Diverse mitogenic agents induce the phosphorylation of two related 42,000-dalton proteins on tyrosine in quiescent chick cells.

Abstract

Incubation of quiescent chicken embryo cells with platelet-derived growth factor, epidermal growth factor, or serum was found to stimulate phosphorylation of two proteins of ca. 42,000 daltons on tyrosine. These proteins are structurally related to each other and to two proteins phosphorylated on tyrosine under similar conditions in mitogen-treated mouse fibroblasts. Three other very different mitogenic agents, the protease trypsin and the chemically unrelated tumor promoters 12-O-tetradecanoyl-phorbol-13-acetate and teleocidin, stimulated phosphorylation of the same proteins. In all cases, phosphotyrosine was detected in these phosphoproteins. Although additional changes in protein phosphorylation were evident, no other proteins were observed by two-dimensional gel electrophoresis which contained increased amounts of phosphotyrosine in mitogen-treated chicken embryo cells. One of these 42,000-dalton proteins was shown previously to be phosphorylated on tyrosine in chicken embryo cells transformed with various retroviruses whose transforming proteins possess tyrosine protein kinase activity. Phosphorylation of the 42,000-dalton proteins could be important in the regulation of cell division.Monolayer cultures of fibroblasts which are sensitive to density-dependent growth inhibition exhibit decreased rates of mitosis, nutrient transport, and macromolecular synthesis as they reach confluence (53,58,62). Most cells in such cultures will recommence DNA synthesis and enter mitosis when exposed to an appropriate growth factor. Mitogens for fibroblasts in culture include the incompletely defined factors in whole unfractionated serum (59) and the following purified growth-stimulatory compounds: platelet-derived growth factor (PDGF; one of the factors contained in serum) (1,36,48), epidermal growth factor (EGF) (2, 31), a number of purified proteases (10, 51), the fibroblast growth factors (30), some prostaglandins (35), insulin (55), and tumor promoters, such as 12-O-tetradecanoyl-phorbol-13-acetate (TPA) (21)(22)(23)54) or the indole alkaloid teleocidin (34). How these diverse agents stimulate renewed growth is poorly understood. There very well may be several rather different biochemical bases of growth stimulation.Increases in protein phosphorylation (57), phospholipid turnover (20), ion and metabolite transport (20,49,53), and protein and RNA synthesis (59) all commence within an hour or so after addition of these growth-stimulatory factors to cells and precede measureable increases in the rate of DNA synthesis. As a result, these processes have each, at one time or another, been attributed the paramount role in the propagation of the mitogenic signal. Because the changes in protein phosphorylation induced by some mitogenic agents are extraordinarily rapid, considerable attention has been focused recently on the possible role of protein kinases in the initiation of the renewed growth.For instance, a serine/threonine-specific protein kinase, C-kinase, copurifies with a protein(s) which binds TPA with high affinity (40...

Protein phosphorylation in a tetradecanoyl phorbol acetate-nonproliferative variant of 3T3 cells.

“…The pleiotypic responses include increased transport of ions and hexoses (18,19,29,39,41), increased activity of rate-limiting enzymes in anabolic pathways (e.g., ornithine decarboxylase) (33,34), and increased phosphorylation of a number of cellular proteins (28,36,39). Of particular relevance to the work reported in this communication are the findings that both EGF and TPA cause increased serine/threonine phosphorylation of the EGF receptor (11,17,20,23), that TPA stimulates phosphorylation of an 80-kDa soluble protein (36), and that all the mitogens (including TPA) cause tyrosyl phosphorylation of a 42-kDa cellular protein (4,16,21,31).…”

Kinases: positive and negative regulators of apoptosis

“…42,43 One substrate tyrosine phosphorylated in a number of different cell types, including hematopoietic cells and both T-and B-lymphocytes, [44][45][46] is a 42-kDa protein. [47][48][49] This protein was initially called MAP kinase. This name was based on the ability of this protein to phosphorylate microtubule associated protein.…”

Inhibition of c-Jun DNA binding by mitogen-activated protein kinase.

“…Members of this family become tyrosine-phosphorylated and activated in cells stimulated with diverse mitogens, including activators of cell surface protein tyrosine kinases and activators of protein kinase C (Cooper et al, 1982(Cooper et al, , 1984Bishop et al, 1983;Gilmore and Martin, 1983;Kohno, 1985;Ray and Sturgill, 1987;Hoshi et al, 1988;Rossomando et al, 1989;Ahn et al, 1990;Ferrell and Martin, 1990;Gotoh et al, 1990b;Haystead et al, 1990; Kyriakis and Avruch, 1990;Nel et al, 1990;Tsao et al, 1990;Anderson et al, 1991;L'Allemain et al, 1991;Tobe et al, 1991;Funasaka et al, 1992 1Abbreviations used: DMEM, Dulbecco's modified Eagle's medium; DTT, dithiothreitol; EGF, epidermal growth factor; ERK, extracellular signal-regulated kinase or early response kinase; HPLC, high-pressure liquid chromatography; MAP, microtubule-associated protein 2 or mitogen-activated protein; MBP, myelin basic protein; MES, 2-(N-morpholino)ethanesulfonic acid; PMSF, phenylmethylsulfonyl fluoride; SDS, sodium dodecyl sulfate; TPA, 12-0-tetradecanoylphorbol-13-acetate; TRE, TPA response element; Tris, tris(hydroxymethyl)aminomethane.…”