2. Immunoglobulin Genetics
History
• Same C region could associate with many V regions
– IgG Ab with different specificities
• Same V region could associate with many C regions
– Same idiotype on IgG and IgM Ab
• V and C regions coded for by separate genes
3. Light Chain Gene Families
Germ line gene organization
Jλ1 Cλ1
E
Jλ2 Cλ2
E
Jλ3 Cλ3
E
Jλ4 Cλ4
E
P
L Vλ1
P
L Vλn
P
L Vλ2
Lambda light chain genes; n=30
P
L V κ 1
P
L V κ n
P
L V κ 2 Jκ2 Jκ3 Jκ5 Cκ
E
Jκ1 Jκ4
Kappa light chain genes; n=300
4. Light Chain Gene Families
Gene rearrangement and expression
L Vκ 1 L Vκ n
L Vκ 2 Jκ2 Jκ3 Jκ5 Cκ
Jκ1 J4
DNA
P P
P E
P P E
L Vκ 1 L Jκ5 Cκ
Vκ 2 Jκ4
DNA
DNA Rearrangement
V C
J
V C
Protein
Transport to ER
Protein
Translation
C
L V J
E
L V C
J
RNA
Transcription
Primary transcript
C
L V J
RNA Processing
RNA
mRNA
5. Heavy Chain Gene Family
Germ line gene organization
Introns separate exons coding for H chain domains
Heavy chain genes; Vn=1000, Dn=15
P
L V1
P
L Vn
P
L V2 D2 D3
D1 Dn
Cµ Cδ Cγ3 Cε
C γ2
C γ1 C γ4 Cα1 Cα2
J2 J3 J5
J1 J4
E
CH1 H CH2 CH3 CH4
6. Heavy Chain Gene Family
Gene rearrangement and expression
P
L V1
P
L Vn
P
L V2 D2 D3
D1 Dn Cµ Cδ
J2 J3 J5
J1 J4
E
DJ rearrangement
P
L Vn
P
L V1
P
L V2
D2
D1 Cµ Cδ
J5
J4
E
DNA
VDJ rearrangement
D2 Cµ Cδ
J5
J4
E
P
L V1
P
L
V2
DNA
Transcription
D2 Cµ Cδ
J5
J4
E
L
V2
RNA
Primary transcript
7. Heavy Chain Gene Family
Gene rearrangement and expression
E
Primary transcript
E
J4
D2
V2 Cµ Cδ
J5
L Cµ Cδ
J5
L D2 J4
V2
RNA Processing
An
mRNA for Cµ
Cµ
L DJ
V Cδ
L DJ
V
An
mRNA for Cδ
Translation
Cµ heavy chain
Cµ
L DJ
V Cδ
L DJ
V
Cδ heavy chain
Transport to ER
Cµ heavy chain
V C
Cµ
DJ
V Cδ
DJ
V
Cδ heavy chain
V C
8. Mechanism of DNA Rearrangements
• Recombination
signal sequences
(RSS)
– Nonmer
– Heptamer
– 1 or 2 turn signals
• Rag-1 and Rag-2
L V
2
J C
1
Lambda light chains
L V
1
J C
2
Kappa light chains
Heavy chains
L V
2
J C
2
1
1
D
Heptamer
CACAGTG
GTGTCAC
Nonamer
ACAAAAACC
TGTTTTTGG
23 bp
23 bp
Two turn
RSS
2
Nonamer
GGTTTTTGT
CCAAAAACA
Heptamer
CACTGTG
GTGACAC
12 bp
12 bp
One turn
RSS
1
9. Order of Ig Gene Expression
+
-
Heavy chain gene
VDJ recombination
Productive rearrangement ?
Yes
No
Transcribe/translate
1st
2nd
or
Apoptosis
Progenitor B cell
Pre B cell
µ µ
µ (δ) Heavy chain
Paternal
Maternal
10. Order of Ig Gene Expression
+
-
Light chain gene
VJ recombination
Productive rearrangement ?
Yes
No
Transcribe/translate
Light chain
4 th
Apoptosis
1st - κ
2nd -κ
3rd λ
4th - λ
or
Pre B cell
Mature B cell
µ µ
µ
µ+ δ
11. Order of Ig Gene Expression
Explains:
• Why an individual B cell only produces one
kind of Ig with one kind of light chain
• Why an individual B cell makes only makes
antibodies of one specificity
• Why there is allelic exclusion
Importance of allelic exclusion??
12. Origin of Antibody Diversity
History
• Definition – Diversity (repitoire) is the total
of all the Ab specificities that an organism
is capable of expressing
• Theories
– Germ line theory
– Somatic mutation theory
13. Origin of Antibody Diversity
Current concepts
• Multiple V genes
• V-J and V-D-J joining
• Junctional diversity
• N region insertions
– Amino acid
sequences not
encoded in the germ
line
D region
T G
G
C C
C
Pro Trp
C C C C C C
V gene
T G
G
D region
C C C C C C
V gene
T G
G
C G
G
C C
C
Pro Arg
C C C C C C
V gene
T G
G
D region
C G
C
C C
C
Pro Pro
14. TdT adds nucleotides
J
D
-GTCAATG J
D
-GTCAATG- +
Ligate
N region insertion Intron
J
D
Generation of N region insertions
D J
Break DNA
J
D- OH
15. Origin of Antibody Diversity
Current concepts
• Multiple V genes
• V-J and V-D-J joining
• Junctional diversity
• N region insertions
• Somatic mutation
• Combinatorial association
• Multispecificity
16. Generation of Diversity
B cell receptor
(Immunoglobulin)
Heavy Light
V gene segments 1000 300
D gene segments 15 -
J gene segments 4 4
N region insertion ++ -
Junctional diversity +++ +
Somatic mutation + +
V x D x J
1000 x 15 x 4
V x J
300 x 4
Total 6 x 104
1.2 x 103
Combinatorial association 7.2 x 107
• Ag independent process • Clonal selection
