BIO 432                                                                      Name_______KEY__________

Introduction to Immunology

Exam 1

 

        Before beginning, please verify that you have 8 pages.

        Please read each question carefully before answering.

 

 

I.               Short Answer (52 points):  General Concepts in Immunology and Cells;

Organs of the Immune System; Antigenicity and Immunogenicity

 

A.   (6 points) Discuss the difference between the two arms of the immune system- innate and adaptive immunity by addressing the differences in the following characteristics:

a.     Response time

b.     Specificity for antigen recognition

c.     Response to repeat infection

 

 

 

The above table is from your textbook and contains the information that should have been covered in the answer to this question.

 

B.    (5 points) Collaboration of innate immunity and the adaptive immune response yields a strong defense against infection.  Discuss how the subclass of pattern recognition receptors, the Toll-like receptors, link innate immunity to the adaptive immune response.

 

Toll-like receptors (TLRs) play an essential role in the recognition of microbial components and the presentation of information to the immune cell about the precise nature of a pathogen.  TLRs are either soluble or cell-associated receptors.  TLRs recognize conserved molecular patterns (pathogen-associated molecular patterns) that are shared by large groups of microorganisms. TLRs may also recognize endogenous ligands induced during an inflammatory response.  Recognition of conserved molecular patterns by TLRs can lead to the activation of intracellular signaling pathways within antigen presenting cells. These intracellular signaling pathways often result in an increase in cytokine secretion and an upregulation of MHC and costimulatory proteins on the surface of the APC.  Increases in MHC molecules and cytokines enable APC to readily activate T cells and stimulate an adaptive immune response to infection.  Cytokine secretion also increases the speed and effectiveness of an inflammatory response at the site of injury.  The ability of TLRs to stimulate both an inflammatory response and activate an antigen-specific adaptive immune response enables these receptors to provide a direct link between innate and adaptive immunity.

 

C.    (12 points) Phagocytosis is an important function of innate immunity and is a necessary precursor to antigen presentation by antigen-presenting cells.  Use the illustration from your text shown below to answer the following questions

  1. List and briefly describe the five steps of phagocytosis.

 

1. Bacterium is attached to pseudopodia

2. Bacterium is ingested into the phagosome.

3. Phagosome fuses with the lysosome forming the phagolysosome.

4. Lysosomal enzymes digest the bacterium.

5. Digestion products are released from the cell.

 

 

  1.  Which cells of the innate immune system are phagocytic?

 

Tissue macrophage, neurotrophils

blood monocytes

(2/3 accepted for credit)
 

5.
 

4.
 

3.
 

2.
 

1.
 

 

 

 

 

 

D.   (8 points)  The adaptive immune response depends on the function and interaction of lymphocytes and antigen-presenting cells.  Provide the following information for lymphocytes listed above by completing the table:

 

 

 

B lymphocyte

T lymphocyte

Location of Origin

Bone Marrow

Bone Marrow

Site of Maturation

Bone Marrow

Thymus

Receptor used for Antigen Recognition

Antibody, Immunoglobulin, BCR

T Cell Receptor, TCR

 

 

Name two of the three types of antigen presenting cells.

1. Macrophage

2. Dendritic cell

 

                                                                                                                                                                                                                                               

  1. (12 points) Antigen presentation to resting T cells results in activation and clonal expansion of T cells.  Answer the following questions concerning T cell activation and function.

 

1.     Two membrane adhesion molecules on T cells regulate the type of MHC molecules recognized by T cells.  What are these two adhesion molecules?

 

                 

                  CD8 and CD4

 

 

2.     It is usually the case that T helper cells and T cytotoxic cells express only one of the membrane adhesion molecules listed above.  Indicate which of these adhesion molecules is most often expressed by T helper cells or T cytotoxic cells.

 

 

                        T helper= CD4

 

 

                        T cytotoxic= CD8

 

 

 

 

 

3.     Which class of MHC proteins is usually recognized by T helper cells and which is usually recognized by T cytotoxic cells?  How does the mode of antigen entry into an antigen presenting cell influence the antigenic peptide-MHC complex that is presented on the surface of the antigen presenting cell?

 

T cytotoxic cells recognize antigen presented on class I MHC molecules. These antigenic peptides are generated by the digestion and presentation of endogenous proteins in the cytoplasm of the cell.  Self-antigens, viral proteins, and proteins of cancerous cell are examples of endogenous proteins whose antigens are presented on class I MHC.

