molecules in the genome, encoding a set of structurally similar proteins with similar functions polymorphism, MHC polymorphism is the presence of multiple alleles of most of the MHC I and II genes in the human population codominant expression. occurs when two different alleles are inherited at the same genetic location (i.e, one allele from mom and a different one from dad), and the products of BOTH of the alleles are expressed. Codominance is the most common pattern in blood group genetics

Positive selection of T cells in the cortex of the thymus involves

selection of thymocytes that have intermediate affinity for self-MHC