Relationship between solvent retention capacity and protein molecular weight distribution, quality characteristics, and breadmaking functionality of hard red spring wheat flour

USDA-ARS?s Scientific Manuscript database

In order to investigate suitability of solvent retention capacity (SRC) test for quality assessment of hard red spring (HRS) wheat flour, ten HRS genotypes from six locations in North Dakota State were analyzed for SRC and flour and breadmaking quality characteristics. The SRC values were significa...

Heterofunctionality interaction with donor solvent coal liquefaction. Final progress report, August 1982-April 1984

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cronauer, D.C.

1984-05-01

This project was undertaken to understand the role of the coal liquefaction solvent through a study of the interaction between the hydrogen donor solvent characteristics and the heterofunctionality of the solvent. Specifically, hydroxyl- and nitrogen-containing solvents were studied and characterized. A series of coal liquefaction experiments were carried out at 450/sup 0/C in a continuous feed stirred-tank reactor (CSTR) to observe the effect of adding phenolics to anthracene oil (AO) and SRC-II recycle solvents. The addition of phenol to AO at a ratio of 5/65 resulted in a nominal increase in coal conversion to THF solubles, but the amount ofmore » asphaltenes more than doubled resulting in a sizable net loss of solvent. The addition of m-cresol to both AO and SRC-II solvents had a positive effect on coal conversion to both THF and pentane solubles (oils). The partial removal of an OH-concentrate from SRC-II solvent was carried out using Amberlyst IRA-904 ion exchange resin. The resin-treated oil was only marginally better than raw SRC-II recycle solvent for coal liquefaction. Hydroaromatics having nitrogen functionality should be good solvents for coal liquefaction considering their effective solvent power, ability to penetrate and swell coal, and their ability to readily transfer hydrogen, particularly in the presence of oxygen functionality. However, these benefits are overshadowed by the strong tendency of the nitrogen-containing species to adduct with themselves and coal-derived materials.« less

Process for producing low-sulfur boiler fuel by hydrotreatment of solvent deashed SRC

DOEpatents

Roberts, George W.; Tao, John C.

1985-01-01

In this invention, a process is disclosed characterized by heating a slurry of coal in the presence of a process-derived recycle solvent and passing same to a dissolver zone, separating the resultant gases and liquid/solid products therefrom, vacuum distilling the liquid/solids products, separating the portions of the liquid/solids vacuum distillation effluent into a solid ash, unconverted coal particles and SRC material having a boiling point above 850.degree. F. and subjecting same to a critical solvent deashing step to provide an ash-free SRC product. The lighter liquid products from the vacuum distillation possess a boiling point below 850.degree. F. and are passed through a distillation tower, from which recycled solvent is recovered in addition to light distillate boiling below 400.degree. F. (overhead). The ash-free SRC product in accompanyment with at least a portion of the process derived solvent is passed in combination to a hydrotreating zone containing a hydrogenation catalyst and in the presence of hydrogen is hydroprocessed to produce a desulfurized and denitrogenized low-sulfur, low-ash boiler fuel and a process derived recycle solvent which is recycled to slurry the coal in the beginning of the process before heating.

Preliminary study of semi-refined carrageenan (SRC) as secondary gelling agent in natural rubber (NR) latex foam

NASA Astrophysics Data System (ADS)

Norhazariah, S.; Azura, A. R.; Azahari, B.; Sivakumar, R.

2017-12-01

Semi-refined carrageenan (SRC) product is considerably cheaper and easier to produce as a natural polysaccharide, which was utilized in food and other product application. However, the application in latex is limited. The aim of this work is to evaluate the SRC produced from low industrial grade seaweed (LIGS) in the latex foam application. The FTIR spectra showed the SRC produced as kappa type carrageenan with lower sulfur content compared to native LIGS. NR latex foam is produced by using the Dunlop method with some modifications. The effect of SRC loading as a secondary gelling agent in NR latex foam is investigated. The density and morphology of the NR latex foam with the addition of the SRC are analyzed. NR latex foam density increased with SRC loading and peaked at 1.8 phr SRC. The addition of SRC has induced the bigger cell size compared to the cell size of the control NR latex foam, as shown in the optical micrograph. It can be concluded that SRC LIGS could be acted as secondary gelling agent in NR latex foam.

Effect of addition of semi refined carrageenan on mechanical characteristics of gum arabic edible film

NASA Astrophysics Data System (ADS)

Setyorini, D.; Nurcahyani, P. R.

2016-04-01

Currently the seaweed is processed flour and Semi Refined Carraagenan (SRC). However, total production is small, but both of these products have a high value and are used in a wide variety of products such as cosmetics, processed foods, medicines, and edible film. The aim of this study were (1) to determine the effect of SRC on mechanical characteristics of edible film, (2) to determine the best edible film which added by SRC with different concentration. The edible film added by SRC flour which divided into three concentrations of SRC. There are 1.5%; 3%; and 4.5% of SRC, then added 3% glycerol and 0.6% arabic gum. The mechanical properties of the film measured by a universal testing machine Orientec Co. Ltd., while the water vapor permeability measured by the gravimetric method dessicant modified. The experimental design used was completely randomized design with a further test of Duncan. The result show SRC concentration differences affect the elongation breaking point and tensile strength. But not significant effect on the thickness, yield strength and the modulus of elasticity. The best edible film is edible film with the addition of SRC 4.5%.

Plan for radionuclide tracer studies of the residence time distribution in the Wilsonville dissolver and preheater

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jolley, R.L.; Begovich, J.M.; Brashear, H.R.

1983-12-01

Stimulus-response measurements using radiotracers to measure residence time distribution (RTD) and hydrodynamic parameters for the preheaters and dissolvers at the Ft. Lewis Solvent Refined Coal (SRC) and the Exxon Donor Solvent (EDS) coal conversion pilot plants are reviewed. A plan is also presented for a series of radioactive tracer studies proposed for the Advanced Coal Liquefaction Facility at Wilsonville, Alabama, to measure the RTD for the preheater and dissolvers in the SRC-I mode. The tracer for the gas phase will be /sup 133/Xe, and /sup 198/Au (on carbonized resin or as an aqueous colloidal suspension) will be used as themore » slurry tracer. Four experimental phases are recommended for the RTD tracer studies: (1) preheater; (2) dissolver with 100% takeoff; (3) dissolver with 100% takeoff and solids withdrawal; and (4) dissolver with 50% takeoff. Eighteen gas-tracer and 22 liquid-tracer injections are projected to accomplish the four experimental phases. Two to four tracer injections are projected for preliminary tests to ensure the capability of safe injection of the radiotracers and the collection of statistically significant data. A complete projected cost and time schedule is provided, including procurement of necessary components, preparation of the radiotracers, assembly and testing of tracer injection apparatus and detection systems, onsite work and tracer injections, laboratory experimentation, data analysis, and report writing.« less

Relationships of flour solvent retention capacity, secondary structure and rheological properties with the cookie making characteristics of wheat cultivars.

PubMed

Kaur, Amritpal; Singh, Narpinder; Kaur, Seeratpreet; Ahlawat, Arvind Kumar; Singh, Anju Mahendru

2014-09-01

The relationships of grain, flour solvent retention capacity (SRC) and dough rheological properties with the cookie making properties of wheat cultivars were evaluated. Cultivars with higher proportion of intermolecular-β-sheets+antiparallel β sheets and lower α-helix had greater gluten strength. The grain weight and diameter positively correlated with the proportion of fine particles and the cookie spread factor (SF) and negatively to the grain hardness (GH) and Na2CO3 SRC. The SF was higher in the flour with a higher amount of fine particle and with a lower Na2CO3 SRC and dough stability (DS). The breaking strength (BS) of cookies was positively correlated to lactic acid (LA) SRC, DS, peak time, sedimentation value (SV), G' and G″. Na2CO3 SRC and GH were strongly correlated. The gluten performance index showed a strong positive correlation with SV, DS, G' and G″. The water absorption had a significant positive correlation with sucrose SRC and LASRC. Cultivars with higher GH produced higher amount of coarse particles in flours that had higher Na2CO3 SRC and lower cookie SF. Copyright © 2014 Elsevier Ltd. All rights reserved.

Coal Technology Program progress report for April 1976

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

In the Hydrocarbonization Research program, two successful experiments were completed in the bench-scale hydrocarbonizer. A settling test at a lower temperature (390/sup 0/F) using 20 percent toluene in Solvent Refined Coal (SRC) Unfiltered Oil (UFO) produced a 30 percent clarified product in 2 hr. Characterization tests include distillation curves for Wilsonville's SRC-UFO and a particle size distribution of Pittsburg and Midway Coal Mining Company's (PAMCO) SRC-UFO. Studies of intermediate-temperature pyrolysis of large blocks have been maintained with char samples continuing to demonstrate pyrophoricity, even after heating to 700/sup 0/C. Simulated distillation analysis of tars produced by the last eight experimentsmore » are being compared with those performed at Laramie upon tars produced by the Hanna No. 2 experiment. In Coal-Fueled MIUS, stainless steel tubing to be used in one of the furnace tube bundles was ordered and the bid package for the furnace completed. Tests continued on the coal feed system and with the cold flow fluidized bed model. For the Synthoil process, flow diagrams, material balances, and utilities requirements were completed for the entire facility. For the Hydrocarbonization process, flowsheets were reviewed for compatibility; equipment lists were brought up to date; and utilities requirements were compiled from the individual flowsheets. The char recovery and storage subsystem flowsheet was completed. (auth)« less

Process for solvent refining of coal using a denitrogenated and dephenolated solvent

DOEpatents

Garg, Diwakar; Givens, Edwin N.; Schweighardt, Frank K.

1984-01-01

A process is disclosed for the solvent refining of non-anthracitic coal at elevated temperatures and pressure in a hydrogen atmosphere using a hydrocarbon solvent which before being recycled in the solvent refining process is subjected to chemical treatment to extract substantially all nitrogenous and phenolic constituents from the solvent so as to improve the conversion of coal and the production of oil in the solvent refining process. The solvent refining process can be either thermal or catalytic. The extraction of nitrogenous compounds can be performed by acid contact such as hydrogen chloride or fluoride treatment, while phenolic extraction can be performed by caustic contact or contact with a mixture of silica and alumina.

Firing of pulverized solvent refined coal

DOEpatents

Derbidge, T. Craig; Mulholland, James A.; Foster, Edward P.

1986-01-01

An air-purged burner for the firing of pulverized solvent refined coal is constructed and operated such that the solvent refined coal can be fired without the coking thereof on the burner components. The air-purged burner is designed for the firing of pulverized solvent refined coal in a tangentially fired boiler.

Effect of wheat flour characteristics on sponge cake quality.

