T & B Cell Activation

B cells require what from T cells

"help" or "second signal"

What is Ab feedback

regulation of humoral responses by Fc receptors

Examples of therapeutics that augment T cell response

Ipilimumab Cancer Melanoma

___molecule binding required for T cell activation

costimulatory

mantle zone

darker staining rim surrounding germinal center --> region of Ag presentation

When CD40L and cytokines from CD4 T cells present,

isotype switching occurs

Recruits ___ that mark proteins including signaling molecules for degradation

ubiquitin

In isotype switching VDJ region remains

unchanged (same antigen specificity)

CTLA-4 has ___ fold higher affinity for B7 than CD28

50

B7 on

APC

Does TCR have intrinsic signaling function

no

Signal transduction pathways result in translocation of active transcription factors to ___ which modulate gene expression

nucleus

Tyrosine kinases such as Lck add

phosphate group to tyrosine residues

Abatacept & Betalacept do what

Bind B7 on APC (block second signal of costimulation) through CD38 and blocks T cell activation used with autoimmunity

Examples of therapeutics that block T cell response

1. Abatacept (autoimmune arthritis) 2. Betalacept (kidney transplant rejection)

Naive T cells require what 2 signals for activation

1. Engagement of TCR/CD3 complex 2. Engagement of costimulatory molecules

Microbes that cross epithelial barriers activate dendritic cells (DCs) to:

1. Increase expression of co- stimulatory molecules B7-1 (CD80) and B7-2 (CD86) 2. Migrate to LN to present phagocytosed and processed protein antigens to T cells 3. Microbes that enter the circulation are captured by dendritic cells in the blood and spleen. Polysaccharide antigens are primarily presented in spleen and mucosal tissues.

These two signals lead to activation of three intracellular signaling pathways

1. PLC->increasedcalcium,calcineurin->NFAT 2. PLC->PKC->NFkB 3. Ras->MAPkinase->AP1

A 5 year old boy has a history of recurrent pneumococcal pneumonia, Pneumocystis carinii pneumonia and bacterial ear infections. He is found to have a mutation in CD40L resulting in a deficiency in expression of CD40L. Are the following statements true or false?

1. Serum contains mostly IgM and very little IgG. True 2. IgM antibodies are of lower affinity for antigen than those of a healthy individual. True 3. Lymph nodes lack well-developed follicles containing germinal centers. True 5. There is no evidence of somatic mutation of IgM variable regions. True False: 4. There is limited diversity generated by VDJ recombination in the repertoire of IgM antibodies. False

Describe Engagement of TCR/CD3 complex

1. TCR on T cell binding MHC: peptide on antigen presenting cell (APC)

CTLA-4 inhibits T cell responses by

1. competitively preventing CD28 binding to B7 2. removing costimulatory molecules from the APC cell surface

4. Which of the following descriptions about affinity maturation is correct? A. Depends on activation induced deaminase (AID)-mediated VDJ recombination B. Depends on activation induced deaminase (AID)-mediated point mutations of Ig V genes C. Depends on T follicular helper cell induced apoptosis of low affinity B cells D. Depends on antigen processing and presentation by dendritic cells within the germinal center E. Depends on the autoimmune regulator (AIRE) induced expression of self antigens in the germinal center

B. Affinity maturation occurs when B cells in the germinal center undergo AID- induced point mutations in Ig V genes, and then B cells whose mutated membrane Ig bind antigen with high affinity are selected by antigen recognition to survive. T follicular helper Copyright © 2015 by Saunders, an imprint of Elsevier Inc. Test Bank 6 cells do not induce B cell apoptosis, but rather provide signals that enhance B cell proliferation, somatic mutation, and survival in the germinal center. VDJ recombination is involved in producing functional Ig genes during B cell development, but it is not involved in germinal center reactions, and is not induced by AID. B cells that cannot bind antigen with high affinity die through a default pathway of apoptosis. Antigen processing and presentation by dendritic cells are required to generate helper T cells outside the follicle, but not to activate B cells in the follicle. Autoimmune regulator (AIRE) is involved in thymic expression of tissue antigens but is not involved in affinity maturation.

