Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “TYROSINE”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,063 records · Page 59Linked to original sources
PubMed1993

Zymosan-induced tyrosine phosphorylations in human monocytes. Role of protein kinase C.

Protein tyrosine phosphorylations are involved in the proliferation and secretory responses of immune cells, but their role in phagocytes is poorly understood. The ability of unopsonized zymosan to induce protein tyrosine phosphorylations was investigated in human monocytes. The addition of zymosan to monocytes resulted in an increase in tyrosine phosphorylation of several endogenous proteins including 28-, 33-, 38-, 42-, 47-, 55- to 60-, 62-, 68-, 90-, 105-, 116-, and 120-kDa proteins; 55- to 60-kDa proteins were the predominant phosphoproteins. Moreover, we studied the effects of tyrphostin 23, a specific tyrosine kinase inhibitor, on stimulated tyrosine phosphorylations and early secretory responses of monocytes, i.e., arachidonic acid release and oxidative metabolism. We showed that tyrphostin inhibited zymosan-stimulated tyrosine phosphorylations and arachidonic acid release, but that it did not affect superoxide generation induced by zymosan. Zymosan binds mainly to CR3 receptor on human monocytes, and CR3 is devoid of intrinsic tyrosine kinase activity. It was predictable that zymosan stimulated a tyrosine kinase distal to the receptor or associated with it. We observed that PMA mimicked zymosan-induced tyrosine phosphorylations, thus suggesting that both agonists used a common transductional pathway implicating the serine/threonine kinase, protein kinase C. The antagonists of protein kinase C, sphingosine and calphostin C, inhibited zymosan-stimulated tyrosine phosphorylations. We suggest that, in human monocytes, zymosan-induced tyrosine phosphorylations are involved in cell responses such as the release of arachidonic acid, and that they require the sequential activation of protein kinase C and cellular protein tyrosine kinases.

Catechols↗
PubMed1998

Redundant and selective roles for erythropoietin receptor tyrosines in erythropoiesis in vivo.

Cytokine receptors have been shown in cell culture systems to use phosphotyrosine residues as docking sites for certain signal transduction intermediates. Studies using various cellular backgrounds have yielded conflicting information about the importance of such residues. The present studies were undertaken to determine whether or not tyrosine residues within the erythropoietin receptor (EPOR) are essential for biologic activity during hematopoiesis in vivo. A variant of the EPOR was constructed that contains both a substitution (R129C) causing constitutive receptor activation as well as replacement of all eight cytoplasmic tyrosines by phenylalanines (cEPORYF). A comparison between animals exposed to recombinant retroviruses expressing cEPOR and cEPORYF showed that efficient red blood cell (RBC) development in vivo is dependent on the pressence of tyrosine residues in the cytoplasmic domain of the EPOR. In addition, an inefficient EPOR tyrosine independent pathway supporting RBC development was detected. Tyrosine add-back mutants showed that multiple individual tyrosines have the capacity to restore full erythropoietic potential to the EPOR as determined in whole animals. The analysis of primary erythroid progenitors transduced with the various cEPOR tyrosine mutants and tyrosine add-backs showed that only tyrosine 343 (Y1) and tyrosine 479 (Y8) were capable of supporting immature burst-forming unit-erythroid progenitor development. Thus, this receptor is characterized by striking functional redundancy of tyrosines in a biologically relevant context. However, selective tyrosine residues may be uniquely important for early signals supporting erythroid development.

Animals↗
PubMed2000

Cellular stress induces the tyrosine phosphorylation of caveolin-1 (Tyr(14)) via activation of p38 mitogen-activated protein kinase and c-Src kinase. Evidence for caveolae, the actin cytoskeleton, and focal adhesions as mechanical sensors of osmotic stress.

