Mechanism of Action of Polypeptide 
Growth Factors 
Linda J. Pike, Ph.D. — Associate Investigator 
Dr. Pike is also Associate Professor in the Department of Biochemistry and Molecular Biophysics at 
Washington University School of Medicine, St. Louis. She received her B.S. degree in chemistry from the 
University of Delaware and her Ph.D. degree in biochemistry from Duke University, where she studied with 
Robert Lefkowitz. Her postdoctoral training was done in the laboratory of Edwin Krebs at the University 
of Washington, Seattle. 
A number of low-molecular-weight polypep- 
tides have been shown to regulate cell 
growth. These growth factors bind to specific re- 
ceptors on the surface of cells. Through a com- 
plex series of reactions, the binding of the growth 
factor to its receptor stimulates the cell to grow 
and divide. Until recently, little was known of the 
mechanism by which growth factors induce cell 
division. It is now recognized, however, that the 
receptors not only bind the appropriate growth 
factor but also have an enzymatic activity. 
The receptors catalyze the transfer of a phos- 
phate group from adenosine triphosphate to tyro- 
sine residues in selected protein substrates. The 
enzyme possessing this activity is called a tyro- 
sine protein kinase. Typically the phosphoryla- 
tion of a protein by a kinase leads to changes in 
the activity of the protein. Although much is 
known about the growth factor receptor kinases, 
the substrates for these enzymes have not been 
identified. 
My laboratory is involved in studies of the 
mechanism by which the binding of epidermal 
growth factor (EGF) to the outside of the cell 
elicits a biological response inside. This is re- 
ferred to as signal transduction. Because they are 
extremely responsive to EGF, the A431 line of 
human epidermal carcinoma cells is used as the 
model system in most of our studies. 
Phosphatidylinositol Metabolism 
One of the earliest responses of A431 cells to 
EGF is an increase in the metabolism of a particu- 
lar phospholipid, phosphatidylinositol. This im- 
portant lipid serves as a precursor for the genera- 
tion of two intracellular compounds that activate 
various enzymes and thereby mediate the effects 
of EGF within the cell. One of the enzymes in- 
volved in phosphatidylinositol metabolism is 
phosphatidylinositol kinase. Since the activity of 
this enzyme is stimulated by EGF, it represents a 
potential substrate for phosphorylation by the 
EGF receptor tyrosine protein kinase. 
We have purified the phosphatidylinositol ki- 
nase from both A431 cells and human placenta. 
This 55-kDa enzyme is active as a monomer — 
that is, as a single polypeptide chain. It phos- 
phorylates phosphatidylinositol on the 4 position 
of the inositol ring and hence is distinct from an- 
other phosphatidylinositol kinase that phosphor- 
ylates the ring on the 3 position. Information re- 
garding the sequence of the amino acids that 
make up the phosphatidylinositol 4-kinase (PI 
4-kinase) was obtained. Molecular biology tech- 
niques were used to isolate and characterize the 
cDNA encoding the PI 4-kinase. From this cDNA 
the amino acid sequence of the entire PI 4-kinase 
was deduced and was found to be unique. The 
sequence showed limited similarity to other pro- 
teins that bind inositol phosphate or carry out the 
phosphorylation of other sugars, suggesting that 
the PI 4-kinase may be derived from enzymes that 
metabolize sugars. Analysis of the types of mRNAs 
present in a variety of tissues that encode the PI 
4-kinase indicated that at least two types of mRNA 
are present and that the amount of message is reg- 
ulated by physiological stimuli. Efforts are now 
directed toward producing a cell line in which 
this PI 4-kinase cDNA is overexpressed, leading 
to elevated levels of PI 4-kinase activity within 
the cell. The effects of this overexpression on 
the growth properties of the cells will be investi- 
gated. This work is supported by a grant from the 
National Institutes of Health. 
Another enzyme involved in phosphatidylino- 
sitol metabolism is a phosphatidylinositol mono- 
phosphate phosphatase. This enzyme catalyzes 
the reverse of the reaction catalyzed by the PI 
4-kinase — that is, it removes the phosphate from 
the 4 position of the inositol ring of phosphatidyl- 
inositol monophosphate. Although this enzyme 
has not been studied previously, its position in 
the metabolic pathway of phosphatidylinositol 
suggests that it may be important in the overall 
regulation of the pathway. We have characterized 
it with respect to its kinetic properties, substrate 
specificity, and response to various inhibitors. 
We have purified the enzyme to a high degree and 
have shown that it is a 95-kDa glycoprotein. Un- 
like other enzymes in this pathway, it appears to 
have an extracellular domain. The possibility that 
its activity is regulated through the binding of an 
extracellular mediator is being investigated. 
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