A 31 year old man from Phoenix presents to his primary care physician with a chronic cough. Laboratory evaluation reveals high titers of IgG specific for cocciodiomycosis. IgM specific for coccidiomycosis is not detectable. Which of the following is the most likely interpretation of the patient's laboratory results? A. Coccidiomycosis infection occurred within the past 2 weeks  B. Coccidiomycosis infection occurred more than 3 months ago C. Serologic evaluation was performed prior to seroconversion D. False positive IgG result from exposure to dust containing coccidiomycosis spores

B. Coccidiomycosis infection occurred more than 3 months ago Correct

major members of ___ and __ families involved in costimulation and regulation

B7 and CD28

What does Ipilimumab do?

Binds CTLA4 on T cell, blocking inhibitory signal & potentiating T cell response It is an: Anti CTLA 4 blocking mAB Keeps T cells turned on (increased T cell response) used in cancer

Upon encountering the appropriate viral peptide:MHC class I complex on an infected cell, A. B cell receptors become crosslinked and B cell activation occurs B. Naïve CD4+ T cells become activated C. Previously activated CD8+ T cells destroy the infected cell D. Naïve CD4+ T cells secrete IL-2

C. Previously activated CD8+ T cells destroy the infected cell - Correct A. B cell receptors become crosslinked and B cell activation occurs - Incorrect. B cell receptors do not recognize peptide:MHC B. Naïve CD4+ T cells become activated - Incorrect. Naïve T cells do not become activated, because the infected cells does not express costimulatory molecules and do not provide second signal for T cell actvation D. Naïve CD4+ T cells secrete IL-2 - Incorrect. IL- 2 secretion occurs after T cell activation, and infected cells do not activate naïve T cells.

Describe Engagement of costimulatory molecules

CD28 on T cell binding B7-1 (CD80) or B7-2 (CD86) on APC

Interaction of CD40 on B cell with _____ on T cell is required for germinal center formation, isotype switching, and affinity maturation

CD40L

3. All of the following protein-protein interactions are involved in activation of naive helper T cells by antigen-presenting cells (APCs) EXCEPT binding of: A. Peptide-MHC complexes on the APC to the TCR on the T cell B. CD4 on the T cell to non-polymorphic regions of class II MHC molecules on theAPC C. Integrins on the Tcell with adhesion ligands on the APC D. B7-2 on the APC with CD28 on the T cell E. CD40L on the T cell with CD40 on the APC

CD40L is not expressed on naive T cells and is only up-regulated subsequent to activation by an antigen-presenting cell (APC). In the naive helper T cell, the TCR binds to the MHC-peptide complex whereas the CD4 coreceptor engages a conserved region on the MHC II molecule. Integrins on the T cell interact with adhesion ligands on the APC. This region of binding between the T cell and the APC is known as the immunologic synapse and also includes costimulatory interactions, such as CD28 on the T cell binding to B7 on the APC.

B cell present antigen to activated ____ to recruit T cell help; CD4 T cell recognizes epitope from same protein

CD4T

CTLA-4 mediates trans-endocytosis: CTLA-4 expressing T cells capture and degrade ___from APC surface

CD80 and CD86 (activating)

Inhibitory signals such as ____ on the T cell binding B7-1 (CD80) or B7-2 (CD86) on the APC shut off the T cell response

CTLA4

Within 24-48 hr after T cell activation via TCR and CD28 stimulation, T cells express inhibitory molecule ___

CTLA4

what are inhibitors receptors of CD28 family

CTLA4 and PD-1

A 57 year old woman with metastatic melanoma is being treated with an anti-CTLA-4 monoclonal antibody. Which of the following best describes the mechanism of action of this drug? A. Suppress the immune response by blocking CTLA-4 interaction with CD80 (B7) B. Suppress the immune response by blocking the CTLA-4 interaction with CD28 between T cells and antigen presenting cells C. Prevent suppression of the immune response by triggering CD80 (B7) expression on antigen presenting cells D. Prevent suppression of the immune response by blocking the interaction of CTLA-4 with CD80 (B7)

D. Prevent suppression of the immune response by blocking the interaction of CTLA-4 with CD80 (B7) - Correct Anti-CTLA-1 monoclonal antibody called ipilimumab prevents suppressive signaling through CTLA-4 on T cells to potentiate T cell activity. A and B are wrong because anti-CTLA-4 does not suppress the immune response. Additionally, B is wrong because CTLA-4 and CD28 are both expressed on T cells and both of these bind CD80 (B7) on APCs. C is wrong because anti-CTLA-4 does not trigger CD80 on APCs. CD80 is induced on dendritic cells after activation CTLA-4 is induced on T cells after T cell activation.