PubMed

Moiraghi, Malena; de la Hera, Esther; Pérez, Gabriela T; Gómez, Manuel

2013-02-01

To select the flour parameters that relate strongly to cake-making performance, in this study the relationship between sponge cake quality, solvent retention capacity (SRC) profile and flour physicochemical characteristics was investigated using 38 soft wheat samples of different origins. Particle size average, protein, damaged starch, water-soluble pentosans, total pentosans, SRC and pasting properties were analysed. Sponge cake volume and crumb texture were measured to evaluate cake quality. Cluster analysis was applied to assess differences in flour quality parameters among wheat lines based on the SRC profile. Cluster 1 showed significantly higher sponge cake volume and crumb softness, finer particle size and lower SRC sucrose, SRC carbonate, SRC water, damaged starch and protein content. Particle size, damaged starch, protein, thickening capacity and SRC parameters correlated negatively with sponge cake volume, while total pentosans and pasting temperature showed the opposite effect. The negative correlations between cake volume and SRC parameters along with the cluster analysis results indicated that flours with smaller particle size, lower absorption capacity and higher pasting temperature had better cake-making performance. Some simple analyses, such as SRC, particle size distribution and pasting properties, may help to choose flours suitable for cake making. Copyright © 2012 Society of Chemical Industry.

Soft wheat and flour products methods review: solvent retention capacity equation correction

USDA-ARS?s Scientific Manuscript database

This article discusses the results of a significant change to calculations made within AACCI Approved methods 56-10 and 56-11, the Alkaline Water Retention Capacity (AWRC) test and the Solvent Retention Capacity (SRC) test. The AACCI Soft Wheat and Flour Products Technical Committee reviewed propos...

Studies of the effect of selected nondonor solvents on coal liquefaction yields

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jolley, R. L.; Rodgers, B. R.; Benjamin, B. M.

The objective of this research program was to evaluate the effectiveness of selected nondonor solvents (i.e., solvents that are not generally considered to have hydrogen available for hydrogenolysis reactions) for the solubilization of coals. Principal criteria for selection of candidate solvents were that the compound should be representative of a major chemical class, should be present in reasonable concentration in coal liquid products, and should have the potential to participate in hydrogen redistribution reactions. Naphthalene, phenanthrene, pyrene, carbazole, phenanthridine, quinoline, 1-naphthol, and diphenyl ether were evaluated to determine their effect on coal liquefaction yields and were compared with phenol andmore » two high-quality process solvents, Wilsonville SRC-I recycle solvent and Lummus ITSL heavy oil solvent. The high conversion efficacy of 1-naphthol may be attributed to its condensation to binaphthol and the consequent availability of hydrogen. The effectiveness of both the nitrogen heterocycles and the polycyclic aromatic hydrocarbon (PAH) compounds may be due to their polycyclic aromatic nature (i.e., possible hydrogen shuttling or transfer agents) and their physical solvent properties. The relative effectiveness for coal conversion of the Lummus ITSL heavy oil solvent as compared with the Wilsonville SRC-I process solvent may be attributed to the much higher concentration of 3-, 4-, and 5-ring PAH and hydroaromatic constituents in Lummus solvent. The chemistry of coal liquefaction and the development of recycle, hydrogen donor, and nondonor solvents are reviewed. The experimental methodology for tubing-bomb tests is outlined, and experimental problem areas are discussed.« less

Oxidative Gelation of Solvent-Accessible Arabinoxylans is the Predominant Consequence of Extensive Chlorination of Soft Wheat Flour

USDA-ARS?s Scientific Manuscript database

Solvent retention capacity (SRC) and Bostwick flow were used to explore the effects of milling yield, extent of chlorination, and flour particle size on cake flour functionality and batter viscosity. The effects of the extent of chlorination were dramatic, but milling yield and additional milling t...

The solvent component of macromolecular crystals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weichenberger, Christian X.; Afonine, Pavel V.; Kantardjieff, Katherine

2015-04-30

On average, the mother liquor or solvent and its constituents occupy about 50% of a macromolecular crystal. Ordered as well as disordered solvent components need to be accurately accounted for in modelling and refinement, often with considerable complexity. The mother liquor from which a biomolecular crystal is grown will contain water, buffer molecules, native ligands and cofactors, crystallization precipitants and additives, various metal ions, and often small-molecule ligands or inhibitors. On average, about half the volume of a biomolecular crystal consists of this mother liquor, whose components form the disordered bulk solvent. Its scattering contributions can be exploited in initialmore » phasing and must be included in crystal structure refinement as a bulk-solvent model. Concomitantly, distinct electron density originating from ordered solvent components must be correctly identified and represented as part of the atomic crystal structure model. Herein, are reviewed (i) probabilistic bulk-solvent content estimates, (ii) the use of bulk-solvent density modification in phase improvement, (iii) bulk-solvent models and refinement of bulk-solvent contributions and (iv) modelling and validation of ordered solvent constituents. A brief summary is provided of current tools for bulk-solvent analysis and refinement, as well as of modelling, refinement and analysis of ordered solvent components, including small-molecule ligands.« less

Integrated process for the solvent refining of coal

DOEpatents

Garg, Diwakar

1983-01-01

A process is set forth for the integrated liquefaction of coal by the catalytic solvent refining of a feed coal in a first stage to liquid and solid products and the catalytic hydrogenation of the solid product in a second stage to produce additional liquid product. A fresh inexpensive, throw-away catalyst is utilized in the second stage hydrogenation of the solid product and this catalyst is recovered and recycled for catalyst duty in the solvent refining stage without any activation steps performed on the used catalyst prior to its use in the solvent refining of feed coal.

Comparison of binding energies of SrcSH2-phosphotyrosyl peptides with structure-based prediction using surface area based empirical parameterization.

PubMed Central

Henriques, D. A.; Ladbury, J. E.; Jackson, R. M.

2000-01-01

The prediction of binding energies from the three-dimensional (3D) structure of a protein-ligand complex is an important goal of biophysics and structural biology. Here, we critically assess the use of empirical, solvent-accessible surface area-based calculations for the prediction of the binding of Src-SH2 domain with a series of tyrosyl phosphopeptides based on the high-affinity ligand from the hamster middle T antigen (hmT), where the residue in the pY+ 3 position has been changed. Two other peptides based on the C-terminal regulatory site of the Src protein and the platelet-derived growth factor receptor (PDGFR) are also investigated. Here, we take into account the effects of proton linkage on binding, and test five different surface area-based models that include different treatments for the contributions to conformational change and protein solvation. These differences relate to the treatment of conformational flexibility in the peptide ligand and the inclusion of proximal ordered solvent molecules in the surface area calculations. This allowed the calculation of a range of thermodynamic state functions (deltaCp, deltaS, deltaH, and deltaG) directly from structure. Comparison with the experimentally derived data shows little agreement for the interaction of SrcSH2 domain and the range of tyrosyl phosphopeptides. Furthermore, the adoption of the different models to treat conformational change and solvation has a dramatic effect on the calculated thermodynamic functions, making the predicted binding energies highly model dependent. While empirical, solvent-accessible surface area based calculations are becoming widely adopted to interpret thermodynamic data, this study highlights potential problems with application and interpretation of this type of approach. There is undoubtedly some agreement between predicted and experimentally determined thermodynamic parameters: however, the tolerance of this approach is not sufficient to make it ubiquitously applicable. PMID:11106171

Firing of pulverized solvent refined coal

DOEpatents

Lennon, Dennis R.; Snedden, Richard B.; Foster, Edward P.; Bellas, George T.

1990-05-15

A burner for the firing of pulverized solvent refined coal is constructed and operated such that the solvent refined coal can be fired successfully without any performance limitations and without the coking of the solvent refined coal on the burner components. The burner is provided with a tangential inlet of primary air and pulverized fuel, a vaned diffusion swirler for the mixture of primary air and fuel, a center water-cooled conical diffuser shielding the incoming fuel from the heat radiation from the flame and deflecting the primary air and fuel steam into the secondary air, and a watercooled annulus located between the primary air and secondary air flows.

Coal technology program progress report, February 1976

DOE Office of Scientific and Technical Information (OSTI.GOV)

None

Final testing of the 20-atm bench-scale system is underway in preparation for experiments with hydrogen. Laboratory-scale testing of a number of inexpensive pure compounds to improve the settling rate of solids in Solvent Refined Coal (SRC) unfiltered oil (UFO), bench-scale testing of the effect of the Tretolite additive on settling, and characterization tests on a new sample of UFO from the PAMCO-SRC process are reported. Experimental engineering support of an in situ gasification process include low-temperature pyrolyses at exceptionally low heating rates (0.3/sup 0/C/min). Highly pyrophoric chars were consistently produced. Aqueous by-products from coal conversion technologies and oil shale retortingmore » have been analyzed directly to determine major organic components. A report is being prepared discussing various aspects of the engineering evaluations of nuclear process heat for coal. A bench-scale test program on thermochemical water splitting for hydrogen production is under consideration. In the coal-fueled MIUS program, preparations for procurement of tubing for the matrix in the fluidized-bed furnace and for fabrication of the furnace continued. Analyses of the AiResearch gas turbine and recuperator under coal-fueled MIUS operating conditions are near completion. Process flow diagrams and heat and material balances were completed for most of the units in the synthoil process. Overall utilities requirements were calculated and the coal preparation flowsheets were finalized. For hydrocarbonization, the flowsheet was revised to include additional coal data. Flowsheets were finalized for the acid gas separation and recycle, and the oil-solids separation. (LTN)« less

The Activation of c-Src Tyrosine Kinase: Conformational Transition Pathway and Free Energy Landscape.

PubMed

Fajer, Mikolai; Meng, Yilin; Roux, Benoît

2017-04-20

Tyrosine kinases are important cellular signaling allosteric enzymes that regulate cell growth, proliferation, metabolism, differentiation, and migration. Their activity must be tightly controlled, and malfunction can lead to a variety of diseases, particularly cancer. The nonreceptor tyrosine kinase c-Src, a prototypical model system and a representative member of the Src-family, functions as complex multidomain allosteric molecular switches comprising SH2 and SH3 domains modulating the activity of the catalytic domain. The broad picture of self-inhibition of c-Src via the SH2 and SH3 regulatory domains is well characterized from a structural point of view, but a detailed molecular mechanism understanding is nonetheless still lacking. Here, we use advanced computational methods based on all-atom molecular dynamics simulations with explicit solvent to advance our understanding of kinase activation. To elucidate the mechanism of regulation and self-inhibition, we have computed the pathway and the free energy landscapes for the "inactive-to-active" conformational transition of c-Src for different configurations of the SH2 and SH3 domains. Using the isolated c-Src catalytic domain as a baseline for comparison, it is observed that the SH2 and SH3 domains, depending upon their bound orientation, promote either the inactive or active state of the catalytic domain. The regulatory structural information from the SH2-SH3 tandem is allosterically transmitted via the N-terminal linker of the catalytic domain. Analysis of the conformational transition pathways also illustrates the importance of the conserved tryptophan 260 in activating c-Src, and reveals a series of concerted events during the activation process.