Environmental stressors have been recently shown to activate intracellular mitogen-activated protein (MAP) kinases, such as p38 MAP kinase, leading to changes in cellular functioning. However, little is known about the downstream elements in these signaling cascades. In this study, we show that caveolin-1 is phosphorylated on tyrosine 14 in NIH 3T3 cells after stimulation with a variety of cellular stressors (i.e. high osmolarity, H2O2, and UV light). To detect this phosphorylation event, we employed a phosphospecific monoclonal antibody probe that recognizes only tyrosine 14-phosphorylated caveolin-1. Since p38 MAP kinase and c-Src have been previously implicated in the stress response, we next assessed their role in the tyrosine phosphorylation of caveolin-1. Interestingly, we show that the p38 inhibitor (SB203580) and a dominant-negative mutant of c-Src (SRC-RF) both block the stress-induced tyrosine phosphorylation of caveolin-1 (Tyr(P)(14)). In contrast, inhibition of the p42/44 MAP kinase cascade did not affect the tyrosine phosphorylation of caveolin-1. These results indicate that extracellular stressors can induce caveolin-1 tyrosine phosphorylation through the activation of well established upstream elements, such as p38 MAP kinase and c-Src kinase. However, heat shock did not promote the tyrosine phosphorylation of caveolin-1 and did not activate p38 MAP kinase. Finally, we show that after hyperosmotic shock, tyrosine-phosphorylated caveolin-1 is localized near focal adhesions, the major sites of tyrosine kinase signaling. In accordance with this localization, disruption of the actin cytoskeleton dramatically potentiates the tyrosine phosphorylation of caveolin-1. Taken together, our results clearly define a novel signaling pathway, involving p38 MAP kinase activation and caveolin-1 (Tyr(P)(14)). Thus, tyrosine phosphorylation of caveolin-1 may represent an important downstream element in the signal transduction cascades activated by cellular stress.

3T3 Cells↗
PubMed2003

Activation of SRC tyrosine kinases in response to ICAM-1 ligation in pulmonary microvascular endothelial cells.

Previous studies demonstrated that ICAM-1 ligation on human pulmonary microvascular endothelial cells (ECs) sequentially induces activation of xanthine oxidase and p38 MAPK. Inhibition of these signaling events reduces neutrophil migration to the EC borders. This study examined the role of SRC tyrosine kinases in ICAM-1-initiated signaling within these ECs. Cross-linking ICAM-1 on tumor necrosis factor-alpha-pretreated ECs induced an increase in the activity of SRC tyrosine kinases. This increase was inhibited by allopurinol (a xanthine oxidase inhibitor), Me2SO (a hydroxyl radical scavenger), or deferoxamine (an iron chelator). Phenylarsine oxide, a tyrosine phosphatase inhibitor, reduced the base-line activity of SRC as well as the increase in SRC activity induced by ICAM-1 cross-linking. Specific inhibition of the protein expression of the SRC homology 2-containing protein-tyrosine phosphatase-2 (SHP-2) by an antisense oligonucleotide prevented the induced SRC activation but had no effect on the basal SRC activity. Activation of SRC tyrosine kinases was accompanied by tyrosine phosphorylation of ezrin at Tyr-146, which was inhibited by PP2, an SRC tyrosine kinase inhibitor. Moreover, PP2 completely inhibited p38 activation, suggesting a role for SRC tyrosine kinases in p38 activation. These data demonstrate that ICAM-1 ligation activates SRC tyrosine kinases and that this activation requires SHP-2 as well as production of reactive oxygen species generated from xanthine oxidase. Activation of SRC tyrosine kinases in turn leads to tyrosine phosphorylation of ezrin, as well as activation of p38, a kinase previously identified to be required for cytoskeletal changes induced by ICAM-1 ligation and for neutrophil migration along the EC surface.

Allopurinol↗
PubMed2005

Receptor-type protein-tyrosine phosphatase-kappa regulates epidermal growth factor receptor function.

