SOMATIC MUTATION & AFFINITY MATURATION OF A CHIMERIC IG
Project Number1R15AI037765-01
Contact PI/Project LeaderPOLLOCK, ROBERTA R
Awardee OrganizationOCCIDENTAL COLLEGE
Description
Abstract Text
The generation of antibody diversity is a complex and highly regulated
process. Immunoglobulin (Ig) gene rearrangement, to form functional
variable (V) region genes (which bind antigen) is the major way this
diversity is initially generated, and occurs before antigen is
encountered. Additional diversity arises during an immune response by
somatic mutation and antigen selection of mutated antibodies with higher
antigen affinity.
The goal of this proposal is to study, in transgenic mice, how expression
of chimeric immunoglobulins composed of rearranged murine variable region
genes and human constant (C) region genes, affects the rearrangement of
endogenous variable region genes and whether such chimeric genes undergo
somatic mutation and affinity maturation. Initial studies will be done on
mice expressing only a chimeric Ig heavy chain; later studies will be done
on mice expressing chimeric heavy and light chains.
Two types of chimeric transgenes will be used. Both the heavy and the
light chain murine variable regions are derived from a hybridoma making an
anti-dansyl response (dansyl is a hapten, which is a small, simple
molecule that contains only one antigenic determinant). The heavy chain
constructs consist of a murine heavy chain anti-dansyl V region and a
human gamma-1 C region, with the murine V/H promoter, intron enhancer, and
plus or minus the 3' alpha enhancer. The light chain construct contains
a murine light chain anti-dansyl V region and a human kappa light chain C
region, with the V/L promoter, intron enhancer, and 3' enhancer. Since
the murine V regions are specific for a known antigen, we can study both
somatic mutation and affinity maturation in these mice. In particular, we
can ask whether a human gamma-1 constant region can send three different
types of signals to murine B lymphocytes; one to shut down endogenous V
gene rearrangement, a second to initiate somatic mutation and a third to
mediate antigen selection.
The specific questions to be asked are: 1) does expression of a human
gamma-1 constant region inhibit rearrangement of endogenous heavy chain V
genes; 2) do the transgenes undergo somatic mutation in these animals; in
other words, can the human gamma-1 gene signal the B lymphocyte that it
has encountered antigen, and in the presence of T lymphocyte help turn on
somatic mutation, and are the transgenes then mutated; and 3) do the
transgenes undergo affinity maturation; are cells expressing mutated
transgenes with increased affinity for dansyl selectively stimulated to
proliferate, resulting in increased antibody affinity over time after
immunization.
Public Health Relevance Statement
Data not available.
NIH Spending Category
No NIH Spending Category available.
Project Terms
B lymphocyteT lymphocyteantibody specificitychimeric proteinsgene mutationgene rearrangementgenetically modified animalshuman genetic material taghybridomasimmunoglobulin genesimmunoglobulinsintronslaboratory mouseleukocyte activation /transformation
National Institute of Allergy and Infectious Diseases
CFDA Code
DUNS Number
046436051
UEI
DCQQX5TRCYN9
Project Start Date
01-June-1995
Project End Date
31-May-1999
Budget Start Date
01-June-1995
Budget End Date
31-May-1999
Project Funding Information for 1995
Total Funding
$103,343
Direct Costs
$75,000
Indirect Costs
$28,343
Year
Funding IC
FY Total Cost by IC
1995
National Institute of Allergy and Infectious Diseases
$103,343
Year
Funding IC
FY Total Cost by IC
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