Relationship between physicochemical characteristics of flour and sugar-snap cookie quality in Korean wheat cultivar

USDA-ARS?s Scientific Manuscript database

The relationship of physicochemical properties of flour, including particle size of flour, damaged starch, SDS-sedimentation volume, gluten content and four solvent retention capacity (SRC) values with cookie baking quality, including cookie diameter and thickness was evaluated using 30 Korean wheat...

Effects of inhalation exposure to SRC-II heavy and middle distillates

DOE Office of Scientific and Technical Information (OSTI.GOV)

Springer, D.L.; Miller, R.A.; Weimer, W.C.

1984-11-01

To expand the data base on potential health effects of coal liquefaction materials, we have performed studies with both solvent refined coal (SRC)-II heavy distillate (HD) and middle distillate (MD). Weight gain for exposed animals was less than that of controls and was dose-related, ranging from no significant difference for animals in the low-exposure group to failure to gain in the high-dose animals. Liver weights increased significantly over controls, and thymus weights decreased for animals sacrificed at 5 and 13 weeks. After both exposure periods, there were significant treatment-related decreases in erythrocyte parameters and in certain types of white bloodmore » cells (WBC). Bone marrow cellularity, and numbers of megakaryocytes consistently decreased, suggesting that bone marrow is a target tissue for high-boiling coal liquids. Microscopic evaluation of tissue indicated exposure-related changes is listed. In contrast to the reported mutagenic and carcinogenic effects observed for the high-boiling coal liquids, middle-boiling-range materials lacked such activity in these assays. These data demonstrate a great deal of similarity in the kinds of effects observed following exposure to middle- and high-boiling-range coal liquids. However, the significance of changes in organ weights and peripheral blood parameters are not always readily apparent following a subchronic study. Because of this, we exposed animals to HD in a manner similar to that for the subchronic experiment and have followed these animals throughout their lives for the development of adverse effects such as reduced longevity and the appearance of tumors. Results from this study will be available for mice in FY 1985 and for rats in FY 1986.« less

Refinement of NMR structures using implicit solvent and advanced sampling techniques.

PubMed

Chen, Jianhan; Im, Wonpil; Brooks, Charles L

2004-12-15

NMR biomolecular structure calculations exploit simulated annealing methods for conformational sampling and require a relatively high level of redundancy in the experimental restraints to determine quality three-dimensional structures. Recent advances in generalized Born (GB) implicit solvent models should make it possible to combine information from both experimental measurements and accurate empirical force fields to improve the quality of NMR-derived structures. In this paper, we study the influence of implicit solvent on the refinement of protein NMR structures and identify an optimal protocol of utilizing these improved force fields. To do so, we carry out structure refinement experiments for model proteins with published NMR structures using full NMR restraints and subsets of them. We also investigate the application of advanced sampling techniques to NMR structure refinement. Similar to the observations of Xia et al. (J.Biomol. NMR 2002, 22, 317-331), we find that the impact of implicit solvent is rather small when there is a sufficient number of experimental restraints (such as in the final stage of NMR structure determination), whether implicit solvent is used throughout the calculation or only in the final refinement step. The application of advanced sampling techniques also seems to have minimal impact in this case. However, when the experimental data are limited, we demonstrate that refinement with implicit solvent can substantially improve the quality of the structures. In particular, when combined with an advanced sampling technique, the replica exchange (REX) method, near-native structures can be rapidly moved toward the native basin. The REX method provides both enhanced sampling and automatic selection of the most native-like (lowest energy) structures. An optimal protocol based on our studies first generates an ensemble of initial structures that maximally satisfy the available experimental data with conventional NMR software using a simplified force field and then refines these structures with implicit solvent using the REX method. We systematically examine the reliability and efficacy of this protocol using four proteins of various sizes ranging from the 56-residue B1 domain of Streptococcal protein G to the 370-residue Maltose-binding protein. Significant improvement in the structures was observed in all cases when refinement was based on low-redundancy restraint data. The proposed protocol is anticipated to be particularly useful in early stages of NMR structure determination where a reliable estimate of the native fold from limited data can significantly expedite the overall process. This refinement procedure is also expected to be useful when redundant experimental data are not readily available, such as for large multidomain biomolecules and in solid-state NMR structure determination.

EXPOSURE TO ENVIRONMENTALLY RELEVANT CONCENTRATIONS OF DIFFERENT NONYLPHENOL FORMULATIONS IN JAPANESE MEDAKA. (R827098)

EPA Science Inventory

The time course of exposure to p-nonylphenol (NP) from two different sources was compared to equalivent exposures of 17--estradiol (E2) and a solvent control (ethanol; EtOH). Japanese medaka were exposed for 4 days to a nomina...

75 FR 71146 - Seamless Refined Copper Pipe and Tube From China and Mexico

Federal Register 2010, 2011, 2012, 2013, 2014

2010-11-22

... Copper Pipe and Tube From China and Mexico Determinations On the basis of the record \\1\\ developed in the... copper pipe and tube (``SRC pipe and tube'') from China and Mexico provided for in subheadings 7411.10.10... Copper Products, LLC, Pine Hall, NC; Mueller Copper Tube Products, Inc. and Mueller Copper Tube Company...

Chemistry and structure of coal-derived asphaltenes, Phase III. Quarterly progress report, January--March 1978

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yen, T. F.

1978-01-01

The solubility limits of Synthoil and PAMCO asphaltenes have been measured as a function of Hildebrand solubility parameters and hydrogen bonding. Solvents with moderate hydrogen bonding capacity such as dioxane, ethyl benzoate and dibutyl phthalate were found to be most effective in dissolving asphaltenes over the widest range of solubility parameters. VPO molecular weight studies of coal liquid derived carbenes, as a function of concentration in the solvent THF, indicate that these fractions are more strongly self-associated than the corresponding asphaltenes, and generally afford high infinite dilution number average molecular weights: Synthoil, 861; HRI H-Coal, 1156; Cat. Inc. SRC, 1228;more » PAMCO SRC, 1054. The variable ESR temperature dependence of the spin intensity for a Synthoil asphaltene-I/sub 2/ charge transfer followed a 1/T (Curie--Weiss) dependence over the temperature range from 25/sup 0/ to -114/sup 0/C suggesting that independent, non-interacting donor and acceptor doublets were formed. Weight percent OH values, determined from 'H NMR analysis of silylated asphaltenes, were found to provide a reasonably linear correlation with the absorbance of the monomeric OH infrared stretching bands of the asphaltenes.« less

Structure refinement of membrane proteins via molecular dynamics simulations.

PubMed

Dutagaci, Bercem; Heo, Lim; Feig, Michael

2018-07-01

A refinement protocol based on physics-based techniques established for water soluble proteins is tested for membrane protein structures. Initial structures were generated by homology modeling and sampled via molecular dynamics simulations in explicit lipid bilayer and aqueous solvent systems. Snapshots from the simulations were selected based on scoring with either knowledge-based or implicit membrane-based scoring functions and averaged to obtain refined models. The protocol resulted in consistent and significant refinement of the membrane protein structures similar to the performance of refinement methods for soluble proteins. Refinement success was similar between sampling in the presence of lipid bilayers and aqueous solvent but the presence of lipid bilayers may benefit the improvement of lipid-facing residues. Scoring with knowledge-based functions (DFIRE and RWplus) was found to be as good as scoring using implicit membrane-based scoring functions suggesting that differences in internal packing is more important than orientations relative to the membrane during the refinement of membrane protein homology models. © 2018 Wiley Periodicals, Inc.

Mobil lube dewaxing technologies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baker, C.L.; McGuiness, M.P.

1995-09-01

Currently, the lube refining industry is in a period of transition, with both hydroprocessing and catalytic dewaxing gathering momentum as replacements for solvent extraction and solvent dewaxing. In addition, lube product quality requirements have been increasing, both in the US and abroad. Mobil has developed a broad array of dewaxing catalytic technologies which can serve refiners throughout the stages of this transition. In the future, lube feedstocks which vary in source and wax content will become increasingly important, requiring an optimized system for highest performance. The Mobil Lube Dewaxing (MLDW) process is the work-horse of the catalytic dewaxing technologies, beingmore » a robust, low cost technology suitable for both solvent extracted and hydrocracked feeds. The Mobil Selective Dewaxing (MSDW) process has been recently introduced in response to the growth of hydroprocessing. MSDW requires either severely hydrotreated or hydrocracked feeds and provides improved lube yields and VI. For refiners with hydrocrackers and solvent dewaxing units, Mobil Wax Isomerization (MWI) technology can make higher VI base stocks to meet the growing demand for very high quality lube products. A review of these three technologies is presented in this paper.« less

Prostate segmentation by sparse representation based classification

PubMed Central

Gao, Yaozong; Liao, Shu; Shen, Dinggang

2012-01-01

Purpose: The segmentation of prostate in CT images is of essential importance to external beam radiotherapy, which is one of the major treatments for prostate cancer nowadays. During the radiotherapy, the prostate is radiated by high-energy x rays from different directions. In order to maximize the dose to the cancer and minimize the dose to the surrounding healthy tissues (e.g., bladder and rectum), the prostate in the new treatment image needs to be accurately localized. Therefore, the effectiveness and efficiency of external beam radiotherapy highly depend on the accurate localization of the prostate. However, due to the low contrast of the prostate with its surrounding tissues (e.g., bladder), the unpredicted prostate motion, and the large appearance variations across different treatment days, it is challenging to segment the prostate in CT images. In this paper, the authors present a novel classification based segmentation method to address these problems. Methods: To segment the prostate, the proposed method first uses sparse representation based classification (SRC) to enhance the prostate in CT images by pixel-wise classification, in order to overcome the limitation of poor contrast of the prostate images. Then, based on the classification results, previous segmented prostates of the same patient are used as patient-specific atlases to align onto the current treatment image and the majority voting strategy is finally adopted to segment the prostate. In order to address the limitations of the traditional SRC in pixel-wise classification, especially for the purpose of segmentation, the authors extend SRC from the following four aspects: (1) A discriminant subdictionary learning method is proposed to learn a discriminant and compact representation of training samples for each class so that the discriminant power of SRC can be increased and also SRC can be applied to the large-scale pixel-wise classification. (2) The L1 regularized sparse coding is replaced by the elastic net in order to obtain a smooth and clear prostate boundary in the classification result. (3) Residue-based linear regression is incorporated to improve the classification performance and to extend SRC from hard classification to soft classification. (4) Iterative SRC is proposed by using context information to iteratively refine the classification results. Results: The proposed method has been comprehensively evaluated on a dataset consisting of 330 CT images from 24 patients. The effectiveness of the extended SRC has been validated by comparing it with the traditional SRC based on the proposed four extensions. The experimental results show that our extended SRC can obtain not only more accurate classification results but also smoother and clearer prostate boundary than the traditional SRC. Besides, the comparison with other five state-of-the-art prostate segmentation methods indicates that our method can achieve better performance than other methods under comparison. Conclusions: The authors have proposed a novel prostate segmentation method based on the sparse representation based classification, which can achieve considerably accurate segmentation results in CT prostate segmentation. PMID:23039673

Prostate segmentation by sparse representation based classification.