Epidermal growth factor receptor (EGFR), the prototypic receptor protein tyrosine kinase, is a major regulator of growth and survival for many epithelial cell types. We report here that receptor-type protein-tyrosine phosphatase-kappa (RPTP-kappa) dephosphorylates EGFR and thereby regulates its function in human keratinocytes. Protein-tyrosine phosphatase (PTP) inhibitors induced EGFR tyrosine phosphorylation in intact primary human keratinocytes and cell-free membrane preparations. Five highly expressed RPTPs (RPTP-beta, delta, kappa, mu, and xi) were functionally analyzed in a Chinese hamster ovary (CHO) cell-based expression system. Full-length human EGFR expressed in CHO cells, which lack endogenous EGFR, displayed high basal (i.e. in the absence of ligand) tyrosine phosphorylation. Co-expression of RPTP-kappa, but not other RPTPs, specifically reduced basal EGFR tyrosine phosphorylation. RPTP-kappa also reduced epidermal growth factor-dependent EGFR tyrosine phosphorylation in CHO cells. Purified RPTP-kappa preferentially dephosphorylated EGFR tyrosines 1068 and 1173 in vitro. Overexpression of wild-type or catalytically inactive RPTP-kappa reduced or enhanced, respectively, basal and EGF-induced EGFR tyrosine phosphorylation in human keratinocytes. Furthermore, siRNA-mediated knockdown of RPTP-kappa increased basal and EGF-stimulated EGFR tyrosine phosphorylation and augmented downstream Erk activation in human keratinocytes. RPTP-kappa levels increased in keratinocytes as cells reached confluency, and overexpression of RPTP-kappa in subconfluent keratinocytes reduced keratinocyte proliferation. Taken together, the above data indicate that RPTP-kappa is a key regulator of EGFR tyrosine phosphorylation and function in human keratinocytes.

Animals↗
PubMed2000

Functional regulation of gamma-aminobutyric acid transporters by direct tyrosine phosphorylation.

Tyrosine phosphorylation regulates multiple cell signaling pathways and functionally modulates a number of ion channels and receptors. Neurotransmitter transporters, which act to clear transmitter from the synaptic cleft, are regulated by multiple second messenger pathways that exert their effects, at least in part, by causing a redistribution of the transporter protein to or from the cell surface. To test the hypothesis that tyrosine phosphorylation affects transporter function and to determine its mechanism of action, we examined the regulation of the rat brain gamma-aminobutyric acid (GABA) transporter GAT1 expressed endogenously in hippocampal neurons and expressed heterologously in Chinese hamster ovary cells. Inhibitors of tyrosine kinases decreased GABA uptake; inhibitors of tyrosine phosphatases increased GABA uptake. The decrease in uptake seen with tyrosine kinase inhibitors was correlated with a decrease in tyrosine phosphorylation of GAT1 and resulted in a redistribution of the transporter from the cell surface to intracellular locations. A mutant GAT1 construct that was refractory to tyrosine phosphorylation could not be regulated by tyrosine kinase inhibitors. Activators of protein kinase C, which are known to cause a redistribution of GAT1 from the cell surface, were additive to the effects of tyrosine kinase inhibitors suggesting that multiple signaling pathways control transporter redistribution. Application of brain-derived neurotrophic factor, which activates receptor tyrosine kinases, up-regulated GAT1 function suggesting one potential trigger for the cellular regulation of GAT1 signaling by tyrosine phosphorylation. These data support the hypothesis that transporter expression and function is controlled by the interplay of multiple cell signaling cascades.

Animals↗
PubMed1994

The role of protein tyrosine phosphorylation in integrin-mediated gene induction in monocytes.

Integrin-mediated cell adhesion, or cross-linking of integrins using antibodies, often results in the enhanced tyrosine phosphorylation of certain intracellular proteins, suggesting that integrins may play a role in signal transduction processes. In fibroblasts, platelets, and carcinoma cells, a novel tyrosine kinase termed pp125FAK has been implicated in integrin-mediated tyrosine phosphorylation. In some cell types, integrin ligation or cell adhesion has also been shown to result in the increased expression of certain genes. Although it seems reasonable to hypothesize that integrin-mediated tyrosine phosphorylation and integrin-mediated gene induction are related, until now, there has been no direct evidence supporting this hypothesis. In the current report, we explore the relationship between integrin-mediated tyrosine phosphorylation and gene induction in human monocytes. We demonstrate that monocyte adherence to tissue culture dishes or to extracellular matrix proteins is followed by a rapid and profound increase in tyrosine phosphorylation, with the predominant phosphorylated component being a protein of 76 kD (pp76). Tyrosine phosphorylation of pp76 and other monocyte proteins can also be triggered by incubation of monocytes with antibodies to the integrin beta 1 subunit, or by F(ab')2 fragments of such antibodies, but not by F(ab) fragments. The ligation of beta 1 integrins with antibodies or F(ab')2 fragments also induces the expression of immediate-early (IE) genes such as IL-1 beta. When adhering monocytes are treated with the tyrosine kinase inhibitors genistein or herbimycin, both phosphorylation of pp76 and induction of IL-1 beta message are blocked in a dose-dependent fashion. Similarly, treatment with genistein or herbimycin can block tyrosine phosphorylation of pp76 and IL-1 beta message induction mediated by ligation of beta 1 integrin with antibodies. These observations suggest that protein tyrosine phosphorylation is an important aspect of integrin-mediated IE gene induction in monocytes. The cytoplasmic tyrosine kinase pp125FAK, although important in integrin signaling in other cell types, seems not to play a role in monocytes because this protein could not be detected in these cells.