 

T helper cells recognize antigen presented on class II MHC molecules.  These antigenic peptides are generated as exogenous antigens are endocytosed or phagocytosed and then processed via the endocytic pathway of antigen presenting cells.

 

 

4.     Briefly describe the specific functions of T helper cells and T cytotoxic cells once they are activated by an antigen presenting cell or stimulatory cytokines.

 

Recognition of antigen by T helper cells can generate the activation of two types of response pathways.  TH1 response produces cytokines that stimulate inflammation, activates T cells (other T helper cells and T cytotoxic cells) and tissue macrophages.  The TH2 response involves the secretion of cytokines that activates B cells and immune responses involving antibodies.  T cytotoxic cells activated by binding to class I MHC-peptide, in the presence of cytokines, differentiate into CTL (cytotoxic T lymphocytes) that can directly lyse altered self-cells.

 

 

  1. (5 points)  White blood cells (leukocytes) differentiate from both myeloid and lymphoid progenitors.   Specific combinations of extracellular (ex. cytokines in HIMs) and intracellular (ex. genes) mechanisms control the specific cell type derived during differentiation. Differentiated leukocytes are functionally diverse. 

Place the number (s) of the leukocyte that is best described by each statement.

           

                        1. Natural killer cell

                        2. Neutrophil

                        3. Basophil

                        4. Dendritic cell

                        5. Macrophage

 

 

__3_____        granulocyte that binds IgE with its high affinity Fce

receptor

 

__1_____        participates in ADCC by recognizing decreased levels of class I MHC on target cell

 

__5_____        antigen presenting cell whose phagocytic activity can be enhanced dramatically by an opsonin

 

__2_____        granulocyte possessing a bilobed  multilobed nucleus and is the primary cell type involved in leukocytosis

 

__4____          differentiates from either a myeloid and lymphoid progenitor and functions as an antigen presenting cell when mature

 

 

I.      ( 4 points)  During inflammation, a Langerhans cell has recognized and phagocytosed an invading bacterium.  This Langerhans cell migrates to the nearest lymph node, differentiates into a mature dendritic cell, and activates a nearby T helper cell with its peptide-class II MHC complex.  Describe how this activated T helper cell might influence the structure and function of a nearby primary follicle.

 

Activated T helper cells secrete cytokines that can activate nearby B cells.  In the primary follicle, small resting B cells are surrounded by a network of follicular dendritic cells that regulate B cell function.  After antigen challenge, such as the recognition of antigen or via activation by T helper cells (example above), the primary follicle begins to enlarge as it forms a  secondary follicle. The secondary follicle is comprised of a ring of concentrically packed B lymphocytes surrounding a germinal center of proliferating B cells, nondividing B cells and some T helper cells, macrophages, and follicular dendritic cells.  Antigen-activated B cells proliferate and generate antibody.  Some of these activated B cells migrate to the germinal center and undergo mitosis and selection. VThose B cells that survive selection in the germinal center produce plasma cells generating high affinity antibodies and memory B cells that respond rapidly to a second encounter with the antigen.

 

 

 

 

 

 

 

II. Fill-in-the-Blank (14 points, 2 points each)

Please place the term on the line that is best described by the definition to its right.

 

A. conjuctivae                   mucous membrane lining the inner surface of the eyelids

 

B. extravasation                term used to describe the migration of a phagocyte   through the endothelial cell wall tissue

 

C. hematopoiesis              developmental process leading to the generation of red and

                                          white blood cells

 

D. spleen                           secondary lymphoid organ functioning in filtering dead or

                                          damaged erythrocytes  and antigen from the blood

 

E.immunogenicity             the ability of an antigen to induce a humoral or cell-mediated immune response

 

F. hapten                           low molecular weight molecule that can be immunogenic

                                          when conjugated to a suitable carrier

 

G. adjuvant                       substances when mixed with antigen and injected into an

                                          organism enhance immunogenicity of the antigen

 

 

III. Short Answer  (34 points): Antibody Structure and Function; Organization and Expression of Immunoglobulin Genes

 

  1. (6 points) Immunoglobulin structure contains two heavy and two light chains.  Please answer the following questions concerning immunoglobulin structure.

 

What are the five classes of heavy chains?

 

                                m , g,  d,  a,  e

 

What are the two classes of light chains?  