PubMed

Gao, Yaozong; Liao, Shu; Shen, Dinggang

2012-10-01

The segmentation of prostate in CT images is of essential importance to external beam radiotherapy, which is one of the major treatments for prostate cancer nowadays. During the radiotherapy, the prostate is radiated by high-energy x rays from different directions. In order to maximize the dose to the cancer and minimize the dose to the surrounding healthy tissues (e.g., bladder and rectum), the prostate in the new treatment image needs to be accurately localized. Therefore, the effectiveness and efficiency of external beam radiotherapy highly depend on the accurate localization of the prostate. However, due to the low contrast of the prostate with its surrounding tissues (e.g., bladder), the unpredicted prostate motion, and the large appearance variations across different treatment days, it is challenging to segment the prostate in CT images. In this paper, the authors present a novel classification based segmentation method to address these problems. To segment the prostate, the proposed method first uses sparse representation based classification (SRC) to enhance the prostate in CT images by pixel-wise classification, in order to overcome the limitation of poor contrast of the prostate images. Then, based on the classification results, previous segmented prostates of the same patient are used as patient-specific atlases to align onto the current treatment image and the majority voting strategy is finally adopted to segment the prostate. In order to address the limitations of the traditional SRC in pixel-wise classification, especially for the purpose of segmentation, the authors extend SRC from the following four aspects: (1) A discriminant subdictionary learning method is proposed to learn a discriminant and compact representation of training samples for each class so that the discriminant power of SRC can be increased and also SRC can be applied to the large-scale pixel-wise classification. (2) The L1 regularized sparse coding is replaced by the elastic net in order to obtain a smooth and clear prostate boundary in the classification result. (3) Residue-based linear regression is incorporated to improve the classification performance and to extend SRC from hard classification to soft classification. (4) Iterative SRC is proposed by using context information to iteratively refine the classification results. The proposed method has been comprehensively evaluated on a dataset consisting of 330 CT images from 24 patients. The effectiveness of the extended SRC has been validated by comparing it with the traditional SRC based on the proposed four extensions. The experimental results show that our extended SRC can obtain not only more accurate classification results but also smoother and clearer prostate boundary than the traditional SRC. Besides, the comparison with other five state-of-the-art prostate segmentation methods indicates that our method can achieve better performance than other methods under comparison. The authors have proposed a novel prostate segmentation method based on the sparse representation based classification, which can achieve considerably accurate segmentation results in CT prostate segmentation.

Three-dimensional quantitative structure-activity relationship studies on novel series of benzotriazine based compounds acting as Src inhibitors using CoMFA and CoMSIA.

PubMed

Gueto, Carlos; Ruiz, José L; Torres, Juan E; Méndez, Jefferson; Vivas-Reyes, Ricardo

2008-03-01

Comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were performed on a series of benzotriazine derivatives, as Src inhibitors. Ligand molecular superimposition on the template structure was performed by database alignment method. The statistically significant model was established of 72 molecules, which were validated by a test set of six compounds. The CoMFA model yielded a q(2)=0.526, non cross-validated R(2) of 0.781, F value of 88.132, bootstrapped R(2) of 0.831, standard error of prediction=0.587, and standard error of estimate=0.351 while the CoMSIA model yielded the best predictive model with a q(2)=0.647, non cross-validated R(2) of 0.895, F value of 115.906, bootstrapped R(2) of 0.953, standard error of prediction=0.519, and standard error of estimate=0.178. The contour maps obtained from 3D-QSAR studies were appraised for activity trends for the molecules analyzed. Results indicate that small steric volumes in the hydrophobic region, electron-withdrawing groups next to the aryl linker region, and atoms close to the solvent accessible region increase the Src inhibitory activity of the compounds. In fact, adding substituents at positions 5, 6, and 8 of the benzotriazine nucleus were generated new compounds having a higher predicted activity. The data generated from the present study will further help to design novel, potent, and selective Src inhibitors as anticancer therapeutic agents.

Analysis of solvent dyes in refined petroleum products by electrospray ionization mass spectrometry

USGS Publications Warehouse

Rostad, C.E.

2010-01-01

Solvent dyes are used to color refined petroleum products to enable differentiation between gasoline, diesel, and jet fuels. Analysis for these dyes in the hydrocarbon product is difficult due to their very low concentrations in such a complex matrix. Flow injection analysis/electrospray ionization/mass spectrometry in both negative and positive mode was used to optimize ionization of ten typical solvent dyes. Samples of hydrocarbon product were analyzed under similar conditions. Positive electrospray ionization produced very complex spectra, which were not suitably specific for targeting only the dyes. Negative electrospray ionization produced simple spectra because aliphatic and aromatic moieties were not ionized. This enabled screening for a target dye in samples of hydrocarbon product from a spill.

Coal Technology Program progress report, March 1976

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

Shakedown tests of the bench-scale hydrocarbonization system were successfully completed. Wyodak coal was fed to the reactor at a rate of 9.9 lb/hr where it was hydrocarbonized at 1050/sup 0/F under 20-atm hydrogen pressure. Laboratory results including settling tests, bench-scale settling tests, and sample ageing tests were continued. Two of ten compounds tested with the laboratory-scale apparatus were effective in increasing settling rates of solids in Solvent Refined Coal unfiltered oil, but bench-scale tests failed to show any improvements in the settling rate over the untreated SRC-UFO. Analytical chemistry efforts involved the removal and concentration of organic components in by-productmore » waters from fossil fuel conversion processes. A sephadex gel is being used to achieve hydrophilic-lipophilic separations in organic mixtures as a step in the analysis of fossil fuel related materials. Engineering Evaluations of the Synthiol and Hydrocarbonization Processes continued with the Synthiol process flow diagrams, heat and material balances, and utilities requirements being completed. Inspection techniques were developed for wear- and process-resistant coatings. Orders were placed for the Incoloy 800 tubing and a smaller quantity of Inconel 600 tubing for the tube matrix in the coal-fueled MIUS fluidized bed. An engineering feasibility review of General Atomic's proposal to ERDA for a bench-scale test program on thermochemical water splitting for hydrogen production was completed. (auth)« less

The National Institute of Neurological Disorders and Stroke and Department of Defense Sport-Related Concussion Common Data Elements Version 1.0 Recommendations.

PubMed

Broglio, Steven P; Kontos, Anthony P; Levin, Harvey; Schneider, Kathryn; Wilde, Elisabeth A; Cantu, Robert C; Feddermann-Demont, Nina; Fuller, Gordon; Gagnon, Isabelle; Gioia, Gerry; Giza, Christopher C; Griesbach, Grace Sophia; Leddy, John J; Lipton, Michael L; Mayer, Andrew; McAllister, Thomas; McCrea, Michael; McKenzie, Lara; Putukian, Margot; Signoretti, Stefano; Suskauer, Stacy J; Tamburro, Robert; Turner, Michael; Yeates, Keith Owen; Zemek, Roger; Ala'i, Sherita; Esterlitz, Joy; Gay, Katelyn; Bellgowan, Patrick S F; Joseph, Kristen

2018-05-02

Through a partnership with the National Institute of Neurological Disorders and Stroke (NINDS), National Institutes of Health (NIH), and Department of Defense (DoD), the development of Sport-Related Concussion (SRC) Common Data Elements (CDEs) was initiated. The aim of this collaboration was to increase the efficiency and effectiveness of clinical research studies and clinical treatment outcomes, increase data quality, facilitate data sharing across studies, reduce study start-up time, more effectively aggregate information into metadata results, and educate new clinical investigators. The SRC CDE Working Group consisted of 34 worldwide experts in concussion from varied fields of related expertise, divided into three Subgroups: Acute (<72 hours post-concussion), Subacute (3 days-3 months post-concussion) and Persistent/Chronic (>3 months post-concussion). To develop CDEs, the Subgroups reviewed various domains, and then selected from, refined, and added to existing CDEs, case report forms and field-tested data elements from national registries and funded research studies. Recommendations were posted to the NINDS CDE Website for Public Review from February 2017 to April 2017. Following an internal Working Group review of recommendations, along with consideration of comments received from the Public Review period, the first iteration (Version 1.0) of the NINDS SRC CDEs was completed in June 2017. The recommendations include Core and Supplemental - Highly Recommended CDEs for cognitive data elements and symptom checklists, as well as other outcomes and endpoints (e.g., vestibular, oculomotor, balance, anxiety, depression) and sample case report forms (e.g., injury reporting, demographics, concussion history) for domains typically included in clinical research studies. The NINDS SRC CDEs and supporting documents are publicly available on the NINDS CDE website https://www.commondataelements.ninds.nih.gov/. Widespread use of CDEs by researchers and clinicians will facilitate consistent SRC clinical research and trial design, data sharing, and metadata retrospective analysis.

Effect of flour particle size and damaged starch on the quality of cookies.

PubMed

Barak, Sheweta; Mudgil, Deepak; Khatkar, B S

2014-07-01

Two wheat varieties 'C 306' and 'WH 542' were milled to obtain flour fractions of different particle sizes. Various physicochemical parameters such as wet and dry gluten, falling number, solvent retention capacity (SRC), alkaline water retention capacity (AWRC) and damaged starch content of the flour fractions were analyzed. The damaged starch values ranged from 5.14% to 14.79% for different flour fractions and increased significantly with decrease in particle size. AWRC and SRC of the flour fractions also increased with decrease in particle size. AWRC(r = 0.659) showed positive correlation and cookie spread ratio (r = -0.826) was strongly negatively correlated with the damaged starch levels. Hardness of the cookies in term of compression force showed increasing trend as damaged starch of the flour fractions increased. Spread ratio of the cookies ranged from 6.72 to 10.12. Wheat flour of particle size greater than 150 μm produced cookies with best quality.

Computational study of Gleevec and G6G reveals molecular determinants of kinase inhibitor selectivity

DOE PAGES

Lin, Yen -Lin; Meng, Yilin; Huang, Lei; ...

2014-10-22

Gleevec is a potent inhibitor of Abl tyrosine kinase but not of the highly homologous c-Src kinase. Because the ligand binds to an inactive form of the protein in which an Asp-Phe-Gly structural motif along the activation loop adopts a so-called DFG-out conformation, it was suggested that binding specificity was controlled by a “conformational selection” mechanism. In this context, the binding affinity displayed by the kinase inhibitor G6G poses an intriguing challenge. Although it possesses a chemical core very similar to that of Gleevec, G6G is a potent inhibitor of both Abl and c-Src kinases. Both inhibitors bind to themore » DFG-out conformation of the kinases, which seems to be in contradiction with the conformational selection mechanism. To address this issue and display the hidden thermodynamic contributions affecting the binding selectivity, molecular dynamics free energy simulations with explicit solvent molecules were carried out. Relative to Gleevec, G6G forms highly favorable van der Waals dispersive interactions upon binding to the kinases via its triazine functional group, which is considerably larger than the corresponding pyridine moiety in Gleevec. Upon binding of G6G to c-Src, these interactions offset the unfavorable free energy cost of the DFG-out conformation. When binding to Abl, however, G6G experiences an unfavorable free energy penalty due to steric clashes with the phosphate-binding loop, yielding an overall binding affinity that is similar to that of Gleevec. Such steric clashes are absent when G6G binds to c-Src, due to the extended conformation of the phosphate-binding loop.« less

Coal liquefaction process using pretreatment with a binary solvent mixture

DOEpatents

Miller, R.N.