Benzoquinones↗
PubMed2001

Analysis of tyrosine phosphorylation-dependent protein-protein interactions in TrkB-mediated intracellular signaling using modified yeast two-hybrid system.

Activated receptor tyrosine kinases induce a large number of tyrosine phosphorylation-dependent protein-protein interactions through which they mediate their various ligand-exerted functions including regulation of proliferation, differentiation and survival. TrkB receptor tyrosine kinase activated by binding of brain-derived neurotrophic factor (BDNF) also stimulates various protein interactions in a tyrosine phosphorylation-dependent manner in neuronal cells. To examine tyrosine phosphorylation-dependent interactions stimulated by active TrkB, we developed a modified yeast two-hybrid system, which we call the yeast two-and-a-half-hybrid system. In this system, yeast was engineered to express a tyrosine kinase domain of TrkB as an effector, in addition to two fusion proteins with GAL4 DNA-binding and GAL4 activation domains as bait and prey proteins, respectively. Using this system with Shp2 as the bait, we demonstrated that Shp2 interacts directly with BIT/SHPS-1 (also called SIRP) and Grb2 depending on tyrosine phosphorylation mediated by TrkB. Furthermore, we screened an adult human brain cDNA library with the yeast two-and-a-half-hybrid system in order to identify other Shp2-binding proteins in TrkB-stimulated tyrosine phosphorylation signaling. We found that fibroblast growth factor receptor substrate 2beta (FRS2beta), also called SNT2, interacts with Shp2 dependently on TrkB-mediated tyrosine phosphorylation of FRS2beta/SNT2. Therefore, we show that the two-and-a-half-hybrid system is a powerful tool for studying tyrosine phosphorylation-dependent protein-protein interactions in intracellular signaling pathways stimulated by TrkB receptor tyrosine kinase.

Adaptor Proteins, Signal Transducing↗
PubMed1983

Decarboxylation of p-tyrosine: a potential source of p-tyramine in mammalian tissues.

The question of the existence of a p-tyrosine decarboxylase pathway for the formation of p-tyramine in mammalian tissues remains unresolved. Development of a sensitive and specific assay for p-tyrosine decarboxylase has permitted demonstration of this activity in rat tissues and human kidney. Tyrosine decarboxylase was purified to electrophoretic homogeneity by pH 5.0 precipitation, ammonium sulfate precipitation, gel filtration, phenyl-Sepharose chromatography, DEAE-Sephacel chromatography, and preparative isoelectric focusing. A specific rabbit antiserum to tyrosine decarboxylase was also obtained. Purified tyrosine decarboxylase possessed a narrow pH dependency with an optimum at 8.0. Benzene and certain other organic solvents dramatically stimulated tyrosine decarboxylase activity of purified enzyme. Purified tyrosine decarboxylase activity also decarboxylated L-DOPA, 5-hydroxytryptophan, 3,4-dihydroxyphenylserine, o-tyrosine, m-tyrosine, phenylalanine, histidine, and tryptophan, which suggested that the purified enzyme was aromatic L-amino acid decarboxylase. This conclusion was supported by a constant ratio of 5-hydroxytryptophan decarboxylase to tyrosine decarboxylase throughout the purification scheme and by parallel immunoprecipitation of decarboxylase activities by the specific antityrosine decarboxylase antisera. Thus, we report that p-tyrosine is decarboxylated by aromatic L-amino acid decarboxylase and that this metabolic transformation may be an important source of p-tyramine in mammalian tissues. In conclusion, neuronal tissues that synthesize catecholamines or serotonin should now be considered capable of synthesizing p-tyramine and other biogenic amines.

Animals↗
PubMed1999

Growth stimulation by serum in Entamoeba histolytica is associated with protein tyrosine dephosphorylation.

