Humoral Immune Responses: B Cell Activation and Antibody Production

Question 1

Naïve B cells express two classes of membrane bound B-cell receptors (BCR), [].

Answer 1

Naïve B cells express two classes of membrane bound B-cell receptors (BCR), immunoglobulin D and M (IgD and IgM).

•The antigen-binding regions of the IgD and IgM are identical; they differ in the constant region of the Heavy chain

Question 2

Antibodies can respond to…

Answer 2

…proteins, polysaccharides (abundant on the surfaces of many microbes), nucleic acids and lipids.

Question 3

T-dependent antigens are derived from [].

Answer 3

T-dependent antigens are derived from proteins.

• T cells are exclusively activated by –and therefore provide help to –protein antigens.
• The B cell is able to internalize the BCR-bound protein antigen, then process and present it on MHC-II to effector CD4 T cells.
• The T cell provides signals back to the B cell to aid in B cell activation, heavy chain isotype switching, and affinity maturation.
• T-dependent responses are primarily mediated by follicular B cells within the follicles of the spleen and lymph nodes.

Question 4

T-independent antigens are derived from polysaccharides, lipids and other non-protein antigens.

Answer 4

T-independent antigens are derived from polysaccharides, lipids and other non-protein antigens.

• Because T cells can only respond to protein peptides presented by MHC, by definition, they are unable to aid B cell responses to most non-protein B cell antigens.
• T-independent responses are primarily mediated by marginal zone B cells in the periphery of the splenic white pulp (where they encounter blood-borne microbes), and B-1 cells in the mucosal tissues.
• Generally, no antibody class switching occurs, thus these responses predominantly produce IgM antibodies.

Question 5

Ig-mediated signal transduction on a B cell requires the bringing together of two or more receptors (termed “cross-linking”). This can be accomplished when either:

Answer 5

(1) two or more antigen-bearing molecules form an aggregate, or
(2) when repeating “multivalent” epitopes on one antigen molecule bind to adjacent membrane Ig molecules on the same B cell

Question 6

Antigen-receptor binding occurs mostly via []; by themselves these forces are weak, and therefore require close proximity and an excellent molecular fit between epitope and receptor to be effective.

Answer 6

Antigen-receptor binding occurs mostly via non-covalent forces; by themselves these forces are weak, and therefore require close proximity and an excellent molecular fit between epitope and receptor to be effective.

Question 7

Affinity

Answer 7

Affinity of an antigen receptor is the strength of the bond between a single epitope and a single antigen binding site on the receptor. Low affinity antibodies are involved in weak binding and dissociate more readily than high affinity antibodies.

Question 8

Avidity

Answer 8

Avidity of a receptor is the strength of the multiple interactions between a multivalent antigen (containing numerous epitopes) and a bivalent or multivalent receptor (Fig. 9-3). If the receptor is bound to the cell surface, avidity means the sum of all interactions.

Question 9

Avidity/Affinity is what predominates in biological systems.

Answer 9

Avidity is what predominates in biological systems.

• Most antigens encountered in nature and presented to B cells contain multiple epitopes. Such an antigen could engage both antigen-binding sites on a bivalent receptor molecule.
• High avidity can supersede low affinity.

-For example, IgM is a low affinity antibody but because of its multivalency (10 antigen-binding sites in the pentamer) it binds antigen more effectively than IgG of higher affinity (but with only 2 antigen-binding sites)

Question 10

Membrane IgM and IgD on naïve B cells have short intracellular cytoplasmic tails unable to transmit activation signals. The receptors are non-covalently associated with [] and [] to form the B cell receptor (BCR) complex, similar to the CD3 complex and the T cell receptor.

Answer 10

Membrane IgM and IgD on naïve B cells have short intracellular cytoplasmic tails unable to transmit activation signals. The receptors are non-covalently associated with Igα and Igβ to form the B cell receptor (BCR) complex, similar to the CD3 complex and the T cell receptor.

•Cross-linking of immunoglobulin (Ig) receptors of B cells by antigen triggers biochemical signals that are transduced by the Ig-associated proteins Igα and Igβ.

Question 11

In addition to the BCR complex, B cells also express complement receptor 2 (CR2 aka CD21), which recognizes the C3d fragment that is the end product of C3 deposition of a pathogen surface.