Antibody responses against which of the following antigens will not be significantly affected in achild born with homozygous mutations in a gene required for T follicular helper cell differentiation? A. Tetanus toxoid B. Influenzahemagglutinin C. Beevenomproteins D. ABO blood group antigens E. Rh factor antigen

D. T cell-independent antigens include polysaccharides, glycolipids and nucleic acids with multiple repeated epitopes, which may cross-link the B cell receptor maximally and thus bypass the need for T cell help. Of the choices given, the best choice is the ABO blood group antigen, because of its polyvalent glycolipid structure. The other antigens are all proteins, antibody responses to which require T cell help, including help from T follicular helper cells.

1. Which one of the following statements about primary and secondary antibody responses is NOT true? A. Antibodies in primary responses generally have lower affinity for antigen than those produced in secondary responses. B. Secondary responses reach peak levels more quickly than primary responses. C. Primary responses require higher concentrations of antigen for initiation than secondary responses. D. Primary responses occur to all types of antigens, but secondary responses mostly occur only to protein antigens. E. Primary responses are characterized by IgG antibodies, whereas secondary responses are dominated by IgM antibodies.

E: In a primary immune response, IgM antibodies are initially produced against antigens. IgG production requires T cell-dependent isotype switching and is seen predominantly in secondary responses. Primary antibody responses can be mounted to any type of antigen, but secondary responses usually require CD4+ T cell help, and therefore the antigen must be a protein. Primary responses do require higher concentrations of antigen for initiation. The affinity of membrane Ig for antigen is lower on naive B cells, which are responsible for primary responses, compared with memory B cells, which are responsible for secondary responses. Secondary responses develop more quickly and produce higher peak levels of antibody as compared with primary responses.

The TCR on a naïve CD8 T cell binds to a viral peptide bound to MHC class I on an epithelial cell. What happens?

Epithelial cells do not express costimulatory molecules, so T cell does not receive second signal and does not get activated.

___: secreted late in primary infection and in secondary infection, most abundant isotype in blood

IgG

The normal B cell responses to polysaccharides are T- independent; they are limited to lower affinity __, activate complement pathway, eliminate the pathogen, but lack strong memory.

IgM

Without signals from CD4 T cells, B cells secrete

IgM

_____: isotype on the surface of mature, naïve B cells, secreted in early, acute primary infection

IgM

Since polysaccharides are T-independent antigens, how can you design a vaccine to generate high-affinity antibodies, isotype switching and robust memory?

In a conjugate vaccine, a small chemical (hapten), in this case a part of the polysaccharide, is conjugated to a foreign protein. Hapten-carrier not part of usual anti- bacteria response, but a way to generate vaccine. Hapten and carrier protein must be covalently attached. Since attached, they will both be at same site where B and T cell activation is occurring. B cells specific for polysaccharide will be adjacent to T cells specific for carrier protein. Protein-specific activated T cells will provide help to B cells in same area regardless of B cell specificity. Thus, polysaccharide-specific B cells will receive T cell help in this vaccine design.

A 50 year old man presents with his first episode of jaundice, nausea and vomiting. Two hepatitis A virus (HAV) serologic tests are available. Total anti- HAV antibodies determines the presence of IgM and IgG specific for HAV. Anti-HAV IgM specifically determines the presence of IgM specific for HAV. In our patient hepatitis A serology reveals total anti-HAV is positive and anti-HAV IgM is negative. u Is hepatitis A infection likely the cause of his current symptoms?