Very little protein tyrosine phosphorylation was observed in growing (exponential-phase) Entamoeba histolytica cells by immunoblotting and quantitative immunofluorescence. After 1 h of serum deprivation, two proteins (42 and 38 kDa in SDS-PAGE) were tyrosine phosphorylated and two more proteins (96 and 63 kDa) also showed tyrosine phosphorylation when examined after 4 h of serum deprivation. Intense enhancements of anti-phosphotyrosine immunofluorescence levels were observed during this period of serum withdrawal. Membrane-associated tyrosine kinase activity reached a peak (3.5-fold increase) 1 h after serum deprivation and decreased thereafter reaching a basal level by 2 h of serum deprivation. Interestingly, tyrosine kinase activities remained unaffected by serum stimulation (2-60 min) of serum-deprived cells. Also, during this period of serum stimulation tyrosine phosphorylated proteins of serum-deprived cells were dephosphorylated. Tyrosine phosphatase activities were suppressed during serum deprivation and on serum addition to serum-deprived cells tyrosine phosphatase activities increased significantly. Our data attest that protein tyrosine phosphorylation was associated with growth inhibition of E. histolytica and serum stimulation of E. histolytica produced tyrosine phosphatase activation and protein tyrosine dephosphorylation.

Animals↗
PubMed1995

Endothelin-1 stimulates tyrosine phosphorylation of p125 focal adhesion kinase in mesangial cells.

Endothelin-1 (ET-1) is known to induce the contraction and proliferation of glomerular mesangial cells. Because ET-1 was found to stimulate the tyrosine phosphorylation of unidentified cellular proteins in cultured mesangial cells, protein tyrosine kinase might serve as one of the important signals leading to various functions of ET-1. Focal adhesion kinase (p125FAK) is a newly identified cytoplasmic protein tyrosine kinase that is activated by the phosphorylation of its own tyrosine residue. Because p125FAK was found to play a role in the signal transduction of not only integrins but also various neurotransmitters, including bombesin, endothelin, and vasopressin in Swiss 3T3 cells and Rat-1 fibroblasts, whether ET-1 could stimulate the tyrosine phosphorylation of p125FAK in glomerular mesangial cells was examined. ET-1 stimulated the tyrosine phosphorylation of p125FAK by threefold to fourfold in cultured mesangial cells. This effect of ET-1 was detected at 1 min and reached a maximum within 5 min and was blocked by BQ-123, an antagonist for ETA receptor. A23187, a calcium ionophore, failed to stimulate the tyrosine phosphorylation of p125FAK, and ET-1 was able to stimulate the tyrosine phosphorylation of p125FAK, even in a calcium-free medium. The activation of protein kinase C (PKC) by phorbol 12, 13-dibutyrate resulted in a stimulation of the tyrosine phosphorylation of p125FAK, and an inhibition of PKC by calphostin C or staurosporine significantly reduced the effect of ET-1. Furthermore, prolonged treatment of the cells with phorbol 12, 13-dibutyrate markedly inhibited the ET-1-induced tyrosine phosphorylation of p125FAK. These results indicate that p125FAK might play a role in a signal transduction system of ET-1 in glomerular mesangial cells and that the ET-1-induced tyrosine phosphorylation of p125FAK is largely dependent on the PKC pathway.

Animals↗
PubMed1994

Sphingosine induces p125FAK and paxillin tyrosine phosphorylation, actin stress fiber formation, and focal contact assembly in Swiss 3T3 cells.

