Phospholipase C-yl (PLC-yl) is a substrate for several receptor tyrosine kinases and its catalytic activity is increased by tyrosine phosphorylation. However, the biological significance of this molecule in normal or malignant human epithelial cell proliferation is unknown. We determined the relative content of PLC-yl in primary human mammary carcinomas and in nonmalignant mammary tissues. By Western blot and immunohistochemistry, considerably higher levels of PLC-yl protein were detectable in the majority of carcinomas and in one of two benign fibroadenomas compared to normal breast tissues. In 18 of 21 carcinomas that contained high levels of PLC-y1, the presence of phosphotyrosine on PLC-yl could also be detected. All carcinomas in which tyrosine phosphorylated PLC-y1 was present also expressed detectable levels of the epidermal growth factor receptor or erbB-2, two tyrosine kinases known to phosphorylate this enzyme. Thus, a high percentage of mammary carcinomas concomitantly display increased levels of receptor tyrosine kinases and a direct tyrosine phosphorylation substrate, thereby potentially amplifying two successive steps in a signal transduction pathway.Recently, several reports have demonstrated that phospholipase C-yl (PLC-yl) is a direct substrate of the epidermal growth factor (EGF) receptor protein tyrosine kinase (1-4). PLC isoenzymes hydrolyze phosphatidylinositol 4,5-bisphosphate to produce two intracellular second messengers: inositol 1,4,5-trisphosphate and 1,2-diacylglycerol (5), which regulate intracellular levels of Ca2l and protein kinase C activity, respectively. As much as 50-70% of the total cellular pool of PLC-yl becomes tyrosine phosphorylated within 1 min after the addition of EGF to cells (2,3,6). Importantly, growth factor-induced tyrosine phosphorylation increases the catalytic activity of 8). In vitro experiments show that the purified EGF receptor phosphorylates PLC-yl but does not significantly phosphorylate the PLC-p and 8 isoenzymes (4). However, not all tyrosine kinases phosphorylate PLC-yl. Although the plateletderived growth factor (PDGF) (3, 9), fibroblast growth factor (10), nerve growth factor (11), and erbB-2 receptors (12, 13) also phosphorylate PLC-yl, insulin and colony-stimulating factor type 1 receptors do not utilize PLC-'yl as a substrate (3,14,15). Taken together these data suggest that phosphorylation of PLC-yl is a biologically important event in growth factor-stimulated inositol phospholipid metabolism and perhaps plays a role in signal transduction for cell proliferation. However, most of these studies have utilized cultured A-431 human squamous carcinoma cells or mouse 3T3 fibroblasts transfected with wild-type human EGF receptors. Studies in human tissues are lacking and, although PLC-yl is ubiquitously distributed in animal tissues, factors that may influence its level of expression are unknown.To provide evidence for a functional role of PLC-'yl in proliferating human epithelial tissues, we have examined PLC-yl levels in benign and malignant ...

A kinome-wide selectivity screen of >20000 compounds with a rich representation of many structural classes has been completed. Analysis of the selectivity patterns for each class shows that a broad spectrum of structural scaffolds can achieve specificity for many kinase families. Kinase selectivity and potency are inversely correlated, a trend that is also found in a large set of kinase functional data. Although selective and nonselective compounds are mostly similar in their physicochemical characteristics, we identify specific features that are present more frequently in compounds that bind to many kinases. Our results support a scaffold-oriented approach for building compound collections to screen kinase targets.

Epidermal growth factor (EGF) treatment of A-431 epidermoid carcinoma cells elicited a redistribution of phospholipase C-gamma 1 (PLC-gamma 1) from a predominantly cytosolic localization to membrane fractions. The temporal coincidence of this redistribution with EGF stimulation of inositol phosphate formation and EGF increased phosphorylation of PLC-gamma 1 suggests that the membrane association of PLC-gamma 1 is a significant event in second messenger transduction.

No abstract

We have previously shown that multiple topical applications, over 11 days, of the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) induces a persistent inflammatory reaction characterized by edema, cell infiltration and epidermal hyperplasia. In order to characterize the cell infiltrate during the establishment of this inflammatory reaction, immunohistochemistry was performed using two monoclonal antibodies: MOMA-2, a macrophage antibody and Thy-1, a pan T-cell antibody. The level of polymorphonuclear leukocytes (PMNs) peaked by day 3 at 160-fold over nontreated controls and then subsided to a 30-fold elevation on days 7-10. By day 4, the number of macrophages increased 2.9-fold over the nontreated control and by day 10 were elevated 6.0-fold over the nontreated control. In comparison, the number of T-cells present by day 7 was significantly elevated 9.5-fold over the nontreated group and peaked at day 8 with a 19-fold elevation relative to nontreated controls. Topical treatment of animals with hydrocortisone valerate resulted in a dramatic (> 60%) reduction in the number of T-cells present in the tissue. In contrast, there was no effect of the steroid on the number of macrophages present in the tissue. The identification of specific cell types and their time course of infiltration is consistent with the development of a chronic inflammatory lesion.