No. Patient has IgG, but not IgM antibodies specific for HAV. Therefore, the patient is immune to HAV from past infection or vaccination. HAV is not cause of current symptoms, and patient needs a work up for other causes of jaundice, nausea and vomiting. u Total positive, IgM positive: Acute infection, requires report and counseling or tested soon after vaccination u Total negative: not immune/susceptible, needs immunization

CD28 on

T cell

These recruit tyrosine phosphatases to terminate signal through

TCR and CD28 (remove phosphate)

affinity maturation

The increase in affinity of the antigen-binding sites of antibodies for the antigen that occurs during the course of an adaptive immune response.

what is mechanism for membrane vs. secreted IgM

alternate RNA processing

Germinal center reactions in T dependent

antibody

Switch regions (S) facilitate formation of DNA loops which are lost from genome -> after isotype switching B cell (can/cannot) go back to making IgM

cannot

Interaction of CD40 on B cell with CD40L on T cell is required for =

germinal center formation, isotype switching, and affinity maturation

Affinity maturation results from

hypermutation in Ig variable regions

Signaling via CD3 complex and z proteins which contain ___residues

immunoreceptor tyrosine- based activation motif (ITAM) tyrosine

Mice in which CTLA-4 gene is knocked out develop

massive accumulation of activated lymphocytes in lymphoid organs and severe systemic inflammatory reactions

change in Ab structure during isotype switching

same antigen specificity different constant region constant region is identical in all antibodies of same isotype

change in antibody structure during affinity maturation

same antigen specificity improved affinity for antigen same constant region

change in Ab structure during switch membrane --> secreted form

same antigen specificity same constant region lost transmembrane & cytoplasmic domains

These are also important ____

therapeutic agents

After having received the signal through the TCR and CD28, the CD8 T cell is activated. The TCR on the activated CD8 T cell binds to a viral peptide bound to MHC class I on an epithelial cell. What happens?

u Once activated, the CD8 T cell only needs to bind to MHC:peptide on the target cell. The CD8 T cell will kill the virally-infected epithelial cell.

2. Which one of the following statements about humoral immune responses is true? A. Naive B cells are required for initiation of primary responses and memory B cells are required for initiation of secondary responses. B. Antibody responses to bacterial polysaccharide antigens require CD4+ helper T cells. C. Heavy chain isotype switching typically occurs in response to bacterial polysaccharide antigens. D. Affinity maturation does not require helper T cells. E. Antibody-secreting cells generated during a humoral immune response live for only a few hours.

A. Humoral responses require antigen-dependent activation of B cells through binding of the antigen to membrane Ig on naive B cells or on memory B cells, for primary or secondary responses, respectively. In most cases, nonprotein antigens do not stimulate isotype switching or affinity maturation because these changes require T cell help, and only protein antigens can stimulate T cells. For both nonprotein and protein antigens, antibody- secreting cells that are generated may live for months, often in the bone marrow.

Which of the following membrane proteins on a B cell generate inhibitory signals that block proliferation and differentiation of the B cell? A. FcγRIIB B. TLR5 C. CR2/CD21 D. membrane IgM E. CD40

ANS: A. Antibody feedback is the mechanism of regulation of humoral immune responses and is mediated by the FcγRIIB receptor, which delivers inhibitory signals into the B cells on binding the Fc portion of IgG. Each of the other proteins listed generate B cell activating signals.

The cytoplasmic tails of the signaling polypeptides found in lymphocyte antigen receptor complexes (BCR and TCR) contain regions that become phosphorylated upon antigen recognition and then bind ZAP family kinases. These regions are called: A. Immunoreceptor tyrosine-based activating motifs (ITAMs) B. Fcregions C. Toll-like IL-1R (TIR) domains D. Complementarity determining regions E. Nod like receptor ( NLR) domains

ANS: A. ITAMs are found in the cytoplasmic tails of CD3 and ζ proteins of the TCR complex and Igα and Iβ proteins of the BCR complex. ITAMs contain pairs of tyrosine residues that become phosphorylated by Src family kinases after antigen recognition, and then serve as docking sites for ZAP70 in T cells or Syk in B cells. ZAP70 and Syk are tyrosine kinases that become active after binding to ITAMs and then phosphorylate adaptor proteins and other kinases.

The germinal center reaction generates long lived plasma cells which secrete protective high affinity B cells for many years. Where do these cells reside? A. Bone marrow B. Thymus C. Parathyroid D. Skin E. Splenic red pulp

ANS: A. Plasmablasts generated in the germinal center migrate to the bone marrow, and differentiate into long-lived plasma cells, which remain active antibody secreting cells for months to years.