1986-10-14

An improved process for thermal solvent refining or hydroliquefaction of non-anthracitic coal at elevated temperatures under hydrogen pressure in a hydrogen donor solvent comprises pretreating the coal with a binary mixture of an aromatic hydrocarbon and an aliphatic alcohol at a temperature below 300 C before the hydroliquefaction step. This treatment generally increases both conversion of coal and yields of oil. 1 fig.

Coal liquefaction process using pretreatment with a binary solvent mixture

DOEpatents

Miller, Robert N.

1986-01-01

An improved process for thermal solvent refining or hydroliquefaction of non-anthracitic coal at elevated temperatures under hydrogen pressure in a hydrogen donor solvent comprises pretreating the coal with a binary mixture of an aromatic hydrocarbon and an aliphatic alcohol at a temperature below 300.degree. C. before the hydroliquefaction step. This treatment generally increases both conversion of coal and yields of oil.

Specific Cx43 phosphorylation events regulate gap junction turnover in vivo

PubMed Central

Solan, Joell L.; Lampe, Paul D.

2014-01-01

Gap junctions, composed of proteins from the connexin gene family, are highly dynamic structures that are regulated by kinase-mediated signaling pathways and interactions with other proteins. Phosphorylation of Connexin43 (Cx43) at different sites controls gap junction assembly, gap junction size and gap junction turnover. Here we present a model describing how Akt, mitogen activated protein kinase (MAPK) and src kinase coordinate to regulate rapid turnover of gap junctions. Specifically, Akt phosphorylates Cx43 at S373 eliminating interaction with zona occludens-1 (ZO-1) allowing gap junctions to enlarge. Then MAPK and src phosphorylate Cx43 to initiate turnover. We integrate published data with new data to test and refine this model. Finally, we propose that differential coordination of kinase activation and Cx43 phosphorylation controls the specific routes of disassembly, e.g., annular junction formation or gap junctions can potentially “unzip” and be internalized/endocytosed into the cell that produced each connexin. PMID:24508467

Zinc sulfide liquefaction catalyst

DOEpatents

Garg, Diwakar

1984-01-01

A process for the liquefaction of carbonaceous material, such as coal, is set forth wherein coal is liquefied in a catalytic solvent refining reaction wherein an activated zinc sulfide catalyst is utilized which is activated by hydrogenation in a coal derived process solvent in the absence of coal.

Catalysts for coal liquefaction processes

DOEpatents

Garg, Diwakar

1986-01-01

Improved catalysts for catalytic solvent refining or hydroliquefaction of non-anthracitic coal at elevated temperatures under hydrogen pressure in a hydrogen donor solvent comprise a combination of zinc or copper, or a compound thereof, and a Group VI or non-ferrous Group VIII metal, or a compound thereof.

Catalysts for coal liquefaction processes

DOEpatents

Garg, D.

1986-10-14

Improved catalysts for catalytic solvent refining or hydroliquefaction of non-anthracitic coal at elevated temperatures under hydrogen pressure in a hydrogen donor solvent comprise a combination of zinc or copper, or a compound thereof, and a Group VI or non-ferrous Group VIII metal, or a compound thereof.

Method of refining cracked oil by using metallic soaps. [desulfurization of cracked oils

DOE Office of Scientific and Technical Information (OSTI.GOV)

Masakichi, M.; Marunouchi, K.K.; Yoshimura, T.

1937-04-13

The method of refining cracked oil consists in dissolving oil-soluble heavy metallic soap of oleic acid in a volatile organic solvent which will disperse homogeneously in cracked oil; pouring the solution thus obtained slowly into cracked oil to effect dispersion naturally and homogeneously at room temperature in the cracked oil. This process serves to react the mercaptans in the cracked oil with the heavy metallic soap by a double decomposition reaction and to precipitate the mercaptans as insoluble metallic salts. The remaining liquid is distilled to separate it from the remaining solvent.

Re-refining of waste petroleum by competing solubility characteristics

NASA Astrophysics Data System (ADS)

Byars, Michael Steven

1998-11-01

The United States produces over 1.3 billion gallons of used oil per year. Of the 1.3 billion gallons about 60% is used as fuel, nearly 20% is dumped into the environment, 13% is placed in landfills, 2% is re-refined into lube oil, and the remaining is either used for other purposes or incinerated. This is a great potential source of lubricating oil. The work presented here is a solvent extraction process using a solvent (highly miscible with the oil) and a co-solvent (slightly miscible with the oil). Extractions using isopropanol, ethanol, methyl tert-butylether and methanol are presented. The criteria used for evaluation of the extraction processes are yield, product viscosity index, and ash percent. The solvent/co-solvent combinations of MTBE and ethanol performed best and had the advantage of a common solvent/co-solvent in all extraction steps. The extraction process that provided the best results was a two step process using a combination solvent of MTBE and ethanol. The used oil was first extracted using MTBE/ethanol. The extracted oil was then contacted with a solvent combination composed of 80% ethanol. This solvent combination extracted the remaining additives from the oil. The recovered oil was nearly 60% by weight with a high viscosity index and no ash content. A preliminary battery limits design and economic analysis of the process was performed. A 500 bbl/day plant would have a capital cost of 1.9 million and an annual operation cost of 310,000. The plant as designed would produce 300 bbl/day of lube feedstock and have an ROI of 19%.

Inert Reassessment Document for 2-methyl-2,4-pentanediol - CAS No. 107-41-5

EPA Pesticide Factsheets

2-methyl-2,4-pentanediol is used as a chemical intermediate, a selective solvent in petroleum refining, a component of hydraulic fluids, an additive for cement, a component of industrial coatings, a solvent for inks, an additive for fuel and lubricants

Preparative separation of six rhynchophylla alkaloids from Uncaria macrophylla wall by pH-zone refining counter-current chromatography.

PubMed

Zhang, Qinghai; Lin, Changhu; Duan, Wenjuan; Wang, Xiao; Luo, Aiqin

2013-12-12

pH-Zone refining counter-current chromatography was successfully applied to the preparative isolation and purification of six alkaloids from the ethanol extracts of Uncaria macrophylla Wall. Because of the low content of alkaloids (about 0.2%, w/w) in U. macrophylla Wall, the target compounds were enriched by pH-zone refining counter-current chromatography using a two-phase solvent system composed of petroleum ether-ethyl acetate-isopropanol-water (2:6:3:9, v/v), adding 10 mM triethylamine in organic stationary phase and 5 mM hydrochloric acid in aqueous mobile phase. Then pH-zone refining counter-current chromatography using the other two-phase solvent system was used for final purification. Six target compounds were finally isolated and purified by following two-phase solvent system composed of methyl tert-butyl ether (MTBE)-acetonitrile-water (4:0.5:5, v/v), adding triethylamine (TEA) (10 mM) to the organic phase and HCl (5 mM) to aqueous mobile phase. The separation of 2.8 g enriched total alkaloids yielded 36 mg hirsutine, 48 mg hirsuteine, 82 mg uncarine C, 73 mg uncarine E, 163 mg rhynchophylline, and 149 mg corynoxeine, all with purities above 96% as verified by HPLC Their structures were identified by electrospray ionization-mass spectrometry (ESI-MS) and 1H-NMR spectroscopy.

Removing micron size particles from coal liquids

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rodgers, B.R.; Westmoreland, P.R.

This paper reports results of an investigation which was undertaken in order to improve purification of liquid fuels obtained by coal liquefaction processes. It is shown that settling and filtration rates increased substantially after agglomeration. And, ground coal was found to be an economical substitute for diatomaceous earth in filtration. The effects of certain solvents on the agglomerating tendencies of solids in the unfiltered oil (UFO) from the SRC and COED processes were determined by oil immersion microscopy. The significant results obtained by these experiments are listed. Economic advantages are presented. 13 references.

European solvent industry group generic exposure scenario risk and exposure tool

PubMed Central

Zaleski, Rosemary T; Qian, Hua; Zelenka, Michael P; George-Ares, Anita; Money, Chris

2014-01-01

The European Solvents Industry Group (ESIG) Generic Exposure Scenario (GES) Risk and Exposure Tool (EGRET) was developed to facilitate the safety evaluation of consumer uses of solvents, as required by the European Union Registration, Evaluation and Authorization of Chemicals (REACH) Regulation. This exposure-based risk assessment tool provides estimates of both exposure and risk characterization ratios for consumer uses. It builds upon the consumer portion of the European Center for Ecotoxicology and Toxicology of Chemicals (ECETOC) Targeted Risk Assessment (TRA) tool by implementing refinements described in ECETOC TR107. Technical enhancements included the use of additional data to refine scenario defaults and the ability to include additional parameters in exposure calculations. Scenarios were also added to cover all frequently encountered consumer uses of solvents. The TRA tool structure was modified to automatically determine conditions necessary for safe use. EGRET reports results using specific standard phrases in a format consistent with REACH exposure scenario guidance, in order that the outputs can be readily assimilated within safety data sheets and similar information technology systems. Evaluation of tool predictions for a range of commonly encountered consumer uses of solvents found it provides reasonable yet still conservative exposure estimates. PMID:23361440

European solvent industry group generic exposure scenario risk and exposure tool.

PubMed

Zaleski, Rosemary T; Qian, Hua; Zelenka, Michael P; George-Ares, Anita; Money, Chris

2014-01-01

The European Solvents Industry Group (ESIG) Generic Exposure Scenario (GES) Risk and Exposure Tool (EGRET) was developed to facilitate the safety evaluation of consumer uses of solvents, as required by the European Union Registration, Evaluation and Authorization of Chemicals (REACH) Regulation. This exposure-based risk assessment tool provides estimates of both exposure and risk characterization ratios for consumer uses. It builds upon the consumer portion of the European Center for Ecotoxicology and Toxicology of Chemicals (ECETOC) Targeted Risk Assessment (TRA) tool by implementing refinements described in ECETOC TR107. Technical enhancements included the use of additional data to refine scenario defaults and the ability to include additional parameters in exposure calculations. Scenarios were also added to cover all frequently encountered consumer uses of solvents. The TRA tool structure was modified to automatically determine conditions necessary for safe use. EGRET reports results using specific standard phrases in a format consistent with REACH exposure scenario guidance, in order that the outputs can be readily assimilated within safety data sheets and similar information technology systems. Evaluation of tool predictions for a range of commonly encountered consumer uses of solvents found it provides reasonable yet still conservative exposure estimates.