Treatment of Swiss 3T3 cells with sphingosine, a potential breakdown product of all sphingolipids, induced tyrosine phosphorylation of multiple substrates including bands of M(r) 110,000-130,000 and M(r) 70,000-80,000. Tyrosine phosphorylation in response to sphingosine occurred in a concentration dependent manner (EC50 = 10 microM) and developed gradually reaching half maximum and maximum effects at 20 and 60 min, respectively. The dihydroenantiomere of sphingosine, DL-threo-dihydrosphingosine, neither induced tyrosine phosphorylation nor interfered with sphingosine-stimulated tyrosine phosphorylation. Focal adhesion kinase (p125FAK) and paxillin were identified as prominent substrates for sphingosine-stimulated tyrosine phosphorylation. Cell permeable ceramides also stimulated tyrosine phosphorylation of the M(r) 110,000-130,000 band as well as p125FAK, but the effect was less pronounced than that of sphingosine. Tyrosine phosphorylation by sphingosine could be dissociated from both protein kinase C activation and Ca2+ mobilization from intracellular stores. Sphingosine stimulated striking actin stress fiber formation and focal adhesion assembly in Swiss 3T3 cells. The kinetics of actin stress fiber formation and tyrosine phosphorylation in response to sphingosine closely paralleled. Cytochalasin D, which disrupts the network of actin microfilaments, completely inhibited sphingosine induced tyrosine phosphorylation. In addition, tyrosine phosphorylation of p125FAK and paxillin in response to sphingosine was completely prevented when cells were stimulated in the presence of platelet-derived growth factor at a concentration (30 ng/ml) that caused disruption of the actin cytoskeleton. Our results demonstrate, for the first time, that sphingosine induces p125FAK and paxillin tyrosine phosphorylation, actin stress fiber formation and focal adhesion assembly in Swiss 3T3 cells.

3T3 Cells↗
PubMed1994

Angiotensin II-induced protein tyrosine phosphorylation in neonatal rat cardiac fibroblasts.

Angiotensin II has been demonstrated to act as a growth factor in rat cardiac fibroblasts. However, the signaling events that lead to fibroblast cell growth in response to angiotensin II remain to be elucidated. This study was designed to determine whether angiotensin II stimulated tyrosine phosphorylation of proteins in cardiac fibroblasts. Immunoblot analysis demonstrated rapid tyrosine phosphorylation of distinct substrates of 125, 95, 46-60, and 44 kDa in response to 10 nM angiotensin II. Tyrosine phosphorylation was maximal at 5 min and persisted for at least 180 min. Additional tyrosine-phosphorylated proteins of 185, 145, and 85 kDa were detected in response to 10 ng/ml platelet-derived growth factor BB. A cluster of 75-80-kDa proteins were phosphorylated in response to angiotensin II, phorbol ester, and platelet-derived growth factor. Angiotensin II-induced tyrosine phosphorylation was unaffected by phorbol ester-sensitive protein kinase C down-regulation and could be partially blocked by pertussis toxin pretreatment. Angiotensin II stimulation resulted in increased cytosolic tyrosine kinase activity which was recovered by immunoprecipitation. Immunoblot analysis demonstrated tyrosine phosphorylation of p44MAPK, and, in addition, we demonstrated for the first time tyrosine phosphorylation of p125FAK, p46SHC, and p56SHC in response to angiotensin II. The finding that angiotensin II and platelet-derived growth factor stimulated tyrosine phosphorylation of p46SHC and p56SHC suggested that this protein may serve as a common tyrosine kinase substrate in the mitogenic signaling cascade induced by G-protein-coupled receptors and growth factors and is consistent with the hypothesis that angiotensin II-induced tyrosine phosphorylation is involved in mitogenic signaling pathways in neonatal rat cardiac fibroblasts.

Adaptor Proteins, Signal Transducing↗
PubMed1998

SHP-1 binds and negatively modulates the c-Kit receptor by interaction with tyrosine 569 in the c-Kit juxtamembrane domain.

The SH2 domain-containing SHP-1 tyrosine phosphatase has been shown to negatively regulate a broad spectrum of growth factor- and cytokine-driven mitogenic signaling pathways. Included among these is the cascade of intracellular events evoked by stem cell factor binding to c-Kit, a tyrosine kinase receptor which associates with and is dephosphorylated by SHP-1. Using a series of glutathione S-transferase (GST) fusion proteins containing either tyrosine-phosphorylated segments of the c-Kit cytosolic region or the SH2 domains of SHP-1, we have shown that SHP-1 interacts with c-Kit by binding selectively to the phosphorylated c-Kit juxtamembrane region and that the association of c-Kit with the larger of the two SHP-1 isoforms may be mediated through either the N-terminal or C-terminal SHP-1 SH2 domain. The results of binding assays with mutagenized GST-Kit juxtamembrane fusion proteins and competitive inhibition assays with phosphopeptides encompassing each c-Kit juxtamembrane region identified the tyrosine residue at position 569 as the major site for binding of SHP-1 to c-Kit and suggested that tyrosine 567 contributes to, but is not required for, this interaction. By analysis of Ba/F3 cells retrovirally transduced to express c-Kit receptors, phenylalanine substitution of c-Kit tyrosine residue 569 was shown to be associated with disruption of c-Kit-SHP-1 binding and induction of hyperproliferative responses to stem cell factor. Although phenylalanine substitution of c-Kit tyrosine residue 567 in the Ba/F3-c-Kit cells did not alter SHP-1 binding to c-Kit, the capacity of a second c-Kit-binding tyrosine phosphatase, SHP-2, to associate with c-Kit was markedly reduced, and the cells again showed hyperproliferative responses to stem cell factor. These data therefore identify SHP-1 binding to tyrosine 569 on c-Kit as an interaction pivotal to SHP-1 inhibitory effects on c-Kit signaling, but they indicate as well that cytosolic protein tyrosine phosphatases other than SHP-1 may also negatively regulate the coupling of c-Kit engagement to proliferation.