6. The most important costimulators for naïve T cell activation are which of the following? A. ICOS Ligand B. B7-1 and B7-2 C. PD-L1 and PD-L2 D. OX40 -Ligand E. FAS Ligand

ANS: B. B7-1 (CD80) and B7-2 (CD86) are the major costimulators required for naïve T cell activation. They are expressed on mature myeloid DCs, and bind to CD28 on the T cells. ICOS-Ligand and OX40 ligand are costimulators for previously activated T cells; their receptors are not expressed on naïve T cells. PD-L1 and PD-L2 bind to PD-1 on T cells, which inhibits T cell activation. FAS Ligand binds to FAS on T cells, and induces apoptotic cell death.

4. Which one of the following statements about MHC-TCR interactions is NOT true? A. Antigen receptors on T cells bind to MHC molecules for only brief periods of time. B. The affinity of most TCRs for peptide-MHC complexes is similar to the affinity of antibodies for their antigens. C. Only 1% or less of the MHC molecules on any antigen-presenting cell (APC) display a peptide recognized by a particular T cell. D. T cells usually require multiple engagements with an APC before a threshold of activation is reached. E. A subthreshold number of MHC-TCR interactions can lead to T cell inactivation.

ANS: B. In general, the TCR binds to peptide-MHC complexes with lower affinity than antigen-antibody interactions. This relatively low-affinity interaction occurs briefly; thus, a T cell may need multiple engagements with the antigen-presenting cell (APC) before a threshold of activation occurs. If this threshold is not reached, the T cell may enter into an inactive state known as anergy. On any given APC, less than 1% of the MHC molecules display the same peptide.

13. A previously healthy young European woman who had not been immunized against measles virus became infected with the virus while traveling in Africa. She developed a rash, high fever, cough, and runny nose, and then fully recovered after 4 days. A blood test performed 2 months later, after her return to Europe, showed the presence of high affinity IgG antibodies specific for measles virus antigens. Which of the following events did NOT occur in germinal centers of this patient? A. Somatic mutation of Ig V genes B. Ig gene V(D)J recombination C. B cell proliferation D. T follicular helper cell interactions with B cells E. Generation of memory B cells

ANS: B. V(D)J recombination occurs in developing B cells in the bone marrow. Germinal centers are sites of differentiation of mature B cells, in response to T cell-dependent protein antigens. The B cell response to protein antigens involves helper T cell signals delivered to B cells via CD40 ligand and cytokines. This results in the initial B cell proliferation and antibody production. Some of the activated B cells move back into the follicle, where there is brisk proliferation of one or a few clones of B cells specific for the inciting antigen. Within the germinal center, specialized T cells called T follicular helper cells express cytokines and CD40 that are required for further B cell activation. The proliferating B cells undergo extensive isotype switching and somatic mutation of the variable genes, followed by selective survival of the high-affinity B cells. Some of these cells become antibody- secreting plasma cells, and others become memory B cells.

CTLA-4-Ig is a soluble recombinant protein containing the ligand binding portion of CTLA-4 fused to the Fc portion of IgG. It is an approved drug used to treat autoimmune diseases. How does it work? A. Binds to CTLA-4 on T cells and induces inhibitory signals B. Binds to CTLA-4 and blocks its ability to bind to B7-1 and B7-2 C. Binds to B7-1 and B7-2 on antigen presenting cells, and blocks their ability to bind to CD28 on T cells D. Binds to inhibitory Fc receptors on B cells and inhibits the production of autoantibodies E. Binds to cytotoxic T lymphocytes and blocks their ability to kill other cells

ANS: C. CTLA-4-Ig is a costimulatory blocker. It has high affinity for B7-1 and B7-2, and therefore blocks costimulation by these molecules, which is required for naïve T cell activation. CTLA-4-Ig is not anti-CTLA-4 and does not bind to CTLA-4. The Ig Fc portion of the drug prolongs half-life, but is not involved in the immune inhibitory function of the drug.

18. Which of the following is NOT true about heavy chain isotype switching? A. It is induced by recombination of Ig constant region DNA segments B. It is defective in individuals lacking functional CD40 or CD40-ligand C. Itisprominentinantibodyresponsestopolysaccharides D. It generates antibodies with diverse effector functions E. Is requires AID, the enzyme that is also involved in somatic mutation of Ig V genes

ANS: C. Isotype switching requires signals from helper T cells, so it is most prominent in antibody responses to protein antigens and not T-independent polysaccharides. The molecular mechanism does involve "switch recombination" of Ig gene segments, which is stimulated by CD40L-CD40 interactions and AID. Antibodies of different ispotypes serve distinct functions.

