Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.10.1 (ERK)
 95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The sialyl-fucosyl-lactosamine-epitope present in sialyl (SA)-Lex (NeuAc alpha 2-3Gal beta 1-4 [Fuc alpha 1-3]GlcNAc beta 1-3Gal beta 1-4Glc-Cer), a carcinoembryonic antigen, has been recognized recently as a ligand for the binding of leukocyte-endothelial cell adhesion molecule 1 (LECAM-1) to myeloid and tumour cell surfaces. We have recently detected the presence of an alpha 1-3 fucosyltransferase (FucT-3) activity in both embryonic chicken brain (ECB) and human colon carcinoma cells (Colo-205) which catalyses the biosynthesis in vitro of SA-Lex and SA-diLex. Fucosyltransferase activities from both sources are stimulated in the presence of divalent cations (Mn2+, Mg2+, Ca2+, Co2+ and Fe2+), although absolute metal requirement is not observed. Substrate specificity studies with this partially purified (ECB, 3000-fold; Colo-205, 100-fold) novel FucT-3 indicate the preference for terminally sialyl-substituted glycolipid acceptors, as observed by the lower Km values when sialyl-neolactotetraosyl ceramide, LM1, (Neu-Gc alpha 2-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4 Glc-Cer; Km = 0.048 mM) and sialyl-norhexaosylceramide, NeuGc-nLc6, (Neu-Gc alpha 2-3Gal beta 1-4 GlcNAc beta 1-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc-Cer; Km = 0.032 mM) were used as substrates. Fucosyltransferase from Colo-205 requires the presence of the acyl group of the ceramide moiety and an acetyl group on glucosamine in the acceptor glycolipid since lyso-LM1 was found to be completely inactive.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Biosynthesis in vitro of SA-Lex and SA-diLex by alpha 1-3 fucosyltransferases from colon carcinoma cells and embryonic brain tissues. 172 78

Adipocytic-cytosolic non-receptor protein tyrosine kinase (CytPTK) when activated can substitute for the insulin receptor tyrosine kinase (InsRTK), in manifesting several insulin effects in insulin-receptor independent fashion. Our aims here were to utilize PolyGlu4Tyr, a good experimental exogenous substrate for protein tyrosine kinases (PTKs) in general, for studying qualitative and quantitative parameters of CytPTKs extracted from different tissue cytosols. At the same time, we would search for a unique specific marker specifically characterizing CytPTKs. High speed supernatants of spleen, thymus, smooth muscle, lung and kidney were found to be rich in CytPTK activities. Their specific activities being 6- to 13-fold that of liver or adipose cytosols. Brain, testis and adrenal cytosols were an intermediate source of CytPTK activity, whereas CytPTK activity of heart and skeletal muscle was low. It was also evaluated that the capacity of the cytosol to phosphorylate PolyGlu4Tyr is 15-50% that of the non-stimulated Triton X-100 extractable plasma membrane PTKs. Fractionation of the cytosols on superose 12 column revealed several CytPTKs within the same tissue, their peaks ranging between 30 and 450 kDa. Immunoblotting analysis showed Fyn and Lyn were present in most tissue cytosols. Upon immunoprecipitation, however, with anti-Fyn or anti-Lyn, negligible amounts (< 2%) of the total cellular CytPTK were precipitated. Thus, these general markers of CytPTKs comprise only a minor proportion of the total intracellular PolyGlu4Tyr phosphorylating capacity. To see whether a specific marker for CytPTK could be detected, we also examined the requirement of CytPTKs for divalent ions, their preferred phosphate donor and their sensitivity to inhibition by known PTK inhibitors. We found that the order of reactivity with divalent cations was Co2+ > Mn2+ > Mg2+, while Zn2+ and Ca2+ did not support CytPTK activity. The best phosphate donor was ATP (ED50 = 5 microM), but other nucleoside 3-phosphates could substitute for ATP at high concentrations. With respect to these parameters, no basic difference exists between cytosolic and plasma-membrane PTKs. The PTK inhibitors, genestein and quercetin, inhibited both cytosolic and membranal PTKs at micromolar concentrations. In contrast, staurosporine was a potent inhibitor of CytPTKs (IC50 5-20 nM) and a poor inhibitor of membranal PTKs (IC50 10-40 microM). One of the conclusions we can draw from this study is that tissue cytosols contain PolyGlu4Tyr phosphorylating capacity in quantities greater than previously assumed and that the low level of phosphotyrosine found in cells is not the result of limited intracellular levels of CytPTKs.
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PMID:Non-receptor cytosolic protein tyrosine kinases from various rat tissues. 749 84

