Regulation of Gene Activity During 
B Cell Development 
Harinder Singh, Ph.D. — Assistant Investigator 
Dr. Singh is also Assistant Professor in the Department of Molecular Genetics and Cell Biology at the Uni- 
versity of Chicago. He received his Ph.D. degree in biochemistry, molecular biology, and cell biology with 
Lawrence Dumas at Northwestern University. His postdoctoral research was done with Phillip Sharp at 
the Massachusetts Institute of Technology, as a Jane Coffin Childs fellow. He remained at MIT as a research 
associate until his move to Chicago. 
MY research interests are focused on the 
analysis of transcriptional regulatory cir- 
cuits that turn genes on or off during the growth 
and differentiation of B lymphocytes, cells that 
produce antibodies. The B cell lineage is a very 
useful model for exploring the molecular basis of 
differential gene activity in mammalian develop- 
ment. We are seeking answers to the following 
questions: What is the nature of the genetic ele- 
ment(s) linked to a target gene that controls its 
transcriptional activity? What is the nature of the 
regulatory protein (s) that recognizes this genetic 
element? How does the regulatory protein on 
binding near its target gene modulate the activity 
of the enzyme complex that transcribes the gene? 
What is the mechanism by which a growth or de- 
velopmental signal is transduced by the regula- 
tory protein (s) to effect gene activity? How does 
the structure of the chromatin within which the 
target gene is packaged influence the function of 
the regulatory protein (s)? 
The heavy- and light-chain genes encoding the 
immunoglobulin (Ig) molecule are selectively 
transcribed in B cells. These genes are assembled 
from gene segments through an ordered series of 
somatic recombination events that occur in a de- 
veloping B cell. The heavy-chain gene locus is the 
first to undergo recombination and transcrip- 
tional activation, thereby defining the pre-B cell 
developmental state. Subsequently one of two 
light-chain gene loci is recombined and ex- 
pressed, resulting in the development of a mature 
B cell. Ig genes contain multiple cis-acting tran- 
scriptional regulatory elements that restrict their 
expression to appropriate stages in the B lineage. 
Previous work by various research groups has 
resulted in the identification, characterization, 
and cloning of a B cell-specific regulatory pro- 
tein, Oct-2. This protein recognizes the octanu- 
cleotide sequence ATTTGCAT, which confers B 
cell specificity to Ig gene promoters. The same 
sequence motif is also a functional component of 
the heavy-chain and /c-light-chain gene en- 
hancers. Oct-2 can activate transcription of a re- 
porter gene linked to an Ig promoter in a non-B 
cell. Thus Oct-2 appears to be both necessary and 
sufficient for regulating the activity of Ig pro- 
moters. The deduced amino acid sequence of 
Oct-2 reveals a region of similarity that is shared 
with three other regulatory proteins — Pit-1, 
Oct-1 , and unc-86. This region is termed the POU 
box and includes a subdomain related to the 
homeobox. 
Current research focuses on the regulatory 
functions of Oct-2 in B cell development, as well 
as the mechanism underlying lineage-restricted 
expression. Oct-2 is likely to represent a member 
of a regulatory gene hierarchy that controls the 
development of B cells from pluripotent hemato- 
poietic stem cells. By studying the regulation of 
Oct-2 expression, we can identify and isolate 
genes interacting with Oct-2 in the hierarchy. 
The murine gene encoding Oct-2 has been iso- 
lated in the laboratory and is being analyzed for 
regulatory sequences that control its expression 
in developing B cells. 
The activity of the Oct-2 gene is regulated dur- 
ing B cell differentiation. The Oct-2 gene is ex- 
pressed at low levels in pre-B cells. Oct-2 ex- 
pression is transcriptionally up-regulated upon 
pre-B cell differentiation, and higher Oct-2 pro- 
tein levels correlate with activation of the Ig k- 
light-chain gene locus. Increased expression of 
Oct-2 in pre-B cells is induced by signaling with 
the B cell mitogen, bacterial lipopolysaccharide, 
as well as the lymphokine interleukin-1 (IL-1). 
Transforming growth factor-jS, an inhibitor of k 
gene induction in pre-B cells, blocks the up- 
regulation of Oct-2 but not the activation of NF- 
kB, another regulatory protein that has been im- 
plicated in the control of the activity of the k 
locus. We propose a model in which the con- 
certed action of increased levels of Oct-2 and ac- 
tivated NF-kB controls the proper stage-specific 
expression of the k locus. Our results suggest that 
NF-kB may also be involved in regulating Oct-2 
transcription during pre-B cell differentiation. 
Therefore this transition in B cell ontogeny may 
be controlled by a cascade of transcriptional 
regulators. 
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