Answer 11

In addition to the BCR complex, B cells also express complement receptor 2 (CR2 aka CD21), which recognizes the C3d fragment that is the end product of C3 deposition of a pathogen surface.

• CR2 (CD21) is present on the B cell in a complex that includes CD19 and CD81, which aid in enhancing signal transduction for B cell activation.
• B cell binding of C3d through CR2 greatly enhances B cell activation (Signal 2). This is another instance in which innate activation in response to the pathogen can help in the adaptive immune response that subsequently develops.

Question 12

3 Signals for B Cell Activation

Answer 12

1. Initial activation of a naïve B cell involves Ag-induced crosslinking/clustering of IgM & IgD BCR on the cell surface

•B cell epitopes come in many flavors and shapes

• protein, carbohydrate, lipid, nucleic acids etc…
• linear or discontinuous (3D)
  2. CR2 (CD21) binds to complement C3d on the pathogen surface

•CD19, CD81 associate with CR2

• signal transduction (phosphorylation of CD19)
  3. cytokines

Question 13

Consequences of B Cell Activation

Answer 13

• Make & secrete IgM Ab (not switched or hypermutated) – particularly if Ag is multivalent and activates C’ (short-lived plasma cells)
• Are ready to receive CD4 T cell signals (help) for further differentiation
• Change the expression of chemokine receptors in order to emigrate from a primary follicle into the T cell zone (CD4 cells do the same)

Question 14

T-B contact happens in [].

Answer 14

T-B contact happens in the LN or the white pulp of the spleen.

Question 15

tivated B cells initially proliferate and differntiate in the [].

Answer 15

Activated B cells initially proliferate & differentiate in the primary follicle and produce plasma cells which make IgM for rapid anti-pathogen response.

Question 16

3 Steps to T-B Cell Collaboration: Step 1

Answer 16

1. CD4 helper T cells and B cells are independently activated by a protein antigen in different areas of the lymphoid organ, and then migrate toward each other.

Question 17

3 Steps to T-B Cell Collaboration: Step 2

Answer 17

2. These T and B cells initially interact with each other outside the follicle in 2 phases:
   a. The newly activated B cells internalize immunoglobulin-bound antigen, and process and present it via the MHC-II pathway for presentation to CD4 T helper cells.
   b. The previously activated CD4 T cells express CD40L and secrete cytokines, which act on the B cell to promote proliferation and differentiation into antibodysecreting plasma cells.

Question 18

3 Steps to T-B Cell Collaboration: Step 3

Answer 18

Some activated “helped” B cells migrate back to the follicle, accompanied by their CD4 helper cells. These T cells are now known as follicular helper T cells (TFH). The B cells undergo a massive wave of proliferation to form a germinal center, where somatic mutation of the variable regions occurs, as well as Ig heavy chain isotype switching. These germinal center reactions are what make the Tdependent antibody response so specialized and effective

Question 19

Activated T cells downregulate [] (which promotes migration to T cell zones) and upregulate [] (which promotes migration to B cell follicles).

Answer 19

Activated T cells downregulate CCR7 (which promotes migration to T cell zones) and upregulate CXCR5 (which promotes migration to B cell follicles).

•Collectively, these cells meet at the edges of the follicle or in the interfollicular areas, where the T cell tries to find a B cell presenting its’ antigen.

Question 20

Activated B cells do the opposite: upregulate [] (to promote migration toward T cell zones) and downregulate [] (to promote migration out of the follicles).

Answer 20

Activated B cells do the opposite: upregulate CCR7 (to promote migration toward T cell zones) and downregulate CXCR5 (to promote migration out of the follicles).

•Collectively, these cells meet at the edges of the follicle or in the interfollicular areas, where the T cell tries to find a B cell presenting its’ antigen.

Question 21

CD4 helper T cells recognize processed and presented peptide antigens (on MHC-II) and costimulatory molecules on the B cell, leading to:

Answer 21

1. Further upregulation of CD40 ligand (CD40L) to engage CD40 on the B cell.
2. Secretion of cytokines that act on the B cell to promote activation and isotype switching.

Question 22

The initial T-B cell interactions occur at the edge of the lymphoid follicles, and result in the production of a low level of relatively [] antibodies. These plasma cells are generally short-lived and produce antibodies for a few weeks – an initial wave to fight the infection.