Cell Membrane↗
PubMed2003

Kinetic analysis of the 4-methylideneimidazole-5-one-containing tyrosine aminomutase in enediyne antitumor antibiotic C-1027 biosynthesis.

The enediyne antitumor antibiotic C-1027 contains an unusual (S)-3-chloro-4,5-dihydroxy-beta-phenylalanine moiety, which requires an aminomutase for its biosynthesis. Previously, we established that SgcC4 is an aminomutase that catalyzes the conversion of L-tyrosine to (S)-beta-tyrosine and employs 4-methylideneimidazole-5-one (MIO) at its active site [Christenson, S. D., Liu, W., Toney, M. D., and Shen, B. (2003) J. Am. Chem. Soc. 125, 6062-6063]. Here, we present a thorough analysis of the properties of SgcC4. L-Tyrosine is the best substrate among those tested and most likely serves as the in vivo precursor for the (S)-3-chloro-4,5-dihydroxy-beta-phenylalanine moiety. The presence of MIO in the active site is supported by several lines of evidence. (1) Addition of ATP or divalent metal ions has no effect on its aminomutase activity. (2) SgcC4 has optimal activity at pH approximately 8.8, similar to the pH optima of MIO-dependent ammonia lyases. (3) SgcC4 is strongly inhibited by sodium borohydride and potassium cyanide, but preincubation with L-tyrosine or 4-hydroxycinnamate largely prevents this inhibition. (4) The difference spectrum between SgcC4 and its S153A mutant shows a positive peak at approximately 310 nm, indicative of MIO. (5) The S153A mutation lowers k(cat)/K(M) 640-fold. The SgcC4-catalyzed conversion of L-tyrosine to (S)-beta-tyrosine proceeds via 4-hydroxycinnamate as an intermediate. The latter also acts as a competitive inhibitor with respect to L-tyrosine and serves as an alternative substrate for the production of beta-tyrosine in the presence of an amino source. A full time course for the SgcC4-catalyzed interconversion between L-tyrosine, beta-tyrosine, and 4-hydroxycinnamate was measured and analyzed to provide estimates for the rate constants in a minimal mechanism. SgcC4 also exhibits a beta-tyrosine racemase activity, but alpha-tyrosine racemase activity was not detected.

Amino Acid Sequence↗
PubMed2006

Increased striatal dopamine synthesis is associated with decreased tissue levels of tyrosine.

Tyrosine levels do not generally affect indices of dopamine (DA) synthesis or efflux under basal conditions, but can do so when DA synthesis is increased. One possibility is that a high rate of DA synthesis depletes the normally adequate pool of endogenous tyrosine. To study this, we administered drugs known to preferentially increase striatal DA synthesis and examined DOPA levels in striatal microdialysate during perfusion with NSD-1015. In additional groups, we also measured DA, tyrosine and large neutral amino acids in striatal microdialysate, as well as in tissue from striatum and medial prefrontal cortex (MPFC). gamma-butyrolactone (GBL) (750 mg/kg i.p.) increased DOPA levels in striatal microdialysate, increased tissue DA levels in the MPFC and striatum, but lowered tissue tyrosine levels only in striatum. In striatal microdialysate, GBL markedly lowered DA levels; tyrosine levels were only marginally lower. Haloperidol (HAL) (1.0 mg/kg s.c.)+/-amfonelic acid (AFA) (5 mg/kg i.p.) increased striatal DOPA accumulation, increased striatal DA efflux, lowered striatal tissue tyrosine levels, but did not affect microdialysate tyrosine levels. There were no consistent changes in levels of other large neutral amino acids. We conclude that increased tyrosine hydroxylation can significantly deplete the endogenous pool of tyrosine. Under such conditions, near normal extracellular tyrosine levels are maintained despite lower tissue levels. The data are consistent with a net transfer of tyrosine from non-DAergic cells to DA terminals in support of DA synthesis.