1. Ionic conductances controlled by type A and type B cholecystokinin (CCK) receptors were studied in neurons of the rat nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus (DMNV), using intracellular and whole-cell patch clamp recordings in current or voltage clamp configuration during bath application of agonists (CCK8, CCK4, BC 264) and antagonists. 2. CCKA receptor-related inhibition was associated with a membrane hyperpolarization and a decrease in input resistance that developed 2-6 min after the arrival of drug into the extracellular medium. These effects were induced by 5 nM CCK8 but not BC 264 and they were blocked by the CCKA antagonist, L-364,718, but not by the CCKB antagonist, L-365,260. 3. CCKA receptor-related inhibition was generated by a potassium current that reversed at a reversal potential E(rev) of -73 +/- 1 (mean +/- SE) mV with bathing potassium concentration [K+]o = 6 mM and at -88 +/- 1 with [K+]o = 3 mM, in agreement with the Nernst equation for potassium ions. 4. CCKB receptor-related excitation was associated with a membrane depolarization and an increase of the input resistance induced by the following agonists at threshold concentrations: CCK8 (0.2 nM) > or = BC 264 (0.4 nM) > CCK4 (10.9 nM). The increase of input resistance was abolished by L-365,260 and was maintained after blockade of the CCKA current by L-364,718. 5. CCKB receptor-related excitation, in the neurons (30% of cases) in which clear response reversal was observed, appeared to be generated by a decrease of a potassium conductance. Responses showed a reversal potential E(rev) of -68 +/- 4 mV with [K+]o = 6 mM and -89 +/- 1 mV with [K+]o = 3 mM, verifying predictions from the Nernst equation applied to potassium ions. However, in 70% of cases, clear reversal was not observed at membrane potentials negative to the theoretical potassium equilibrium potential EK. 6. In voltage clamp studies, CCK8 induced a 181 +/- 17 pA inward current associated with a 26 +/- 4% decrease in the instantaneous current (I(ins)) generated by hyperpolarizing voltage steps. This effect on I(ins) was demonstrated in the absence of effects on the outward noninactivating potassium current (IM) and on the inward noninactivating cationic current (IQ). 7. CCKB receptor-mediated excitation was not suppressed by cobalt, a blocker of calcium currents, and was not associated with a change of the calcium-dependent potassium current (IK(Ca)).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Cholecystokinin-gated currents in neurons of the rat solitary complex in vitro. 750 60

The interaction between the cobalt complex of a bleomycin functional model and d(CGCAATTGCG)2 was determined by 2D NMR methods. The intermolecular NOE cross peaks between ligand protons and the DNA minor groove protons suggest that the cobalt complex of AMPHIS-NET binds in the minor groove of DNA at the central AATT site. Also, the NOEs interactions of H8 pyridine proton and H2 imidazole proton in the metal-binding domain with H4' sugar proton of C9 and H4' sugar proton of A15 demonstrate that the sixth ligand site of cobalt complex, which is occupied by a solvent molecule, faces towards the minor groove of DNA. This binding model is in accord with the observed non-diffusion DNA cleavage by the metal-complexes of the bleomycin functional model compounds.
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PMID:Interaction of the cobalt complex of a bleomycin functional model with d(CGCAATTGCG)2: evidence of minor groove binding by 2D NMR methods. 754 92

Smooth muscle cells, macrophages, glial cells, keratinocytes, and transformed cells have been established as synthesis sites for vascular endothelial growth factor (VEGF). The modulating effects of VEGF are essentially limited to endothelial cells (ECs), the only cell type consistently shown to express VEGF receptors. VEGF has thus been considered to act exclusively via a paracrine pathway. We sought to determine whether the role of human ECs might, under selected conditions, extend beyond that of a target to involve contingency synthesis of VEGF. In both unstimulated human umbilical vein ECs (HUVECs) and human derma-derived microvascular ECs (HMECs), Northern analysis detected no VEGF transcripts. Phorbol-12-myristate 13-acetate (10(-7) M) treatment, however, induced VEGF mRNA expression in both HUVECs and HMECs, peaking at 3 and 6 h, respectively, and returning to undetectable levels by 12 h. In vitro exposure of HUVECs to a hypoxic environment (pO2 = 35 mm of mercury) for 12, 24, and 48 h and exposure of HMECs for 6, 12, 24, and 48 h induced VEGF mRNA in a time-dependent fashion. Re-exposure to normoxia (pO2 = 150 mm of mercury) for 24 h after 24 h of hypoxia returned VEGF mRNA transcripts to undetectable levels in HUVECs. Cobalt chloride and nickel chloride treatment each induced VEGF mRNA in ECs. Cycloheximide treatment further augmented expression of VEGF mRNA induced by cobalt chloride, nickel chloride, and hypoxia in HUVECs. VEGF protein production in hypoxia HUVECs was demonstrated immunohistochemically. Conditioned media from hypoxic HUVECs caused a 2-fold increase in the incorporation of tritiated thymidine. Finally, immune precipitates of anti-KDR probed with anti-Tyr(P) antibodies demonstrated evidence of receptor autophosphorylation in hypoxic but not normoxic HUVECs. These findings thus establish the potential for an autocrine pathway that may augment and/or amplify the paracrine effects of VEGF in stimulating angiogenesis.
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PMID:Hypoxia induces vascular endothelial growth factor in cultured human endothelial cells. 853 83

