"Microbiology/Immunology Development of B Cells" MARY

Question 1

T/F: The clonal selection theory states that each naive B-cell produces an immunoglobulin of unique specificity.

Answer 1

True. ONE antibody produced by ONE B-cell, if it survives the process of making the antibody at all.

Question 2

The J region is also known as:

Answer 2

the Joining gene segment, part of the gene “choices” for the developing B cell’s heavy chain (Hc).

Question 3

The D region is also known as:

Answer 3

The Diversity segment, part of the gene “choices” for the developing B cell’s heavy chain (Hc).

Question 4

The components of Hcs (heavy chains) consist of what segments?

Answer 4

V (variable), D (diversity), J (Joining) and C (constant).

Question 5

What special enzymes bring together the V, D and J regions in the creation of the Hc?

Answer 5

The recombinase genes Rag 1 and Rag 2. This is important.

Question 6

T/F: Every heavy chain has a signal sequence section that can join to every light chain that is made, helping to contribute to the large diversity of immunoglobulin products.

Answer 6

False! Each immunoglobulin chain has signal sequences, and those signal sequences determine which other chains it can match to. If a maturing B-cell has made a Hc that does not match the Lc it has made, the B-cell is useless and commits suicide.

Question 7

Give the overall steps in the maturation of B-cell, from early pro-B-cell to immature B-cell.

Answer 7

1. Early pro-B-cell - rearrangement of D and J regions on both chromosomes.
2. Late pro-B-cell - H chain rearrangement –> V-DJ rearrangement on 1st chromosome
   2a. If successful, becomes pre-B-cell
   2b. If Step 2 unsuccessful –> V-DJ rearrangement on 2nd chromosome
   2c. If successful, becomes pre-B-cell. If unsuccessful, apoptosis
3. Pre-B cell - rearrange K on 1st chromosome –> if successful, done! and B-cell expresses miu and k, IgM is done!
   3a. If unsuccessful - rearrange K on 2nd chromosome –> if successful, done! and B-cell expresses miu and k, IgM is done!
   3b. If still unsuccessful with both k’s, pre-b-cell rearranges lambda on 1st chromosome, if successful, B cell shows IgM with miu and lambda!
   3c. If still unsuccessful, pre-b-cell tries to rearrange with lambda on 2nd chromosome, if successful, B cell shows IgM with miu and lambda!
   3d. A tired and discouraged pre-b-cell commits suicide if neither k’s nor lambdas fit.

Question 8

Joining of D and J is imprecise, and there is also a special mechanism to increase diversity at the CDR3 site. Explain the steps which create junctional diversity, starting with the cleaving of the RSS (recognition signal sequence) heptamer.

Answer 8

1. The cleavage of both D and J region heptamer RSS’s by RAG complex creates DNA hairpins.
2. RAG then opens the hairpins on the extruding DNA, yielding “P” palindromic sequences.
3. TdT (terminal deoxynucleotidyl transferase) adds N nucleotides are added randomly to spice things up.
4. The strands with their P section and new N section pair.
5. The non-matched N nucleotides are removed by an exonuclease.
6. Any remaining gaps (for example, over the P section) are filled in by DNA synthesis and ligation, yielding a unique P-N-P junctional site.

Question 9

Where are N nucleotides found in antibodies?

Answer 9

Only in heavy chains, and they are only found in the developing B-cell, up to the pro-B-cell stage.

Question 10

T/F: Joining of D to J and V-D in Hc gene formation can involve multiple reading frames, as most Ds can be read in all three reading frames.

Answer 10

True!

Question 11

A B-cell that has never encountered its cognate antigen is considered:

Answer 11

Naive or virgin. A B-cell that has met and joined with its cognate antigen is considered “experienced” ;)

Question 12

What are the three routes a B-cell can pursue if it matures?

Answer 12

1. Class switching
2. Somatic hypermutation
3. The “career decision” when a B-cell can become an antibody factory (plasma cell) or a memory cell.

Question 13

What determines the class of an antibody?

Answer 13

The constant region of its heavy chain (Fc), the “tail” of the Ab. *Therefore, class switching involves the changing of this constant region.

Question 14

When naive B-cells are first activated, they mainly make antibodies of what class?

Answer 14

IgM, probably the first class of Abs to evolve, even found in lower vertebrates.

Question 15

What might be the advantage to having IgM be produced first in an infection?

Answer 15

Because IgM is like 5 IgGs together, a big snowflake of a molecule, it has a higher chance of activating the complement cascade or “fixing complement.” IgM does this through bringing C1 complexes close together so their inhibitors fall off and they can be activated (complement chain reaction of sorts on the surface of the invader).
Additionally, IgMs can bind to virtually any surface of a virus, preventing its entry into cells.

Question 16

What are some advantages that IgG antibodies have over IgM?

Answer 16

1. Subclasses of IgG, based on the Fc region, that further specialize their function
2. Can cross the placental barrier and provide fetus with early protection.
3. Longer half life than IgM (3 weeks vs. 1 day)

Question 17

IgG antibodies are also called:

Answer 17

gamma globulins

Question 18

What is generally considered the most abundant class of antibody in the human body?

Answer 18

IgA! IgG is usually the most abundant class in the blood, however IgA guards the mucosal surfaces (digestive, respiratory, reproductive) of the body and exists in abundance, more than all other classes combined. 
IgA is also helpful to the neonate because it is secreted in breast milk.

Question 19

What is the main influence of class switching for B-cells?

Answer 19

Certain cytokines or combinations of cytokines influence a B-cell to switch from one class to another.

Question 20

How can the right cytokines “magically” be present for a B-cell to get the correct class-switching instructions?

Answer 20

T-helper cells make these cytokines, more on this in the next lecture.

Question 21

What is different between the B-cell that is displaying its winning Ab and the Ab it secretes?

Answer 21

As the B-cell becomes an antibody factory, the transcripts exclude the transmembrane domains so the Ab can be free in the blood.

Question 22

Conversion to what class of antibody does not involve class switching from IgM, and why?

Answer 22

IgD, because this is a simple RNA splicing of the IgM and IgD constant regions, the same way that displayed Ab and secreted Ab are spliced differently.
It should be noted here that the function of IgD is not well understood, if it has a function at all.

Question 23

What is the all-star enzyme involved in class switching AND somatic hypermutation? Dr. Taffet indicated there will likely be a test question on this!

Answer 23

AID, activation-induced cytidine deaminase

Question 24

Talk to me about somatic hypermutation. What’s happening and where?

Answer 24

The variable region of the Ab (or Ig in the notes), and only the variable region, is undergoing an unusually high rate of mutation. This is particularly evident in the CDR1 and CDR2 regions, which directly contact residues with the CDR3 region of the antigen. It is influenced by the enzyme AID.

Question 25

When does somatic hypermutation happen?

Answer 25

Usually after class-switching

Question 26

For maturing B-cells to continue to proliferate, they must be continually re-stimulated by binding to their cognate antigen, meaning that the whole process of somatic mutation of the variable region happens over and over again. How does this help increase immunity to an invader?

Answer 26

As the B-cell becomes fine tuned, only those B-cells who express increasing affinity for the cognate antigen will continue to proliferate. This whole process is called “affinity maturation.”