B Cells 4

Question 1

describe the amount of protein-coding genes relative to the amount of Ig and TCRs

what does this mean?

Answer 1

a lottttttt more Ig/TCRs compared to the number of protein-coding genes

means there is something allowing genes to change so they can produce more variable proteins

Question 2

are heavy and light chain genes on antibodies encoded on the same or separate chromosomes?

Answer 2

separate chromosomes

Question 3

what is somatic recombination?

Answer 3

rearrangement of V, J, C (and D) gene segments in Ig

Question 4

describe the process of somatic recombination from when we inherit genes (2 steps)

Answer 4

1. our germline DNA from our parents contains many segments of each V, J, C (and D) segment
2. during B cell development, somatic recombination selects one of each

Question 5

what are the segments of the Ig light chain?

Answer 5

Variable
Joining
Constant

Question 6

what are the segments of the Ig heavy chain?

Answer 6

Variable
Diversity
Joining
Constant

Question 7

how many CDR loops are there per light or heavy chain? how many chains are there in Ig? therefore, how many CDR loops are there in total?

Answer 7

3 CDR

4 variable chains in Ig

12 CDR in total

Question 8

describe where each of the 3 CDR loops are encoded and which is most variable?

Answer 8

CDR 1 and 2 are encoded in the V segments of light and heavy chains

CDR 3 is encoded in:
- joining of V-J segments of light chain
- joining of V,D,J segments of heavy chain

Question 9

describe the loci of the LIGHT CHAIN

Answer 9

two different loci, each with different constant regions:
1. KAPPA chain
2. GAMMA chain

Question 10

what does each locus/chain of the light chain include?

Answer 10

has many V and J regions

Kappa only has 1 C

Question 11

which locus/chain of the light chain will be expressed?

Answer 11

either the full kappa OR gamma chain will be expressed and silence the other one

Question 12

describe the loci of the HEAVY CHAIN

Answer 12

1 locus

Question 13

what does the locus of the heavy chain include?

Answer 13

many different V, D, J, C regions

Question 14

What do the C regions on the heavy chain correspond to? and why does this make sense?

Answer 14

corresponds to diff isotypes –> IgM, IgD, IgG, etc

makes sense bc each isotype is differentiated by its number of constant regions

Question 15

what are the steps for B cell receptor maturation?

Answer 15

1. precursor B cell has inherited genes
2. somatic recombination selects segments
3. recombinase proteins join gene segments together
4. transcription, splicing
5. mature naive B cell

Question 16

is BCR maturation reversible or irreversible? why?

Answer 16

IRREVERSIBLE

bc segments that are not chosen will get deleted

Question 17

what is combinatorial diversity?

how is it different than somatic recombination?

Answer 17

THE DIVERSITY that is a result of diff combos of V, D, J regions

somatic recombination is the PROCESS, combinatorial diversity is the DIVERSITY

Question 18

what does RSS stand for?

Answer 18

Recombination Signal Sequence

Question 19

what is the role of RSS?

Answer 19

flanks each antibody gene segment to direct recombinase enzymes to pair different segments together for recombination

Question 20

what is the structure of RSS? (3)

Answer 20

1. conserved nonamer
2. conserved heptamer
3. 12 or 23 bp spacer btwn the nonamer and heptamer

Question 21

what is the 12/23 rule?

Answer 21

a 12-bp RSS must pair with a 23-bp RSS for recombination to occur

ex. 12-bp RSS binds J and combines with 23-bp RSS

Question 22

what happens when the 12 and 23 RSS bring segments together?

Answer 22

every segment between them that were not selected is put in a loop that is cut out (irreversibly)

Question 23

what gene activates recombination?

Answer 23

RAG –> RAG1, RAG2

Question 24

what does RAG do?

Answer 24

recognizes and cuts DNA at Ig-encoding region where RSS have brought segments together

Question 25

what is the signal joint? what happens to it?

Answer 25

the loop that is excised / deleted from the chromosome

Question 26

what is the coding joint?

Answer 26

coding regions of selected V and J regions that remain

Question 27

what is junctional diversity?

Answer 27

during recombination, nucleotides may be added or removed at the junctions btwn:
- Heavy: V and D, D and J
- Light: V and J

Question 28

when does junctional diversity occur?

Answer 28

once RAG picks each segment, nucleotides are added and removed during ligation when the segments join

Question 29

what happens to the ends of the coding joint when the signal joint is cut off?

Answer 29

junctional diversity! repair proteins will bind the DNA hairpin and ARTEMIS will open the DNA hairpin

Question 30

what is ARTEMIS

Answer 30

an endonuclease that binds and opens the DNA hairpin

Question 31

where does hairpin cleavage occur?

Answer 31

at random place

Question 32

what are the different ways the hairpin can be opened? (3)

Answer 32

1. cut with a blunt end
2. 5’ overhang
3. 3’ overhang

Question 33

what is the role of exonucleases in hairpin cleavage?

Answer 33

exonucleases may REMOVE nucleotides on each side of the coding joint

Question 34

what can be added to the coding joint? where?

Answer 34

ADD palindromic nucleotide sequences added to overhangs

Question 35

what is a palindromic nucleotide sequence?

Answer 35

complementary DNA added where 5’ - 3’ sequence is same as 3’ - 5’

Question 36

what does Terminal Deoxynucleotidyl Transferase do?

what chain does it act on?

how does this affect CDRs?

Answer 36

add up to 20 NON-TEMPLATE encoded to the cleaved chain

acts on HEAVY chain primarily

it is the reason why CDRs vary in length

Question 37

what happens after TdT adds nucleotides?

Answer 37

repair enzymes trim off any nonmatching/unpaired nucleotides, fill in remaining single-stranded gaps, and ligate the new DNA