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1.
Fibroblastic cultures derived from normal human tissues undergo a finite number of population doublings when serially subcultivated in vitro (see refs 1, 2 for reviews). Epidermal growth factor (EGF) serves as a mitogen for early doubling level cultures of the human fetal lung-derived cell strain, WI-38, under serum-free conditions. The ability of cells from late doubling level cultures to respond mitogenically to EGF is lost, however, despite undiminished binding of EGF throughout the replicative lifespan. The ultimate effects of EGF, that is DNA synthesis and mitosis (see ref. 4 for review), occur after a sequence of events initiated by binding of ligand to specific cellular receptors. The receptor for EGF has been characterized as a 145,000-165,000 (145 K-165 K) molecular weight doublet, and, like the receptors for platelet-derived growth factor and insulin, and the transforming proteins of certain of the RNA tumour viruses, is a tyrosine-specific protein kinase with autophosphorylating activity. Moreover, several of the cellular target molecules of tyrosine phosphorylation have been found to be substrates for two or more of these kinases. The hypothesis that tyrosine phosphorylation underlies a common mechanism of growth control prompted us to ask whether the loss of responsiveness to EGF by late doubling level WI-38 cells is accompanied by altered expression of the EGF receptor, and specifically whether changes occur in the ability of receptors from populations of cells of various in vitro ages to catalyse tyrosine autophosphorylation. We show here that autophosphorylating activity is absent from the EGF receptor of cells which have lost their mitogenic responsiveness to EGF.  相似文献   

2.
Syk is a protein tyrosine kinase that is widely expressed in haematopoietic cells. It is involved in coupling activated immunoreceptors to downstream signalling events that mediate diverse cellular responses including proliferation, differentiation and phagocytosis. Syk expression has been reported in cell lines of epithelial origin, but its function in these cells remains unknown. Here we show that Syk is commonly expressed in normal human breast tissue, benign breast lesions and low-tumorigenic breast cancer cell lines. Syk messenger RNA and protein, however, are low or undetectable in invasive breast carcinoma tissue and cell lines. Transfection of wild-type Syk into a Syk-negative breast cancer cell line markedly inhibited its tumour growth and metastasis formation in athymic mice. Conversely, overexpression of a kinase-deficient Syk in a Syk-positive breast cancer cell line significantly increased its tumour incidence and growth. Suppression of tumour growth by the reintroduction of Syk appeared to be the result of aberrant mitosis and cytokinesis. We propose that Syk is a potent modulator of epithelial cell growth and a potential tumour suppressor in human breast carcinomas.  相似文献   

3.
High proportion of Ig-producing cells making autoantibody in normal mice   总被引:14,自引:0,他引:14  
E J Steele  A J Cunningham 《Nature》1978,274(5670):483-484
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4.
H Riedel  T J Dull  J Schlessinger  A Ullrich 《Nature》1986,324(6092):68-70
The cell surface receptors for insulin and epidermal growth factor (EGF) appear to share a common evolutionary origin, as suggested by structural similarity of cysteine-rich regions in their extracellular domains and a highly conserved tyrosine-specific protein kinase domain. Only minor similarity is found outside this catalytic domain, as expected for receptors that have different ligand specificities and generate different biological signals. The EGF receptor is a single polypeptide chain but the insulin receptor consists of distinct alpha and beta subunits that function as an alpha 2 beta 2 heterotetrameric receptor complex. Provoked by this major structural difference in two receptors that carry out parallel functions, we have designed a chimaeric receptor molecule comprising the extracellular portion of the insulin receptor joined to the transmembrane and intracellular domains of the EGF receptor to investigate whether one ligand will activate the tyrosine kinase domain of the receptor for the other ligand. We show here that the EGF receptor kinase domain of the chimaeric protein, expressed transiently in simian cells, is activated by insulin binding. This strongly suggests that insulin and EGF receptors employ closely related or identical mechanisms for signal transduction across the plasma membrane.  相似文献   

5.
M S Collett  A F Purchio  R L Erikson 《Nature》1980,285(5761):167-169
The protein responsible for malignant transformation by avian sarcoma viruses (ASVs) has been identified as a phosphoprotein of molecular weight 60,000 designated pp60src (refs 1--4). It has been suggested that this protein has a functional role in cellular transformation involving the phosphorylation of cellular proteins, for it was discovered that specific immunoprecipitates from ASV-transformed cells that contain pp60src catalysed the transfer of phosphate from [gamma-32P]ATP to the heavy chain of rabbit immunoglobulin. Additional studies involving the cell-free synthesis of the ASV src protein further demonstrated that the presence of the src polypeptide correlated with that presence of a phosphotransferase activity. Our studies, involving the biochemical purification of this protein, have demonstrated that the ASV-transforming gene product, pp60src, is itself a protein kinase. We have purified the pp60src protein approximately 5,000-fold using either conventional ion-exchange chromatography or immunoaffinity chromatography. The resultant partially purified preparations contain a cyclic AMP-independent protein kinase activity. We report here that the soluble phosphotransferase activity of partially purified pp60src results in the phosphorylation of exclusively tyrosine residues in a variety of proteins that serve as substrates.  相似文献   