Effect of dynamic surface polarization on the oxidative stability of solvents in nonaqueous Li-O 2 batteries

NASA Astrophysics Data System (ADS)

Khetan, Abhishek; Pitsch, Heinz; Viswanathan, Venkatasubramanian

2017-09-01

Polarization-induced renormalization of the frontier energy levels of interacting molecules and surfaces can cause significant shifts in the excitation and transport behavior of electrons. This phenomenon is crucial in determining the oxidative stability of nonaqueous electrolytes in high-energy density electrochemical systems such as the Li-O2 battery. On the basis of partially self-consistent first-principles Sc G W0 calculations, we systematically study how the electronic energy levels of four commonly used solvent molecules, namely, dimethylsulfoxide (DMSO), dimethoxyethane (DME), tetrahydrofuran (THF), and acetonitrile (ACN), renormalize when physisorbed on the different stable surfaces of Li2O2 , the main discharge product. Using band level alignment arguments, we propose that the difference between the solvent's highest occupied molecular orbital (HOMO) level and the surface's valence-band maximum (VBM) is a refined metric of oxidative stability. This metric and a previously used descriptor, solvent's gas phase HOMO level, agree quite well for physisorbed cases on pristine surfaces where ACN is oxidatively most stable followed by DME, THF, and DMSO. However, this effect is intrinsically linked to the surface chemistry of the solvent's interaction with the surface states and defects, and depends strongly on their nature. We conclusively show that the propensity of solvent molecules to oxidize will be significantly higher on Li2O2 surfaces with defects as compared to pristine surfaces. This suggests that the oxidative stability of a solvent is dynamic and is a strong function of surface electronic properties. Thus, while gas phase HOMO levels could be used for preliminary solvent candidate screening, a more refined picture of solvent stability requires mapping out the solvent stability as a function of the state of the surface under operating conditions.

Refining of metallurgical-grade silicon

NASA Technical Reports Server (NTRS)

Dietl, J.

1986-01-01

A basic requirement of large scale solar cell fabrication is to provide low cost base material. Unconventional refining of metallurical grade silicon represents one of the most promising ways of silicon meltstock processing. The refining concept is based on an optimized combination of metallurgical treatments. Commercially available crude silicon, in this sequence, requires a first pyrometallurgical step by slagging, or, alternatively, solvent extraction by aluminum. After grinding and leaching, high purity qualtiy is gained as an advanced stage of refinement. To reach solar grade quality a final pyrometallurgical step is needed: liquid-gas extraction.

Catalytic coal liquefaction process

DOEpatents

Garg, D.; Sunder, S.

1986-12-02

An improved process for catalytic solvent refining or hydroliquefaction of non-anthracitic coal at elevated temperatures under hydrogen pressure in a solvent comprises using as catalyst a mixture of a 1,2- or 1,4-quinone and an alkaline compound, selected from ammonium, alkali metal, and alkaline earth metal oxides, hydroxides or salts of weak acids. 1 fig.

Catalytic coal liquefaction process

DOEpatents

Garg, Diwakar; Sunder, Swaminathan

1986-01-01

An improved process for catalytic solvent refining or hydroliquefaction of non-anthracitic coal at elevated temperatures under hydrogen pressure in a solvent comprises using as catalyst a mixture of a 1,2- or 1,4-quinone and an alkaline compound, selected from ammonium, alkali metal, and alkaline earth metal oxides, hydroxides or salts of weak acids.

A New Strategy to Refine Crude Indian Sardine Oil.

PubMed

Charanyaa, S; Belur, Prasanna D; Regupathi, I

2017-05-01

Current work aims to develop a refining process for removing phospholipids, free fatty acids (FFA), and metal ions without affecting n-3 polyunsaturated fatty acid (n-3 PUFA) esters present in the crude Indian sardine oil. Sardine oil was subjected to degumming with various acids (orthophosphoric acid, acetic acid, and lactic acid), conventional and membrane assisted deacidification using various solvents (methanol, ethanol, propanol and butanol) and bleaching with bleaching agents (GAC, activated earth and bentonite) and all the process parameters were further optimized. Degumming with 5%(w/w) ortho phosphoric acid, two stage solvent extraction with methanol at 1:1 (w/w) in each stage and bleaching with 3% (w/w) activated charcoal loading, at 80ºC for 10 minutes resulted in the reduction of phospholipid content to 5.66 ppm from 612.66 ppm, FFA to 0.56% from 5.64% with the complete removal of iron and mercury. Under these conditions, the obtained bleached oil showed an enhancement of n-3 PUFA from 16.39 % (11.19 Eicosapentaenoic acid (EPA) + 5.20 Docosahexaenoic acid (DHA)) to 17.91 % (11.81 EPA + 6.1 DHA). Replacing conventional solvent extraction with membrane deacidification using microporous, hydrophobic polytetrafluoroethylene membrane (PTFE), resulted in a lesser solvent residue (0.25% (w/w)) in the deacidified oil. In view of lack of reports on refining of n-3 PUFA rich marine oils without concomitant loss of n-3 PUFA, this report is significant.

Space Station automation and robotics

NASA Technical Reports Server (NTRS)

1987-01-01

A group of fifteen students in the Electrical Engineering Department at the University of Maryland, College Park, has been involved in a design project under the sponsorship of NASA Headquarters, NASA Goddard Space Flight Center and the Systems Research Center (SRC) at UMCP. The goal of the NASA/USRA project was to first obtain a refinement of the design work done in Spring 1986 on the proposed Mobile Remote Manipulator System (MRMS) for the Space Station. This was followed by design exercises involving the OMV and two armed service vehicle. Three students worked on projects suggested by NASA Goddard scientists for ten weeks this past summer. The knowledge gained from the summer design exercise has been used to improve our current design of the MRMS. To this end, the following program was undertaken for the Fall semester 1986: (1) refinement of the MRMS design; and (2) addition of vision capability to our design.

Literature survey of properties of synfuels derived from coal

NASA Technical Reports Server (NTRS)

Reynolds, T. W.; Niedzwiecki, R. W.; Clark, J. S.

1980-01-01

A literature survey of the properties of synfuels for ground-based gas turbine applications is presented. Four major concepts for converting coal into liquid fuels are described: solvent extraction, catalytic liquefaction, pyrolysis, and indirect liquefaction. Data on full range syncrudes, various distillate cuts, and upgraded products are presented for fuels derived from various processes, including H-coal, synthoil, solvent-refined coal, donor solvent, zinc chloride hydrocracking, co-steam, and flash pyrolysis. Some typical ranges of data for coal-derived low Btu gases are also presented.

New biphasic solvent system based on cyclopentyl methyl ether for the purification of a non-polar synthetic peptide by pH-zone refining centrifugal partition chromatography.

PubMed

Amarouche, Nassima; Boudesocque, Leslie; Borie, Nicolas; Giraud, Matthieu; Forni, Luciano; Butte, Alessandro; Edwards, Florence; Renault, Jean-Hugues

2014-06-01

A new type 1 ternary biphasic system composed of cyclopentyl methyl ether, dimethylformamide and water was developed, characterized and successfully used for the purification of a lipophilic, protected peptide by pH-zone refining centrifugal partition chromatography. The protected peptide is an 8-mer, key intermediate in bivalirudin (Angiomax®) synthesis and shows a very low solubility in the solvents usually used in liquid chromatography. All ionic groups, except the N-terminal end of the peptide, are protected by a benzyl group. The purification of this peptide was achieved with a purity of about 99.04% and a recovery of 94% using the new ternary biphasic system cyclopentyl methyl ether/dimethylformamide/water (49:40:11, v/v) in the descending pH-zone refining mode with triethylamine (28 mM) as the retainer and methanesulfonic acid (18 mM) as the eluter. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Chemical analysis and biological testing of materials from the EDS coal liquefaction process: a status report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Later, D.W.; Pelroy, R.A.; Wilson, B.W.

1984-05-01

Representative process materials were obtained from the EDS pilot plant for chemical and biological analyses. These materials were characterized for biological activity and chemical composition using a microbial mutagenicity assay and chromatographic and mass spectrometric analytical techniques. The two highest boiling distillation cuts, as well as process solvent (PS) obtained from the bottoms recycle mode operation, were tested for initiation of mouse skin tumorigenicity. All three materials were active; the crude 800/sup 0 +/F cut was substantially more potent than the crude bottoms recycle PS or 750 to 800/sup 0/F distillate cut. Results from chemical analyses showed the EDS materials,more » in general, to be more highly alkylated and have higher hydroaromatic content than analogous SRC II process materials (no in-line process hydrogenation) used for comparison. In the microbial mutagenicity assays the N-PAC fractions showed greater activity than did the aliphatic hydrocarbon, hydroxy-PAH, or PAH fractions, although mutagenicity was detected in certain PAH fractions by a modified version of the standard microbial mutagenicity assay. Mutagenic activities for the EDS materials were lower, overall, than those for the corresponding materials from the SRC II process. The EDS materials produced under different operational modes had distinguishable differences in both their chemical constituency and biological activity. The primary differences between the EDS materials studied here and their SRC II counterparts used for comparison are most likely attributable to the incorporation of catalytic hydrogenation in the EDS process. 27 references, 28 figures, 27 tables.« less

Spectroscopic characterization of the SH2- and active site-directed peptide sequences of a bivalent Src kinase inhibitor.

PubMed

Desamero, Ruel Z B; Kang, Jeonghee; Dol, Chrystel; Chinwong, Justina; Walters, Karim; Sivarajah, Thulashie; Profit, Adam A

2009-07-01

The spectral properties of the SH2 and active site-directed sequences of the bivalent Src kinase inhibitor Ac-EELL(F5)Phe-(GABA)3-pYEEIE-amide (1) have been determined. Ac-pYEEIE-amide (2) and AcEELL(F5)Phe-amide (3), as well as the amino acids phosphotyrosine (pTyr) and pentafluorophenylalanine (F5)Phe, have been characterized by electronic absorption, fluorescence, and vibrational spectroscopy. Specific and unique marker bands that originate from the phosphate group of pTyr and the fluorinated aromatic ring of (F5)Phe have been identified, with the latter showing some solvent dependence. Peptide 2 was found to have excitation and emission wavelengths emanating from pTyr at 268 and 295 nm, respectively, whereas peptide 3 displayed excitation and emission peaks attributable to (F5)Phe at 274 and 315 nm, respectively. Fourier transform infrared (FT-IR) analysis of the amino acid pTyr identified distinct marker bands at approximately 930, 1090, and 1330 cm(-1) that could be attributed to the phosphate group. These markers were also observed in the IR spectrum of peptide 2. Likewise, peptide 3 displayed a characteristic C-F stretching mode at 961 cm(-1) due to the presence of (F5)Phe, including two C-F reporting ring modes at 1509 and 1527 cm(-1). Identifying and monitoring spectroscopic changes in these marker bands may afford a means to observe the molecular interactions that occur when peptides 1-3 bind to the Src kinase.