8. At the peak of a CD8+ T cell response to a new microbe, what is the fold increase in the number of microbe-specific T cells in an individual compared to the number of naïve CD8+ T cells specific for the microbe before infection? A. ~100 B. ~1000 C. ~10000 D. ~100,000

ANS: D. CD8+ T cells undergo tremendous clonal expansion in response to antigen stimulation. CD4+ clonal expansion is less, on the order of 1000 to 10,000 fold.

9. Which of the following is an example of G protein-coupled receptor (GPCR)? A. T cell antigen receptor (TCR) B. The B cell antigen receptor (BCR) C. CD4 D. Interleukin-2 (IL-2) receptor E. Chemokine receptor CCR7

ANS: E. All chemokine receptors, including CCR7, are GPCRs. The lymphocyte antigen receptors (TCR, BCR), CD4, and IL-2 receptor signal via associated proteins or non- receptor tyrosine kinases.

12. A previously healthy young European woman who had not been immunized against measles virus became infected with the virus while traveling in Africa. She developed a rash, high fever, cough, and runny nose, and then fully recovered after 4 days. A blood test performed 2 months later, after her return to Europe, showed the presence of high affinity IgG antibodies specific for measles virus antigens. Which of the following proteins was NOT necessary for the production of these high affinity IgG antibodies? A. CD40 Ligand B. Activation induced deaminase (AID) C. Class II MHC D. CXCR5 E. C reactive protein (CRP)

ANS: E. CRP, an acute phase reactant induced by the liver in response to innate inflammatory cytokines, has little influence on B cell activation. All the other proteins listed are essential for the germinal center reaction that generates B cells producing high affinity IgG antibodies. CD40 ligand expressed on helper T cells binds to CD40 on B cells in the germinal center, providing signals that induce expression of AID, which is required for both class switching from IgM to IgG (and other isotypes). AID is also required for somatic hypermutation of Ig variable genes, leading to increased affinity of the antibodies produced in response to an infection. Class II MHC is essential for a B cell to present protein antigens to and receive helper signals from CD4+ T cells, which are required for isotype switching and affinity maturation. CXCR5 is a chemokine required for attracting B cells and T follicular helper cells into the germinal center in response to the chemokine CXCL13.

11. In the T cell calcium signaling pathway induced by antigen recognition, which enzyme and transcription factor are activated? A. IκB kinase, NFκB B. Jun kinase (JNK), AP-1 C. Erk kinase, Fos D. JAK3, STAT4 E. Calcineurin, NFAT

ANS: E. In the calcium pathway, TCR signaling leads to an increase in cytosolic calcium ion (Ca++) concentration, which activates the phosphatase calcineurin, leading to dephosphorylation of the transcription factor NFAT. Dephosphorylated NFAT enters the nucleus and stimulates transcription of various genes that promote T cell proliferation and differentiation. Calcineurin inhibitors, such as cyclosporin and tacrolimus, are widely used as immunosuppressant drugs for transplant recipients

5. Most naïve T cell activation occurs where and in response to what? A. In the skin in response to cytokines B. In the infected tissues in response to antigen presentation by macrophages C. In the blood in response to antigen presentation by monocytes D. In the thymus in response to antigen presentation by thymic medullary epithelial cells E. In secondary lymphoid organs in response to antigen presentation by dendritic cells

ANS: E. Naive T cells home to lymph nodes, and dendritic cells (DCs) carrying antigens from sites of infection drain into lymph nodes. The chemokine CCR7 directs the co- localization of both cell types. Other types of antigen presenting cell do not activate naive T cells efficiently because they do not migrate to where naïve T cells are, and/or they do not express the costimulatory molecules required for naïve T cell activation.q

The TCR on a naïve CD8 T cell binds to a viral peptide bound to MHC class I on an activated dendritic cell which also expresses B7-1 (CD80) and B7-2 (CD86). What happens?

Activated dendritic cells express MHC and costimulatory molecules, so T cell receives two signals and is activated.