Genetic Mechanisms Involved in the Generation of the Antibody Repertoire 
elements that govern the assembly process. Most 
notably, we have shown that genetic regulatory 
elements previously thought to control only the 
level of gene expression can also serve to control 
specific gene rearrangement. 
Currently we are attempting to generate sev- 
eral novel mouse models to study factors in- 
volved in lymphocyte differentiation. One ap- 
proach is to produce a mutant mouse line that 
cannot produce endogenous antibody molecules. 
To accomplish this, we employed new methods 
that permit introduction of desired genetic muta- 
tions into specific genes in mouse embryonal 
stem cell lines to create cells that lack germline 
gene segments necessary for forming functional 
antibody genes. We used these mutant cell lines 
to create chimeric mice that contain cells lacking 
the antibody gene cassettes. Such chimeric mice 
should generate mutant mice that inherit in their 
germline an inability to make endogenous 
antibodies. 
We will use the mutant animals to study the 
role of antibody gene products in regulating anti- 
body gene assembly and lymphocyte develop- 
ment. A potential practical use of such mice may 
be achieved by breeding them with a transgenic 
mouse line that we have engineered to contain 
functional human antibody gene cassettes. The 
hope is that the hybrid animal will now rely on 
the human antibody genes for its immune system, 
providing a more effective method of generating 
tailor-made human antibodies. 
We also are interested in the mechanism by 
which certain intracellular messenger molecules 
(lymphokines or interleukins) affect the develop- 
ment and action of B lymphocytes. Certain T lym- 
phocytes regulate aspects of the immune re- 
sponse; some of these produce lymphokines that 
stimulate activated B lymphocytes (during sec- 
ondary differentiation) to produce antigen- 
specific antibodies that differ in where they go in 
the body and how they mediate their protective 
activity (referred to as different effector activi- 
ties). The difference in these antibodies results 
from the use of different gene cassettes to en- 
code, not the antigen-binding portion, but the ef- 
fector portion of the antibody molecule. This phe- 
nomenon results from a different type of gene 
recombination assembly event, in which one part 
of an already active antibody gene is replaced 
with a different one. We have shown that particu- 
lar lymphokines actually direct this gene rear- 
rangement event by giving a B lymphocyte a sig- 
nal to prepare a specific effector gene cassette for 
a rearrangement event. Most recently we have 
shown that the ultimate targets of the lympho- 
kines are novel elements that control the tran- 
scription of the effector gene cassette. 
Finally, we are interested in the molecular sig- 
nals that direct precursor lymphocytes through 
the primary differentiation process. One focus of 
this work is on three related but distinct genes, 
the N-, L-, and c-myc genes. These genes encode 
proteins that operate in the nucleus of the cell, 
presumably to regulate the expression of other 
genes. The members of this gene family contrib- 
ute to the generation of various types of tumors 
when their expression is altered as the result of 
mutations. We have found that individual 
members of this family are expressed at specific 
points during the differentiation of lymphocytes. 
For example, the N-myc gene is expressed only in 
lymphocyte precursors where expression levels 
are greatly increased following interaction of 
these cells with a factor that specifically stimu- 
lates their growth. 
To define further the function of this gene fam- 
ily, we have introduced the N-myc and L-myc 
genes into transgenic mice in forms in which they 
are incorrectly expressed throughout lympho- 
cyte development (normally they are expressed 
only very early in lymphocyte development). 
Such incorrect expression of these two genes 
leads, respectively, to defects in B or T lympho- 
cyte development, including frequent generation 
of lymphocyte tumors. The genetic mechanisms 
by which incorrect expression of the deregulated 
myc genes affects normal lymphocyte develop- 
ment are being analyzed. 
As a complementary approach, we used the 
gene-targeting methodologies outlined above to 
create mutant mice that lack functional N-myc 
genes. Loss of N-myc gene function appears to be 
lethal at an early stage of murine development. 
Therefore, to study normal function, we are at- 
tempting to rescue development in the N-myc 
mutant mice lacking functional endogenous 
genes by breeding these animals with mice that 
carry functional N-myc transgenes that are ex- 
pressed in various cell lineages. 
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