The Organization and
Expression of Ig
Genes
Justin Walter
Immunology, Fall 2008
Introduction
The vertebrate immune system is capable
of responding to an essentially infinite
array of foreign antigens
Variable vs. constant regions
Basis for variability organization and
expression of Ig genes
Overview
Historical perspective – early theories
“Multigene” organization of Ig genes
Variable region gene rearrangements
Basics of mechanism – key players: DNA
signal sequences, specific enzymes
Generation of Ab diversity
seven primary routes
Historical perspective
Ig sequence analysis revealed many
dilemmas
Extreme diversity of Ab specificity
Variable regions vs constant regions
Isotypes with similar Ag specificity but
differing heavy-chain constant regions
Historical perspective
Proponents of the one-gene-one-protein
paradigm had trouble reconciling this model with
the oddities of Igs.
This led to an initial “germ-line theory” which
suggested that a significant portion of the
genome is dedicated solely to Ab coding.
Argument: the immune system is THAT important
Historical perspective
In contrast, “somatic-variation theories”
emerged which suggested the opposite:
Relatively small amount of Ig genes
Specificity arises from mutation and/or
recombination
Historical perspective
Dreyer & Bennet (1965)
Two-gene, one-protein model
No precedent in any biological system
Historical perspective
Tonegawa & Hozumi (1976)
Compare Ig DNA from embryonic (germline)
and adult myeloma (somatic) cells
Experimental data suggested that during
differentiation, the V and C genes undergo
rearrangement.
1987 Nobel prize
Tonegawa & Hozumi
Multigene Organization of Ig genes
Each “class” of Ig components (kappa,
lambda, heavy) encoded by separate
multigene families on different
chromosomes
Each family contains several coding
sequences, or gene segments
Multigene Organization of Ig genes
Multigene Organization of Ig genes
Κ & λ light chains: V (variable), J (joining), and C
(constant) gene segments
Heavy chains: V, D (diversity), J, and C gene
segments
A leader (L) sequence also precedes each V
segment.
Gene segments discovered by comparing DNA
sequences with amino acid sequences of Igs
Tonegawa, again.
Organization of Ig germ-line
gene segments (mouse)
Pre-rearrangement!
Variable-region rearrangement
Multifaceted process, produces mature B
cells which are “committed” to express
specific Ab
Specificity of Ab determined by the sequence
of its rearranged variable genes.
Light-chain rearrangements
V-J rearrangements
Specific “allowed” rearrangements differ from
species to species, but a “big-picture” view
can suffice
Rearrangement occurs in ordered steps but
can be considered as random events which
result in the random determination of Ab
specificity
Kappa light-chain
rearrangement & RNA
processing
Leader sequence targets
nascent protein to ER
and is subsequently
cleaved
Heavy-chain rearrangements
Requires two separate rearrangement events
D-Jjoining
V-DJ joining
Differential polyadenylation & RNA splicing can
result in mRNA with either Cu or Cδ
heavy chain genes
B cells can express BOTH IgM and IgD with identical
Ag specificity on its surface
Heavy-chain
rearrangement
The focus here is on
IgM IgD B cells…
Mechanism of Variable region
DNA rearrangements
Recombination signal sequences direct recombination
Mechanism of Variable region
DNA rearrangements
Recombination signal sequences direct recombination
Vλ, JK, VH, JH VK, Jλ, DH
Mechanism of Variable region
DNA rearrangements
A one-turn RSS can only join with a two-
turn RSS
Why might this be?
Mechanism of Variable region
DNA rearrangements
Gene segments are joined by a class of
enzymes called recombinases
Two recombination-activating genes
encode proteins which act together to
mediate V-(D)-J joining
RAG-1
RAG-2
a. Same b. opposite
transcriptional transcriptional
orientation orientation
(most common)
Addn. of P-
nucleotides
accomplished
with repair
enzymes
?
Mechanism of Variable region
DNA rearrangements
Rearrangements may be productive or
nonproductive.
Mechanism of Variable region
DNA rearrangements
Allelic exlusion
Heavy-chain genes only expressed from one
chromosome
Light-chain genes only expressed from one
chromosome
Essential for specificity
Expression of both alleles would result in a
“multispecific” B cell
Mechanism of Variable region
DNA rearrangements
Allelic exlusion
Generation of Ab diversity
Multiple germ-line gene segments
Combinatorial V-(D)-J joining
Junctional flexibility
P-region nucleotide addition
N-region nucleotide addition
Somatic hypermutation
Combinatorial association of light and
heavy chains
Possibly as high as 1010!
Junctional flexibility
P/N-addition
Somatic hypermutation
Nucleotide replacement, mediated by
activation-induced cytidine deaminase
(AID)
Also plays a key role in class switching
Frequency of 10-3 per bp per generation
100,000X the rate of spontaneous mutation!
Approx 1 mutation every 2 cell divisions
Somatic hypermutation