Regulation of Gene Expression in Steroid 
Hormone Biosynthesis 
Keith L. Parker, M.D., Ph.D. — Assistant Investigator 
Dr. Parker is also Associate Professor of Medicine and Biochemistry at Duke University Medical Center. 
After attending Williams College, he earned his M.D. and Ph.D. degrees in genetics at Washington Univer- 
sity, studying with Donald Shreffler. He served as intern and resident in internal medicine at Parkland 
Memorial Hospital. He then moved to the Department of Genetics at Harvard Medical School, where he 
was a postdoctoral fellow with Jonathan Seidman. Dr. Parker's next move was to the faculty of Duke 
University Medical Center. 
THE adrenal gland plays essential roles in the 
body's ability to respond to stress. Two dif- 
ferent parts of the gland, an inner medulla and an 
outer cortex, produce discrete agents of this re- 
sponse. The medulla produces epinephrine and 
norepinephrine, which are released very rapidly, 
preparing the organism for immediate physical 
activity. In contrast, the cortex produces steroid 
hormones, which are released more slowly and 
exert prolonged effects. 
These adrenal steroids constitute two major 
classes: glucocorticoids, which are synthesized 
by the inner zone of the cortex and control carbo- 
hydrate metabolism, and mineralocorticoids, 
which are made in the outer zone and regulate 
salt and water balance. Both classes of steroid 
hormones are formed from cholesterol by the se- 
quential action of a related group of steroido- 
genic enzymes. Certain of these enzymes, such as 
the cholesterol side-chain cleavage enzyme 
(SCC), are expressed in all steroidogenic tissues. 
In contrast, steroid 1 1 jS-hydroxylase (11/3- 
OHase) and 2 1 -hydroxylase (21-OHase) are only 
expressed in the adrenal cortex and largely deter- 
mine the unique ability of this tissue to make glu- 
cocorticoids and mineralocorticoids. The physio- 
logical regulators of these two classes of adrenal 
steroids differ markedly despite the shared role of 
the enzymes in their biosynthesis. 
We are interested in defining the events that 
control the expression of the adrenal steroido- 
genic enzymes. These studies have addressed two 
major questions. First, what mechanisms direct 
the expression of these related genes within adre- 
nocortical cells? Second, what determines the 
functional differentiation of the adrenal cortex 
into mineralocorticoid- and glucocorticoid- 
producing zones? 
Our studies of gene regulation have focused on 
the 5'-flanking regions of these genes. This part, 
termed the promoter region, contains most se- 
quences important in transcriptional regulation 
of other genes. We first showed that the 5'-flank- 
ing regions of the steroidogenic enzymes retained 
all information required for adrenal-selective and 
hormonally inducible gene expression. 
Next, in a combination of structural and func- 
tional studies, we identified a protein, termed 
steroidogenic regulatory protein, that played a 
major role in regulating the expression of the 
steroidogenic enzymes. This protein was only 
present in nuclear extracts from steroidogenic 
cell lines, suggesting that it contributed to the 
cell-selective expression of these genes. More- 
over, steroidogenic regulatory protein appeared 
to interact with the promoter regions of all three 
steroidogenic enzymes, suggesting that it coordi- 
nates the expression of this entire network of en- 
zymes in adrenal cells. 
Now that this major regulatory protein has 
been identified, our next goal is to understand 
how it affects the expression of 21-OHase, 1 1(8- 
OHase, and SCC. These studies require sufficient 
amounts of protein for biochemical analysis, and 
we have therefore initiated studies using cow ad- 
renal glands as a source of protein. These experi- 
ments demonstrated that bovine protein behaved 
very similarly to steroidogenic regulatory protein 
isolated from mouse adrenocortical tumor cells. 
By sequential purification steps, we have mark- 
edly enriched the bovine protein to a degree that 
should permit determination of its amino acid 
sequence. 
Attempts are also under way to raise antibodies 
against the partially purified bovine protein. The 
combination of specific antibodies and amino 
acid sequence data should allow us to clone the 
gene encoding this key regulator. By comparing 
the primary structure of steroidogenic regulatory 
protein with that of previously described tran- 
scriptional regulatory proteins, such as those of 
the steroid hormone receptor, we may gain new 
insights into the mechanisms that regulate the ad- 
renal steroidogenic enzymes. The availability of 
specific probes and antibodies to steroidogenic 
regulatory protein will further permit us to study 
the mechanisms that regulate its expression. 
These studies will provide new insights into the 
basis for tissue-specific differences in the produc- 
tion of steroid hormones. They may also increase 
our understanding of the general mechanisms of 
tissue-specific gene expression. 
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