The interaction between the cobalt(III) complex of a bleomycin functional model (AMPHIS-NET) and the oligonucleotide d(CGCAATTGCG)2 and the structural features of the 1:1 ligand-DNA complex have been determined by high-resolution two-dimensional nuclear magnetic resonance methods and restrained molecular dynamics calculations. The intermolecular nuclear Overhauser effect (NOE) cross-peaks between ligand protons and the DNA minor groove protons suggest that the cobalt(III) complex of AMPHIS-NET binds in the minor groove of DNA at the central AATT site. The NOE connectivities also clearly indicate that the H8 pyridine proton and the H2 imidazole proton in the metal-binding domain interact with the H4' sugar proton of C19 and the H4' sugar proton of A5, respectively, which defines a structure where the metal binding moiety of Co(III).AMPHIS-NET participates in binding to the DNA and extends into the region two base pairs beyond the central AATT site in the minor groove. This binding model is in accord with the consistently observed nondiffusion DNA cleavage in locations two to three residues beyond the end of AT-rich binding sites induced by the corresponding iron(II) complexes of AMPHIS-NET and other AMPHIS-lexitropsin hybrids of the bleomycin functional model compounds.
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PMID:Solution structure studies of the cobalt complex of a bleomycin functional model bound to d(CGCAATTGCG)2 by two-dimensional nuclear magnetic resonance methods and restrained molecular dynamics simulation. 895 Apr 86

This study examined the expression of EPO, VEGF and VEGF receptor gene under conditions of reduced oxygen supply in primary cultures of rat hepatocytes, and compared it with the expression of these genes in hypoxic rat livers in vivo. To this end we exposed male Sprague-Dawley rats to hypoxia (10% and 8% O2), carbon monoxide (0.1% CO) or injected cobalt chloride (60 mg/kg CoCl2) subcutaneously. For the in vitro experiments we used primary cultures of rat hepatocytes which were kept at high (20% O2) and low (1% O2) oxygen tensions for three hours. The EPO mRNA was up-regulated by hypoxia in vitro and in vivo about 10-fold. The VEGF mRNA was up-regulated fivefold in the hepatocytes only, whereas the in vivo mRNA levels remained unchanged. The mRNA levels of flt-1 were up-regulated threefold by 8% O2 in livers, dependent on the strength of hypoxia (10% caused no changes in flt-1 gene expression) and on the kind of hypoxic stimulus (8% O2 was as effective as 0.1% CO and more effective than cobalt). The mRNA levels of flk-1/KDR and flt-4 remained unchanged in the liver. In vitro there were no changes in the mRNA levels of flt-1, flt-4 and flk-1/KDR. Consequently, the in vivo regulation of VEGF, which might be modulated by induction of flt-1 receptor gene expression, differs from the in vitro cell culture situation and might be different from the EPO regulation in vivo.
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PMID:Induction of VEGF and VEGF receptor gene expression by hypoxia: divergent regulation in vivo and in vitro. 902 20

