Developmental Control of Gene Expression 
Rudolf Grosschedl, Ph.D. — Assistant Investigator 
Dr. Grosschedl is also Assistant Professor of Microbiology and Immunology and of Biochemistry and Bio- 
physics at the University of California, San Francisco. He completed his undergraduate studies on the 
replication oflambdoid bacteriophages in the laboratory of Gerd Hobom in Freiburg, West Germany. His 
graduate studies, on the regulation of histone gene expression, were carried out in the laboratory of Max 
Birnsttel in Zurich, Switzerland. Dr. Grosschedl spent his postdoctoral years with David Baltimore at the 
Massachusetts Institute of Technology and the Whitehead Institute. 
THE process of terminal differentiation turns a 
multipotential cell into a cell that carries out 
a particular function or synthesizes a specific 
product. The lymphoid B lineage ultimately 
generates a cell that secretes antibody. During 
cell differentiation, genes that encode the anti- 
body or associated proteins are expressed in a de- 
fined cell type-specific and temporally ordered 
pattern. 
Transcription of the jx immunoglobulin (Ig) 
gene encoding the heavy chain of the antibody 
can be detected in virtually all lymphocytes. By 
contrast, the k Ig light-chain gene is transcribed 
only in late-stage B cells, and the mb-1 gene en- 
coding an antibody-associated protein is ex- 
pressed only in early-stage B cells. The goal of our 
research is to gain some insight into the molecu- 
lar mechanisms that mediate the developmental 
control of lymphoid-specific gene expression. 
Regulation of Ig Gene Expression in a 
Transgenic Model 
Our general experimental approach is to intro- 
duce intact rearranged Ig genes into cultured lym- 
phoid and nonlymphoid cells and into the germ- 
line of mice. Comparison of transcription of the 
exogenous wild-type and mutant genes in differ- 
ent cell types and at various stages of develop- 
ment allow^s the identification of DNA sequences 
that are instrumental for tissue-specific and tem- 
poral gene regulation. 
Transfer of a rearranged wild-type ^ gene into 
the mouse germline indicated that the tissue- 
specific expression pattern is more complex than 
anticipated from previous observations. Instead 
of a simple "on-state" in lymphoid tissues and 
"off-state" in all nonlymphoid tissues, expres- 
sion of both the endogenous n locus and the ^l 
transgene was also detected in skeletal muscle. 
Moreover, the ^l transgene was reproducibly ex- 
pressed at a low level in several other nonlym- 
phoid tissues. Analysis of the expression pattern 
of mutant )u transgenes containing intragenic de- 
letions or point mutations in nuclear factor-bind- 
ing sites indicated a control by multiple positive 
and negative regulatory mechanisms. 
Consistent with previous transfection data, /it 
transgene expression in lymphoid tissues was de- 
pendent upon the intragenic enhancer and the 
binding site for the Oct-transcription factors in 
the promoter. Neither of these regulatory se- 
quences, however, was required for expression 
in skeletal muscle, which is governed by a mus- 
cle-specific control region located 3' of the 
enhancer. 
Low-level expression in other nonlymphoid 
tissues was independent of the Oct-binding site 
but requires function of the intragenic enhancer. 
Mutation of Es and E5 factor-binding sites in the 
enhancer increased this low-level expression to a 
high level similar to that found in lymphoid tis- 
sues. The off-state of the transgene in liver and 
fibroblasts, however, was not affected by this mu- 
tation, suggesting negative regulation by at least 
two mechanisms. Taken together, a plethora of 
regulatory information appears to be required for 
directing the complex tissue-specific expression 
pattern of the ^l gene in the whole animal. 
To determine the molecular basis for the se- 
quential expression of Ig heavy- and light-chain 
genes during B cell differentiation, we generated 
chimeric Ig ;u gene constructs, in which individ- 
ual regulatory sequences have been replaced by 
the corresponding regulatory sequences from the 
K light-chain gene. Gene transfer of these chi- 
meric transgenes into the mouse germline and 
analysis of their temporal expression pattern in- 
dicated that replacement of the ^ enhancer with 
the K enhancer decreased the level of expression 
specifically in early-stage B cells and delayed, at 
least in part, the appearance of transgene 
transcripts. 
Likewise, replacement of the ^l promoter with 
the K promoter decreased expression specifically 
in early-stage B cells. Neither the k enhancer nor 
the K promoter, however, was sufficient for con- 
ferring upon the jix gene the precise temporal ex- 
pression pattern of the endogenous k gene or an 
intact K transgene, suggesting that the combina- 
tion of the K enhancer and k promoter may be 
required for the proper temporal regulation of Ig 
genes. 
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