4-Butyrolactone↗
PubMed1994

Overexpression of the tyrosine phosphatase PTP1B is associated with human ovarian carcinomas.

OBJECTIVE: Our purpose was to determine whether protein tyrosine phosphatase 1B is overexpressed in ovarian cancers, possibly altering the balance of intracellular tyrosine phosphorylation. STUDY DESIGN: The expression of protein tyrosine phosphatase 1B was assayed in frozen sections from 54 human ovarian carcinomas and seven normal ovaries by immunochemical staining with monoclonal antibody AE4-2J, which is specific for protein tyrosine phosphatase 1B. The expression of protein tyrosine phosphatase 1B-specific messenger ribonucleic acid in tumors was determined by Northern analysis. The results were analyzed statistically by means of Fisher's exact test. RESULTS: Minimal staining was observed in normal ovarian epithelium. In contrast, 43 of 54 (79.6%) tumors displayed increased protein tyrosine phosphatase 1B expression, which is statistically associated with malignancy. Overexpression was associated with the expression of the p185c-erbB-2, p170EGFR, and p165mCSFR growth factor receptor protein tyrosine kinases. Protein tyrosine phosphatase 1B messenger ribonucleic acid expression was inconsistently increased in tumor cells. CONCLUSION: Increased expression of protein tyrosine phosphatase 1B in ovarian cancers that also express protein tyrosine kinases suggests that protein tyrosine phosphatase 1B may play a role in the growth regulation of ovarian cancers.

Female↗
PubMed1998

Cytokine-induced protein tyrosine phosphorylation is essential for cytokine priming of human eosinophils.

BACKGROUND: Human eosinophils are strongly modulated by the eosinophilotrophic cytokines IL-5, IL-3, and granulocyte-macrophage colony-stimulating factor (GM-CSF). A clear intracellular effect of these cytokines is the induction of tyrosine phosphorylation of multiple cellular substrates. However, the relevance of tyrosine phosphorylation for eosinophil functioning has not been established. OBJECTIVE: In this study we have investigated dose-response and time curves of IL-5-, IL-3-, and GM-CSF-induced tyrosine phosphorylation in eosinophils. Moreover, we have evaluated the importance of IL-5-induced tyrosine phosphorylation for priming of human eosinophils. METHODS: Cytokine-induced tyrosine phosphorylation was monitored on western blot with an antiphosphotyrosine antibody (4G10). To probe the relevance of tyrosine phosphorylation for priming, eosinophils were primed with IL-5 in the presence of the tyrosine kinase inhibitor herbimycin A. Platelet activating factor (PAF) was used as a control priming agent. Subsequently, the eosinophils were incubated with serum-treated zymosan (STZ) to activate the respiratory burst. Binding of STZ was determined by FACS analysis. RESULTS: IL-5-, IL-3-, and GM-CSF-induced tyrosine phosphorylation was found at concentrations that primed eosinophil effector mechanism (median effective dose values: approximately 5.10(-11) mol/L, approximately 5.10(-10) mol/L, and approximately 5.10(-12) mol/L for IL-5, IL-3, and GM-CSF, respectively). Cytokine-induced tyrosine phosphorylation was transient with an optimum value at 15 minutes. IL-5 priming of STZ-induced activation of the respiratory burst was blocked by herbimycin A, whereas PAF still primed this response. In fact, herbimycin A inhibited IL-5 priming of STZ binding to human eosinophils. On the other hand, PAF priming of STZ binding was not affected by herbimycin A. Both IL-5-induced and PAF-induced tyrosine phosphorylation were inhibited by herbimycin A. CONCLUSION: These data demonstrate for the first time that IL-5 priming of opsonized particle-induced responses is mediated by tyrosine kinase activity in human eosinophils.

Benzoquinones↗