6.
D R Kaplan  D Martin-Zanca  L F Parada 《Nature》1991,350(6314):158-160
Nerve growth factor (NGF) is a neurotrophic factor responsible for the differentiation and survival of sympathetic and sensory neurons as well as selective populations of cholinergic neurons. NGF binds to specific cell-surface receptors but the mechanism for transduction of the neurotrophic signal is unknown. Several experiments using the NGF-responsive pheochromocytoma cell line, PC12, have implicated tyrosine phosphorylation in NGF-mediated responses, although no NGF-specific tyrosine kinases have been identified. Here we show that NGF induces tyrosine phosphorylation and tyrosine kinase activity of the trk proto-oncogene product, a tyrosine kinase receptor whose expression is restricted in vivo to neurons of the sensory spinal and cranial ganglia of neural crest origin. Tyrosine phosphorylation of trk by NGF is rapid, specific and occurs with picomolar quantities of factor, indicating that the response is mediated by physiological amounts of NGF. Activation of the trk tyrosine kinase receptor provides a possible mechanism for signal transduction by NGF.  相似文献   

7.
酪氨酸激酶Btk是非受体酪氨酸家族的成员,它由PH结构域、TH结构域、SH3结构域、SH2结构域和催化结构域5部分组成.Btk参与多种信号通路,对细胞的增殖、分化和凋亡起着重要的调控作用.Btk的突变可导致X连锁无丙种球蛋白血症,一直以来都是研究热点.笔者将围绕Btk的结构、功能、X连锁无丙种球蛋白血症的临床表现等方面的内容加以综述,着重探讨Btk参与B细胞信号通路、TLR信号通路和肥大细胞脱颗粒等过程的具体机制.  相似文献   

8.
Ecto-protein kinase activity on the external surface of neural cells   总被引:14,自引:0,他引:14  
Y H Ehrlich  T B Davis  E Bock  E Kornecki  R H Lenox 《Nature》1986,320(6057):67-70
ATP is secreted in association with neurotransmitters at certain synapses and neuromuscular junctions. Extracellular ATP is known to exert potent effects on the activity of cells in the nervous system, where it can act as a neurotransmitter or as a modulator regulating the activity of other neurohormones. We have suggested that such modulation may involve the activity of extracellular protein phosphorylation systems. It is well known that intracellular protein kinases are important in the regulation of various neuronal functions, but protein kinases which use extracellular ATP to phosphorylate proteins localized at the external surface of the plasma membrane (ecto-protein kinases) have not been demonstrated in neuronal cells. Here we present direct evidence for the existence of an ecto-protein kinase and demonstrate endogenous substrates for its activity at the surface of intact neural cells. The phosphorylation of one of these surface proteins is selectively stimulated during cell depolarization. In addition, neuronal cell adhesion molecules (N-CAMs) appear to be among the substrates of ecto-protein kinase activity. These results suggest a role for surface protein phosphorylation in regulating specific functions of developing and mature neurones.  相似文献   

9.
cdc2 is a catalytic subunit of a protein kinase complex, called the M-phase promoting factor, that induces entry into mitosis and is universal among eukaryotes. In HeLa cells, cdc2 is shown to be the most abundant phosphotyrosine-containing protein and its phosphotyrosine content is subject to cell-cycle regulation. One site of cdc2 tyrosine phosphorylation in vivo is selectively phosphorylated by pp60c-src in vitro.  相似文献   

10.
11.
T Itoh  M Ikebe  G J Kargacin  D J Hartshorne  B E Kemp  F S Fay 《Nature》1989,338(6211):164-167
Phosphorylation of myosin light chains by a calmodulin-myosin light-chain kinase (MLCK) pathway is considered to be responsible for coupling increased calcium concentration with contraction in smooth muscle. This simple view has, however, recently been questioned. To test this hypothesis directly, we microinjected individual smooth muscle cells with modulators of the MLCK pathway while measuring contraction and calcium-ion concentration. Injection of a constitutively active proteolyzed form of MLCK causes contraction but no change in calcium concentration. By contrast, injection of peptide inhibitors of MLCK blocks contraction in response to K+ depolarization, despite the fact that the change in calcium concentration in response to stimulation was enhanced over controls. These results provide a direct demonstration at the level of a single cell that activation of the calmodulin-MLCK pathway is both necessary and sufficient to trigger contraction of smooth muscle.  相似文献   

