Normal and Oncogenic Forms of the src Gene Product 
by cellular hormones. Platelets and the released 
products carry out specific functions in clot for- 
mation and wound healing. Platelets are an ideal 
system to study cellular events involved in the 
transduction of extracellular signals. The src pro- 
tein represents a large percentage of total cell 
protein (0.2-0.4 percent); thus this enzyme may 
regulate important events at the plasma mem- 
brane. We have recently found that four protein 
tyrosine kinases closely related to c-src are also 
expressed in platelets and that thrombin and 
other activators of platelet functions cause a 
rapid increase in the phosphorylation of multiple 
proteins on tyrosine. 
We have found that the phosphorylation of sev- 
eral of these proteins requires platelet aggrega- 
tion and the interaction between the blood cell 
adhesion protein fibrinogen and its receptor on 
the platelet surface. Tyrosine phosphorylation of 
these proteins is not necessary for secretion of the 
intracellular platelet granules (the other major 
function of peripheral blood platelets). These 
studies suggest that tyrosine phosphorylation 
may be involved in triggering intracellular events 
that are induced by fibrinogen-mediated platelet 
aggregation. We are attempting to purify these 
proteins, to identify their function in platelets, 
and to determine how the phosphorylation of 
these proteins on tyrosine residues affects their 
functional activity. 
The expression of high levels of the src protein 
in terminally differentiated cells, such as neurons 
and platelets, indicates that c-src is not exclu- 
sively involved in events that directly regulate 
cellular proliferation. It is difficult to reconcile 
the tumorigenic activity of the RSV src gene prod- 
uct with such a role for the c-src protein in termi- 
nally differentiated cells. Mutational changes in 
the v-src gene probably deregulate and activate 
the tyrosine kinase activity of the v-src gene prod- 
uct, thus allowing inappropriate phosphoryla- 
tion of cellular substrates that are involved in 
growth regulation. The identification of these 
substrates is one of the most difficult and impor- 
tant problems in this field. 
Regulation of the src Protein Activation of 
Tumorigenic Potential 
Evidence suggests that the c-src protein is 
tightly regulated in normal cells and that muta- 
tional alterations in this protein cause a deregula- 
tion and constitutive activation of tyrosine pro- 
tein kinase activity. We are using site-directed 
mutagenesis to define regions of the src molecule 
that affect the kinase activity of this enzyme, to 
identify regions that are important for interaction 
with other cellular proteins that mediate src- 
induced transformation. 
We have focused on the amino-terminal half of 
the src protein that lies outside of the catalytic 
domain of the molecule, in two highly conserved 
regions (SH2 and SH3) that are shared with several 
other cellular proteins. Deletion of either of 
these regions does not prevent transformation by 
oncogenic forms of src but does interfere with 
the interaction of src with several protein sub- 
strates. In addition, deletion of the SH2 or SH3 
domain in wild-type c-src activates the kinase ac- 
tivity and transformation potential of this proto- 
oncogene. We are currently examining the im- 
portance of these regions in events other than 
oncogenic transformation. 
We have also identified a novel protein sub- 
strate that is phosphorylated in v-src-transformed 
cells. This protein, connexin43, is the major com- 
ponent of gap junctions, which mediate cell-to- 
cell communication. The phosphorylation of 
connexin43 correlated with the ability of v-src to 
cause a reduction in junctional communication 
between cells, suggesting that phosphorylation 
of connexin43 by the v-src protein may be re- 
sponsible for the inhibition of junctional commu- 
nication in v-src-transformed cells. 
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