Answer 22

The initial T-B cell interactions occur at the edge of the lymphoid follicles, and result in the production of a low level of relatively low affinity antibodies. These plasma cells are generally short-lived and produce antibodies for a few weeks – an initial wave to fight the infection.

Question 23

Some of the T cells that interact successfully with B cells express high levels of the chemokine receptor [], which leads to their trafficking into the B cell- rich follicles. These cells are now referred to as [].

Answer 23

Some of the T cells that interact successfully with B cells express high levels of the chemokine receptor CXCR5, which leads to their trafficking into the B cell- rich follicles. These cells are now referred to as follicular-helper T cells (TFH).

•some of the B cells that interact successfully with T cells also migrate back into the lymphoid follicle where they begin to rapidly proliferate in response to further interactions with TFH cells (again – these cells must continue to be specific for the same antigen, although not the same epitope).

Question 24

This region of the follicle that contains actively proliferating B cells is known as the [], and is the site of antibody heavy chain [] and [] to create more effective, high affinity antibodies of the correct class for the type of pathogen being fought.

Answer 24

This region of the follicle that contains actively proliferating B cells is known as the germinal center, and is the site of antibody heavy chain (isotype) switching and affinity maturation to create more effective, high affinity antibodies of the correct class for the type of pathogen being fought.

Question 25

Heavy chain switching is induced by a combination of [] and [] mediated signals.

Answer 25

Heavy chain switching is induced by a combination of cytokine and CD40L mediated signals.

Question 26

[] – B cells that enter the germinal center (Spleen, LN).

Answer 26

Centrocytes – B cells that enter the germinal center (Spleen, LN)

Question 27

activation-induced cytidine deaminase (AID)

Answer 27

At the same time the enzyme activation-induced cytidine deaminase (AID) is involved in inducing point mutations in the antigen-binding regions (CDRs) of the molecule which results in somatic hypermutation.

Question 28

The take home lesson is that VDJ segments do not join to C region genes at the DNA level whether it is during classical gene re-arrangement or class switching. The joining of VDJ to C to form VDJC occurs only at the [].

Answer 28

The take home lesson is that VDJ segments do not join to C region genes at the DNA level whether it is during classical gene re-arrangement or class switching. The joining of VDJ to C to form VDJC occurs only at the RNA level.

Question 29

affinity maturation

Answer 29

• Affinity maturation is the process in which the affinity of the antibody to antigen is increased after prolonged or repeated exposure to the antigen. It is essentially a process in which the activated B cells attempt to make a better antibody.
• Affinity maturation occurs after the activated B cells and TFH have formed germinal centers. As these B cells rapidly divide, they randomly incorporate point mutations (also via the enzyme AID, see previous section) in a process termed somatic hypermutation.

Question 30

somatic hypermutation

Answer 30

•Somatic hypermutation results in clonal progeny with variations in the original immunoglobulin receptor as a result of these point mutations – this can result in B cells with widely variable affinity for the original antigen.

Question 31

follicular dendritic cells (FDC)

Answer 31

• After somatic hypermutation, B cells will undergo apoptosis unless they can still recognize antigen, which is presented to them by follicular dendritic cells (FDC).
• FDCs reside in the germinal center and capture and present antigen/antibody complexes (via Fc or complement receptors).
• Such complexes are formed during the early stages of the humoral response with the first waves of antibody produced.
• This antigen presentation by FDCs is extremely long-lived, allowing the process of affinity maturation to continue long after the initial infection is controlled.

Question 32

plasmablasts

Answer 32

•After they receive a survival signal in the germinal center, many of these B cells (often called plasmablasts) enter into the circulation and tend to migrate to the bone marrow, where they may survive for years, while continuing to secrete high affinity antibodies.

Question 33

memory B cells

Answer 33

• A small portion of these germinal center B cells do not differentiate into plasmablasts, but rather circulate through the blood and various tissues as long-lived memory B cells, able to respond to the next encounter with that antigen.
• Many memory B cells also reside in the bone marrow.

Question 34

T-independent humoral immunity

Answer 34

When Polysaccharides, lipids and other (non-protein) antigens elicit antibody responses without CD4 T cell help.

Question 35

T-independent antibody responses are very important in recognizing and binding repeating [] on [].

Answer 35

T-independent antibody responses are very important in recognizing and binding repeating polysaccharide structures on extracellular bacteria.