                                                                     k,  l

 

What two forces stabilize an immunoglobulin fold domain?

 

Hydrophobic interactions and a disulfide bond

 

                  Which immunoglobulin heavy chain(s) lacks a hinge region?     m, e

 

 

 

 

  1. (4 points) What is the significance of complementarity determining regions

(CDR’s)?   

CDR’s are sites of hypervariability important for the diversity and specificity required for immunoglobulin antigen recognition

 

Where are CDR’s located? 

 

                        Variable regions of light and heavy chains

 

How many CDRs are present on an IgG molecule?

 

                                    12

 

 

  1. (4 points)  Why is an antibody that binds a single nucleotide unlikely to bind double-stranded DNA?

                                                       

                                                              shape of the binding site - pit vs groove

 

 

  1. (4 points) During the cleavage phase of V(D)J recombination, RAG proteins function to introduce DNA double strand breakage through three main stages (synapsis, nicking and hairpin formation).   How do RAG proteins distinguish between different gene segments to generate the coding segments that will be brought together during the joining phase?

 

Based on the recombination signal sequence: each type of segment is flanked by one form of RSS - either a 12-RSS or a 23-RSS

 

 

  1. (6 points) List the three stages of the joining phase of V(D)J recombination. 

 

1. Opening of DNA hairpins: hairpin intermediates resolved by Artemis/DNA-PKcs

2. Nontemplated (N) nucleotide addition: TdT tails coding DNA, optional and restricted to H chain

3. DSB repair: signal ends ligated and coding ends joined, involves Ku70/Ku80 and XRCC4/DNA Ligase IV

 

Coding joints formed during the joining phase are formed through “imprecise joining”.  Discuss one benefit and one drawback to “imprecise joining”.

                        Benefit- diversity

            Drawback- possible generation of a stop-codon and unproductive V(D)J recombination

 

 

  1. (6 points) Below is a list of all sources contributing to antibody diversity.   Fill in the table below and indicate by writing “yes” or “no” across from the source whether each source contributes to antibody specificity.

 

            Sources                                                              Impact antigenic specificity?

            Multiple V, D and J segments                                                Yes

            Combinatorial rearrangement                                      Yes

            Junctional diversity (N,P, exonuclease)                      Yes

            Random H and L chain pairing                                   Yes

            Somatic Hypermutation (SHM)                                 Yes

            Allotypic variation                                                     No

            Class switch recombination (CSR)                             No

            Alternative RNA splicing                                           No

 

            Which of the above occur after antigenic stimulation?

 

                        SHM, CSR, and Alternative RNA splicing

 

  1. (4 points) What two primary forces help ensure that the B cell receptors on a given B cell have only one antigenic specificity?

 

 

            Gene rearrangement is developmentally and temporally ordered: i.e. H before L, and only one chromosome rearranged at a time

 

 

 

IV. Bonus (10 points)

 

 

    1. (2 points)  Mammals are thought to have evolved bone marrow as the site for the generation of lymphocytes from a condition found in jawless fish such as the lamprey.  Name the site of hematopoiesis and lymphocyte generation in lamprey.  What was the first group of vertebrates to develop bone marrow as the site of lymphocyte generation?

 

GALT, amphibians

 

 

    1. (2 points)  DiGeorge’s syndrome is a congenial defect resulting in absence of a thymus.  Briefly describe how this condition influences the function of the adaptive immune response.

No T cells would be produced. The individual would suffer from  severe immunodeficiency due to the lack of cell-mediated adaptive immune responses.

 

    1.  (2 points)  Why is the Ig-a/Ig-b heterodimer necessary for the function of a mature B cell?

 

This heterodimer contains the cytoplasmic domain required for BCR intracellular signaling.

 

 

4.  (4  points) You are analyzing the immunoglobulin response to immunization in mice by examining the sequence of rearranged immunoglobulin V genes in B cell clones you have obtained.  In one clone, you have derived the putative IgH V, D and J germline gene segment usage by sequence comparison.  Shown are the coding regions of the segments (RSSs would be lost after V(D)J recombination).  Align the germline gene segments (top) to the rearranged gene (bottom) and explain how the coding junctions present might have been formed.  Note: only the top strand of DNA is shown.

 


 

 

 

 


                             

 

 

 

 

 

 

 


    N    P                                               Exonuclease