12.
13.
T J O'Dell  E R Kandel  S G Grant 《Nature》1991,353(6344):558-560
Long-term potentiation (LTP) in the hippocampus is thought to contribute to memory formation. In the Ca1 region, LTP requires the NMDA (N-methyl-D-aspartate) receptor-dependent influx of Ca2+ and activation of serine and threonine protein kinases. Because of the high amount of protein tyrosine kinases in hippocampus and cerebellum, two regions implicated in learning and memory, we examined the possible additional requirement of tyrosine kinase activity in LTP. We first examined the specificity in brain of five inhibitors of tyrosine kinase and found that two of them, lavendustin A and genistein, showed substantially greater specificity for tyrosine kinase from hippocampus than for three serine-threonine kinases: protein kinase A, protein kinase C, and Ca2+/calmodulin kinase II. Lavendustin A and genistein selectively blocked the induction of LTP when applied in the bath or injected into the postsynaptic cell. By contrast, the inhibitors had no effect on the established LTP, on normal synaptic transmission, or on the neurotransmitter actions attributable to the actions of protein kinase A or protein kinase C. These data suggest that tyrosine kinase activity could be required postsynaptically for long-term synaptic plasticity in the hippocampus. As Ca2+ calmodulin kinase II or protein kinase C seem also to be required, the tyrosine kinases could participate postsynaptically in a kinase network together with serine and threonine kinases.  相似文献   

14.
The Drosophila gene torso encodes a putative receptor tyrosine kinase   总被引:27,自引:0,他引:27  
The maternal gene torso, required for determination of anterior and posterior terminal structures in the Drosophila embryo, was cloned using P-element tagging. Genetic evidence suggests that the action of the gene product is spatially restricted to the terminal regions; the torso messenger RNA, however, is evenly distributed. Structural similarities of the predicted torso protein with growth-factor receptor tyrosine kinases suggest that the spatial restriction of torso activity results from a localized activation of the torso protein at the anterior and posterior egg pole.  相似文献   

15.
Tyrosine-dependent increase of tyrosine hydroxylase in neuroblastoma cells   总被引:3,自引:0,他引:3  
T Lloyd  X O Breakefield 《Nature》1974,252(5485):719-720
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16.
The human met oncogene is related to the tyrosine kinase oncogenes   总被引:3,自引:0,他引:3  
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17.
A non-radioisotopic method was developed for the assay of epidermal growth factor receptor (EGFR). A peptide with twenty amino acid residues around Tyr 1173, the major phosphorylation site of EGFR, was cloned as a GST fusion protein and used as substrate. Anti-phosphoty-rosine monoclonal antibody PY99 was used for the determination of the extent of phosphorylation. Both the specificity and the sensitivity were substantially higher than that of the existing method. Km value of the fusion protein is much lower (10 μmol/L) than that of the synthetic peptide (110 μmol/L). The method can be applied to the measurement of the tyrosine kinase activity of c-erb B2 (Neu/HER2).  相似文献   

18.
19.
Malignant melanoma in Xiphophorus fish hybrids is caused by the activity of a dominant oncogene Tu. By combining genetic and molecular approaches, we have isolated the melanoma oncogene. We show that its level of expression correlates with the degree of malignancy of the tumour. The corresponding proto-oncogene is developmentally regulated. The Tu gene codes for a novel receptor tyrosine kinase which is closely related to the receptor for epidermal growth factor.  相似文献   

20.
Obese syndromes of genetic origin or experimentally induced are characterized by resistance to insulin both in vivo (association of hyperglycaemia and hyperinsulinaemia) and in vitro. Thus, skeletal muscle of obese mice, which is the most important target organ for the action of insulin, displays a reduced response to insulin. This hormonal resistance cannot be explained by the moderate decrease in the number of insulin receptors found in obese animals. In fact, it is generally believed that a biochemical event occurring very early after binding of insulin to its receptor, which is the first step in insulin action, is defective in obesity. One of the earliest post-binding events so far recognized, and which is thought to have a key role in cellular signalling by the insulin receptor, is the insulin-stimulated phosphorylation of its receptor. In an effort to localize the defect responsible for the insulin resistance in obesity, we have studied the insulin receptor protein kinase activity and we show here that insulin receptors from skeletal muscles of insulin-resistant obese mice have an altered kinase activity for phosphorylation of both the receptor itself and of exogeneous substrates.  相似文献   

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