Co-exposure to cadmium, cobalt, lead and other heavy metals occurs in many occupational settings, such as pigment and batteries production, galvanization and recycling of electric tools. However, little is known about interactions between several heavy metals. In the present study we determined DNA single strand break (DNA-SSB) induction and repair capacity for 8-oxoguanine in mononuclear blood cells of 78 individuals co-exposed to cadmium (range of concentrations in air: 0.05-138.00 micro g/m(3)), cobalt (range: 0-10 micro g/m(3)) and lead (range: 0-125 micro g/m(3)). Exposure to heavy metals was determined in air, blood and urine. Non-parametric correlation analysis showed a correlation between cadmium concentrations in air with DNA-SSB (P = 0.001, R = 0.371). Surprisingly, cobalt air concentrations correlated even better (P < 0.001, R = 0.401), whereas lead did not correlate with DNA-SSB. Logistic regression analysis including 11 possible parameters of influence resulted in a model showing that cobalt in air, cadmium in air, cadmium in blood and lead in blood influence the level of DNA-SSB. The positive result with cobalt was surprising, since exposure levels were much lower compared with the TRK-value of 100 micro g/m(3). To examine, whether the positive result with cobalt is stable, we applied several logistic regression models with two blocks, where all factors except cobalt were considered preferentially. All strategies resulted in the model described above. Logistic regression analysis considering also all possible interactions between the relevant parameters of influence finally resulted in the following model: Odds ratio = 1.286(Co in air) x 1.040(Cd in air) x 3.111(Cd in blood) x 0.861(Pb in air) x 1.023(Co in air x Pb in air). This model correctly predicts an increased level of DNA-SSB in 91% of the subjects in our study. One conclusion from this model is the existence of more than multiplicative effects for co-exposures of cadmium, cobalt and lead. For instance increasing lead air concentrations from 1.6 to 50 micro g/m(3) in the presence of constant exposures to cobalt and cadmium (8 micro g/m(3) and 3.8 micro g/m(3)) leads to an almost 5-fold increase in the odds ratio, although lead alone does not increase DNA-SSB. The mechanism behind these interactions might be repair inhibition of oxidative DNA damage, since a decrease in repair capacity will increase susceptibility to reactive oxygen species generated by cadmium or cobalt. Indeed, repair of 8-oxoguanine decreased with increasing exposures and inversely correlated with the level of DNA-SSB (P = 0.001, R = -0.427). Protein expression patterns of individuals exposed to cobalt concentrations of approximately 10 micro g/m(3) were compared with those of unexposed individuals using two-dimensional gel electrophoresis. Qualitative and apparent quantitative alterations in protein expression were selective and certainly occurred in <0.1% of all proteins. In conclusion, the hazard due to cobalt exposure - that has been classified only as IIB by the IARC - seems to be underestimated, especially when individuals are co-exposed to cadmium or lead. Co-exposure may cause genotoxic effects, even if the concentrations of individual heavy metals do not exceed TRK-values.
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PMID:Occupational exposure to heavy metals: DNA damage induction and DNA repair inhibition prove co-exposures to cadmium, cobalt and lead as more dangerous than hitherto expected. 1297 68

The molecular mechanism of hypoxia-induced apoptosis has not been clearly elucidated. In this study, we investigated the involvement of extracellular signal-regulated protein kinase (ERK 1/2) in hypoxia-induced apoptosis using cobalt chloride in HeLa human cervical cancer cells. The cobalt chloride was used for the induction of hypoxia, and its IC(50) was 471.4 microM. We demonstrated the DNA fragmentation after incubation with concentrations more than 50 microM cobalt chloride for 24 h, and also evidenced the morphological changes of the cells undergoing apoptosis with electron microscopy. Next, we examined the signaling pathway of cobalt chloride-induced apoptosis in HeLa cells. ERK1/2 activation occurred 6 and 9 h after treatment with 600 microM cobalt chloride. Meanwhile, the pretreatment of the MEK 1 inhibitor (PD98059) completely blocked the cobalt chloride-induced ERK 1/2 activation. At the same time, the activated ERK 1/2 translocated into the nucleus and phosphorylated its transcriptional factor, c-Jun. In addition, the pretreatment of PD98059 inhibited the cobalt chloride-induced DNA fragmentation and apoptotic cell death. These results suggest that cobalt chloride is able to induce apoptotic activity in HeLa cells, and its apoptotic mechanism may be associated with signal transduction via ERK 1/2.
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PMID:Cobalt chloride-induced apoptosis and extracellular signal-regulated protein kinase activation in human cervical cancer HeLa cells. 1453 30

Using tight-seal recordings from rat spinal cord slices, intracellular labelling and computer simulation, we analysed the mechanisms of spike frequency adaptation in substantia gelatinosa (SG) neurones. Adapting-firing neurones (AFNs) generated short bursts of spikes during sustained depolarization and were mostly found in lateral SG. The firing pattern and the shape of single spikes did not change after substitution of Ca2+ with Co2+, Mg2+ or Cd2+ indicating that Ca2+-dependent conductances do not contribute to adapting firing. Transient KA current was small and completely inactivated at resting potential suggesting that adapting firing was mainly generated by voltage-gated Na+ and delayed-rectifier K+ (KDR) currents. Although these currents were similar to those previously described in tonic-firing neurones (TFNs), we found that Na+ and KDR currents were smaller in AFNs. Discharge pattern in TFNs could be reversibly converted into that typical of AFNs in the presence of tetrodotoxin but not tetraethylammonium, suggesting that lower Na+ conductance is more critical for the appearance of firing adaptation. Intracellularly labelled AFNs showed specific morphological features and preserved long extensively branching axons, indicating that smaller Na+ conductance could not result from the axon cut. Computer simulation has further revealed that down-regulation of Na+ conductance represents an effective mechanism for the induction of firing adaptation. It is suggested that the cell-specific regulation of Na+ channel expression can be an important factor underlying the diversity of firing patterns in SG neurones.
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PMID:Mechanism of spike frequency adaptation in substantia gelatinosa neurones of rat. 1523 88


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