4-Hydroxynonenal activates Src through a non-canonical pathway that involves EGFR/PTP1B

PubMed Central

Zhang, Hongqiao; Forman, Henry Jay

2015-01-01

Src, a non-receptor protein tyrosine kinase involved in many biological processes, can be activated through both redox-dependent and independent mechanisms. 4-Hydroxy-2-nonenal (HNE) is a lipid peroxidation product that is increased in pathophysiological conditions associated with Src activation. This study examined how HNE activates human c-Src. In the canonical pathway Src activation is initiated by dephosphorylation of pTyr530 followed by conformational change that causes Src auto-phosphorylation at Tyr419 and its activation. HNE increased Src activation in both dose- and time-dependent manner, while it also increased Src phosphorylation at Tyr530 (pTyr530 Src), suggesting that HNE activated Src via a non-canonical mechanism. Protein tyrosine phosphatase 1B inhibitor (539741), at concentrations that increased basal pTyr530 Src, also increased basal Src activity and significantly reduced HNE-mediated Src activation. The EGFR inhibitor, AG1478, and EGFR silencing, abrogated HNE-mediated EGFR activation and inhibited basal and HNE-induced Src activity. In addition, AG1478 also eliminated the increase of basal Src activation by a PTP1B inhibitor. Taken together these data suggest that HNE can activate Src partly through a non-canonical pathway involving activation of EGFR and inhibition of PTP1B. PMID:26453921

v-Src-driven transformation is due to chromosome abnormalities but not Src-mediated growth signaling.

PubMed

Honda, Takuya; Morii, Mariko; Nakayama, Yuji; Suzuki, Ko; Yamaguchi, Noritaka; Yamaguchi, Naoto

2018-01-18

v-Src is the first identified oncogene product and has a strong tyrosine kinase activity. Much of the literature indicates that v-Src expression induces anchorage-independent and infinite cell proliferation through continuous stimulation of growth signaling by v-Src activity. Although all of v-Src-expressing cells are supposed to form transformed colonies, low frequencies of v-Src-induced colony formation have been observed so far. Using cells that exhibit high expression efficiencies of inducible v-Src, we show that v-Src expression causes cell-cycle arrest through p21 up-regulation despite ERK activation. v-Src expression also induces chromosome abnormalities and unexpected suppression of v-Src expression, leading to p21 down-regulation and ERK inactivation. Importantly, among v-Src-suppressed cells, only a limited number of cells gain the ability to re-proliferate and form transformed colonies. Our findings provide the first evidence that v-Src-driven transformation is attributed to chromosome abnormalities, but not continuous stimulation of growth signaling, possibly through stochastic genetic alterations.

Lack of Csk-mediated negative regulation in a unicellular SRC kinase.

PubMed

Schultheiss, Kira P; Suga, Hiroshi; Ruiz-Trillo, Iñaki; Miller, W Todd

2012-10-16

Phosphotyrosine-based signaling plays a vital role in cellular communication in multicellular organisms. Unexpectedly, unicellular choanoflagellates (the closest phylogenetic group to metazoans) possess numbers of tyrosine kinases that are comparable to those in complex metazoans. Here, we have characterized tyrosine kinases from the filasterean Capsaspora owczarzaki, a unicellular protist representing the sister group to choanoflagellates and metazoans. Two Src-like tyrosine kinases have been identified in C. owczarzaki (CoSrc1 and CoSrc2), both of which have the arrangement of SH3, SH2, and catalytic domains seen in mammalian Src kinases. In Capsaspora cells, CoSrc1 and CoSrc2 localize to punctate structures in filopodia that may represent primordial focal adhesions. We have cloned, expressed, and purified both enzymes. CoSrc1 and CoSrc2 are active tyrosine kinases. Mammalian Src kinases are normally regulated in a reciprocal fashion by autophosphorylation in the activation loop (which increases activity) and by Csk-mediated phosphorylation of the C-terminal tail (which inhibits activity). Similar to mammalian Src kinases, the enzymatic activities of CoSrc1 and CoSrc2 are increased by autophosphorylation in the activation loop. We have identified a Csk-like kinase (CoCsk) in the genome of C. owczarzaki. We cloned, expressed, and purified CoCsk and found that it has no measurable tyrosine kinase activity. Furthermore, CoCsk does not phosphorylate or regulate CoSrc1 or CoSrc2 in cells or in vitro, and CoSrc1 and CoSrc2 are active in Capsaspora cell lysates. Thus, the function of Csk as a negative regulator of Src family kinases appears to have arisen with the emergence of metazoans.

pH-zone-refining elution-extrusion countercurrent chromatography: Separation of hydroxyanthraquinones from Cassiae semen.

PubMed

Bu, Zhisi; Lv, Liqiong; Li, Xingnuo; Chu, Chu; Tong, Shengqiang

2017-11-01

Seven hydroxyanthraquinones were successfully separated from the traditional Chinese medicinal herb Cassiae semen by conventional and pH-zone-refining countercurrent chromatography with an environmentally friendly biphasic solvent system, in which elution-extrusion mode was investigated for pH-zone-refining countercurrent chromatography for the first time. A two-phase solvent system composed of n-hexane/ethyl acetate/ethanol/water (5:3:4:4, v/v/v/v) was used for the conventional countercurrent chromatography while the same system with a different volume ratio n-hexane/ethyl acetate/ethanol/water (3:5:2:6, v/v/v/v) was used for pH-zone-refining countercurrent chromatography, in which 20 mmol/L of trifluoroacetic acid was added in the organic phase as a retainer and 15 mmol/L of ammonia was added to the aqueous phase as an eluter. A 400 mg crude sample could be well separated by pH-zone-refining countercurrent chromatography, yielding 53 mg of aurantio-obtusin, 40 mg of chryso-obtusin, 18 mg of obtusin, 24 mg of obtusifolin, 10 mg of emodin, and 105 mg of the mixture of chrysophanol and physcion with a purity of over 95.8, 95.7, 96.9, 93.5, 97.4, 77.1, and 19.8%, as determined by high-performance liquid chromatography. Furthermore, the difference in elution sequence between conventional and pH-zone-refining mode was observed and discussed. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Dasatinib is preclinically active against Src-overexpressing human transitional cell carcinoma of the urothelium with activated Src signaling.

PubMed

Levitt, Jonathan M; Yamashita, Hideyuki; Jian, Weiguo; Lerner, Seth P; Sonpavde, Guru

2010-05-01

Dasatinib is an orally administered multitargeted kinase inhibitor that targets Src family tyrosine kinases, Abl, c-Kit, and PDGFR. A preclinical study was conducted to evaluate dasatinib alone or combined with cisplatin for human transitional cell carcinoma (TCC). Expression of Src in a human TCC tissue microarray was evaluated by immunohistochemistry. The activity of dasatinib and/or cisplatin was evaluated in six human TCC cell lines. Western blot was done to assess Src and phosphorylated-Src (p-Src) expression. The activity of dasatinib alone and in combination with cisplatin was determined in murine subcutaneous xenografts. Sixty-two percent to 75% of human TCC expressed Src. Dasatinib displayed significant antiproliferative activity at nanomolar concentrations against two human TCC cell lines (RT4 and Hu456) that exhibited high Src and p-Src expression and were cisplatin-resistant. RT4 cells were the most sensitive and displayed the highest level of Src pathway activation (p-Src/Src ratio). Dasatinib downregulated p-Src in either sensitive or resistant cells. TCC cells that were sensitive to cisplatin (5637 and TCC-SUP) were highly resistant to dasatinib and exhibited low Src expression. Dasatinib showed antitumor activity in RT4 murine xenografts, and the combination of dasatinib and cisplatin was significantly more active than placebo. Combination dasatinib plus cisplatin significantly inhibited proliferation and promoted apoptosis in vivo. In conclusion, dasatinib displayed significant preclinical antitumor activity against Src-overexpressing human TCC with active Src signaling and was highly active in combination with cisplatin in vivo. Further clinical development might be warranted in selected human subjects.

Patient-derived models of acquired resistance can identify effective drug combinations for cancer.

PubMed

Crystal, Adam S; Shaw, Alice T; Sequist, Lecia V; Friboulet, Luc; Niederst, Matthew J; Lockerman, Elizabeth L; Frias, Rosa L; Gainor, Justin F; Amzallag, Arnaud; Greninger, Patricia; Lee, Dana; Kalsy, Anuj; Gomez-Caraballo, Maria; Elamine, Leila; Howe, Emily; Hur, Wooyoung; Lifshits, Eugene; Robinson, Hayley E; Katayama, Ryohei; Faber, Anthony C; Awad, Mark M; Ramaswamy, Sridhar; Mino-Kenudson, Mari; Iafrate, A John; Benes, Cyril H; Engelman, Jeffrey A

2014-12-19

Targeted cancer therapies have produced substantial clinical responses, but most tumors develop resistance to these drugs. Here, we describe a pharmacogenomic platform that facilitates rapid discovery of drug combinations that can overcome resistance. We established cell culture models derived from biopsy samples of lung cancer patients whose disease had progressed while on treatment with epidermal growth factor receptor (EGFR) or anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors and then subjected these cells to genetic analyses and a pharmacological screen. Multiple effective drug combinations were identified. For example, the combination of ALK and MAPK kinase (MEK) inhibitors was active in an ALK-positive resistant tumor that had developed a MAP2K1 activating mutation, and the combination of EGFR and fibroblast growth factor receptor (FGFR) inhibitors was active in an EGFR mutant resistant cancer with a mutation in FGFR3. Combined ALK and SRC (pp60c-src) inhibition was effective in several ALK-driven patient-derived models, a result not predicted by genetic analysis alone. With further refinements, this strategy could help direct therapeutic choices for individual patients. Copyright © 2014, American Association for the Advancement of Science.

Aldosterone rapidly activates Src kinase in M-1 cells involving the mineralocorticoid receptor and HSP84.

PubMed

Braun, Sabine; Lösel, Ralf; Wehling, Martin; Boldyreff, Brigitte

2004-07-16

We investigated the effect of aldosterone on Src kinase. In the kidney cell line, M-1 aldosterone leads to a >2-fold transient activation of Src kinase seen as early as 2 min after aldosterone administration. Maximal Src kinase activation was measured at an aldosterone concentration of 1 nM. In parallel to activation, autophosphorylation at Tyr-416 of Src kinase increased. Src kinase activation was blocked by spironolactone. Aldosterone led to increased association of Src with HSP84. Furthermore, rapamycin blocked aldosterone-induced Src activation. We conclude that Src activation by aldosterone is mediated through the mineralocorticoid receptor and HSP84.

Coal Liquefaction desulfurization process

DOEpatents

Givens, Edwin N.

1983-01-01

In a solvent refined coal liquefaction process, more effective desulfurization of the high boiling point components is effected by first stripping the solvent-coal reacted slurry of lower boiling point components, particularly including hydrogen sulfide and low molecular weight sulfur compounds, and then reacting the slurry with a solid sulfur getter material, such as iron. The sulfur getter compound, with reacted sulfur included, is then removed with other solids in the slurry.

Roles of Raft-Anchored Adaptor Cbp/PAG1 in Spatial Regulation of c-Src Kinase

PubMed Central

Oneyama, Chitose; Suzuki, Takashi; Okada, Masato

2014-01-01

The tyrosine kinase c-Src is upregulated in numerous human cancers, implying a role for c-Src in cancer progression. Previously, we have shown that sequestration of activated c-Src into lipid rafts via a transmembrane adaptor, Cbp/PAG1, efficiently suppresses c-Src-induced cell transformation in Csk-deficient cells, suggesting that the transforming activity of c-Src is spatially regulated via Cbp in lipid rafts. To dissect the molecular mechanisms of the Cbp-mediated regulation of c-Src, a combined analysis was performed that included mathematical modeling and in vitro experiments in a c-Src- or Cbp-inducible system. c-Src activity was first determined as a function of c-Src or Cbp levels, using focal adhesion kinase (FAK) as a crucial c-Src substrate. Based on these experimental data, two mathematical models were constructed, the sequestration model and the ternary model. The computational analysis showed that both models supported our proposal that raft localization of Cbp is crucial for the suppression of c-Src function, but the ternary model, which includes a ternary complex consisting of Cbp, c-Src, and FAK, also predicted that c-Src function is dependent on the lipid-raft volume. Experimental analysis revealed that c-Src activity is elevated when lipid rafts are disrupted and the ternary complex forms in non-raft membranes, indicating that the ternary model accurately represents the system. Moreover, the ternary model predicted that, if Cbp enhances the interaction between c-Src and FAK, Cbp could promote c-Src function when lipid rafts are disrupted. These findings underscore the crucial role of lipid rafts in the Cbp-mediated negative regulation of c-Src-transforming activity, and explain the positive role of Cbp in c-Src regulation under particular conditions where lipid rafts are perturbed. PMID:24675741

SH2 Ligand-Like Effects of Second Cytosolic Domain of Na/K-ATPase α1 Subunit on Src Kinase.

PubMed

Banerjee, Moumita; Duan, Qiming; Xie, Zijian

2015-01-01

Our previous studies have suggested that the α1 Na/K-ATPase interacts with Src to form a receptor complex. In vitro binding assays indicate an interaction between second cytosolic domain (CD2) of Na/K-ATPase α1 subunit and Src SH2 domain. Since SH2 domain targets Src to specific signaling complexes, we expressed CD2 as a cytosolic protein and studied whether it could act as a Src SH2 ligand in LLC-PK1 cells. Co-immunoprecipitation analyses indicated a direct binding of CD2 to Src, consistent with the in vitro binding data. Functionally, CD2 expression increased basal Src activity, suggesting a Src SH2 ligand-like property of CD2. Consistently, we found that CD2 expression attenuated several signaling pathways where Src plays an important role. For instance, although it increased surface expression of Na/K-ATPase, it decreased ouabain-induced activation of Src and ERK by blocking the formation of Na/K-ATPase/Src complex. Moreover, it also attenuated cell attachment-induced activation of Src/FAK. Consequently, CD2 delayed cell spreading, and inhibited cell proliferation. Furthermore, these effects appear to be Src-specific because CD2 expression had no effect on EGF-induced activation of EGF receptor and ERK. Hence, the new findings indicate the importance of Na/K-ATPase/Src interaction in ouabain-induced signal transduction, and support the proposition that the CD2 peptide may be utilized as a Src SH2 ligand capable of blocking Src-dependent signaling pathways via a different mechanism from a general Src kinase inhibitor.

Identification and functional characterization of an Src homology domain 3 domain-binding site on Cbl.

PubMed

Sanjay, Archana; Miyazaki, Tsuyoshi; Itzstein, Cecile; Purev, Enkhtsetseg; Horne, William C; Baron, Roland

2006-12-01

Cbl is an adaptor protein and ubiquitin ligase that binds and is phosphorylated by the nonreceptor tyrosine kinase Src. We previously showed that the primary interaction between Src and Cbl is mediated by the Src homology domain 3 (SH3) of Src binding to proline-rich sequences of Cbl. The peptide Cbl RDLPPPPPPDRP(540-551), which corresponds to residues 540-551 of Cbl, inhibited the binding of a GST-Src SH3 fusion protein to Cbl, whereas RDLAPPAPPPDR(540-551) did not, suggesting that Src binds to this site on Cbl in a class I orientation. Mutating prolines 543-548 reduced Src binding to the Cbl 479-636 fragment significantly more than mutating the prolines in the PPVPPR(494-499) motif, which was previously reported to bind Src SH3. Mutating Cbl prolines 543-548 to alanines substantially reduced Src binding to Cbl, Src-induced phosphorylation of Cbl, and the inhibition of Src kinase activity by Cbl. Expressing the mutated Cbl in osteoclasts induced a moderate reduction in bone-resorbing activity and increased amounts of Src protein. In contrast, disabling the tyrosine kinase-binding domain of full-length Cbl by mutating glycine 306 to glutamic acid, and thereby preventing the previously described binding of the tyrosine kinase-binding domain to the Src phosphotyrosine 416, had no effect on Cbl phosphorylation, the inhibition of Src activity by full-length Cbl, or bone resorption. These data indicate that the Cbl RDLPPPP(540-546) sequence is a functionally important binding site for Src.

Efficient production of fatty acid methyl ester from waste activated bleaching earth using diesel oil as organic solvent.

PubMed

Kojima, Seiji; Du, Dongning; Sato, Masayasu; Park, Enoch Y

2004-01-01

Fatty acid methyl ester (FAME) production from waste activated bleaching earth (ABE) discarded by the crude oil refining industry was investigated using fossil fuel as a solvent in the esterification of triglycerides. Lipase from Candida cylindracea showed the highest stability in diesel oil. Using diesel oil as a solvent, 3 h was sufficient to obtain a yield of approximately 100% of FAME in the presence of 10% lipase from waste ABE. Kerosene was also a good solvent in the esterification of triglycerides embedded in the waste ABE. Fuel analysis showed that the FAME produced using diesel oil as a solvent complied with the Japanese diesel standard and the 10% residual carbon amount was lower than that of FAME produced using other solvents. Use of diesel oil as solvent in the FAME production from the waste ABE simplified the process, because there was no need to separate the organic solvent from the FAME-solvent mixture. These results demonstrate a promising reutilization method for the production of FAME, for use as a biodiesel, from industrial waste resources containing waste vegetable oils.

Genesis Solar Wind Sample 61422: Experiment in Variation of Sequence of Cleaning Solvent for Removing Carbon-Bearing Contamination

NASA Technical Reports Server (NTRS)

Allton, J. H.; Kuhlman, K. R.; Allums, K. K.; Gonzalez, C. P.; Jurewicz, A. J. G.; Burnett, D. S.; Woolum, D. S.

2015-01-01

The recovered Genesis collector fragments are heavily contaminated with crash-derived particulate debris. However, megasonic treatment with ultra-pure-water (UPW; resistivity (is) greater than18 meg-ohm-cm) removes essentially all particulate contamination greater than 5 microns in size [e.g.1] and is thus of considerable importance. Optical imaging of Si sample 60336 revealed the presence of a large C-rich particle after UPW treatment that was not present prior to UPW. Such handling contamination is occasionally observed, but such contaminants are normally easily removed by UPW cleaning. The 60336 particle was exceptional in that, surprisingly, it was not removed by additional UPW or by hot xylene or by aqua regia treatment. It was eventually removed by treatment with NH3-H2O2. Our best interpretation of the origin of the 60336 particle was that it was adhesive from the Post-It notes used to stabilize samples for transport from Utah after the hard landing. It is possible that the insoluble nature of the 60336 particle comes from interaction of the Post-It adhesive with UPW. An occasional bit of Post-It adhesive is not a major concern, but C particulate contamination also occurs from the heat shield of the Sample Return Capsule (SRC) and this is mixed with inorganic contamination from the SRC and the Utah landing site. If UPW exposure also produced an insoluble residue from SRC C, this would be a major problem in chemical treatments to produce clean surfaces for analysis. This paper reports experiments to test whether particulate contamination was removed more easily if UPW treatment was not used.

p68 Sam is a substrate of the insulin receptor and associates with the SH2 domains of p85 PI3K.

PubMed

Sánchez-Margalet, V; Najib, S

1999-07-23

The 68 kDa Src substrate associated during mitosis is an RNA binding protein with Src homology 2 and 3 domain binding sites. A role for Src associated in mitosis 68 as an adaptor protein in signaling transduction has been proposed in different systems such as T-cell receptors. In the present work, we have sought to assess the possible role of Src associated in mitosis 68 in insulin receptor signaling. We performed in vivo studies in HTC-IR cells and in vitro studies using recombinant Src associated in mitosis 68, purified insulin receptor and fusion proteins containing either the N-terminal or the C-terminal Src homology 2 domain of p85 phosphatidylinositol-3-kinase. We have found that Src associated in mitosis 68 is a substrate of the insulin receptor both in vivo and in vitro. Moreover, tyrosine-phosphorylated Src associated in mitosis 68 was found to associate with p85 phosphatidylinositol-3-kinase in response to insulin, as assessed by co-immunoprecipitation studies. Therefore, Src associated in mitosis 68 may be part of the signaling complexes of insulin receptor along with p85. In vitro studies demonstrate that Src associated in mitosis 68 associates with the Src homology 2 domains of p85 after tyrosine phosphorylation by the activated insulin receptor. Moreover, tyr-phosphorylated Src associated in mitosis 68 binds with a higher affinity to the N-terminal Src homology 2 domain of p85 compared to the C-terminal Src homology 2 domain of p85, suggesting a preferential association of Src associated in mitosis 68 with the N-terminal Src homology 2 domain of p85. This association may be important for the link of the signaling with RNA metabolism.

SGT1 is required in PcINF1/SRC2-1 induced pepper defense response by interacting with SRC2-1

PubMed Central

Liu, Zhi-qin; Liu, Yan-yan; Shi, Lan-ping; Yang, Sheng; Shen, Lei; Yu, Huan-xin; Wang, Rong-zhang; Wen, Jia-yu; Tang, Qian; Hussain, Ansar; Khan, Muhammad Ifnan; Hu, Jiong; Liu, Cai-ling; Zhang, Yang-wen; Cheng, Wei; He, Shui-lin

2016-01-01

PcINF1 was previously found to induce pepper defense response by interacting with SRC2-1, but the underlying mechanism remains uninvestigated. Herein, we describe the involvement of SGT1 in the PcINF1/SRC2-1-induced immunity. SGT1 was observed to be up-regulated by Phytophthora capsici inoculation and synergistically transient overexpression of PcINF1/SRC2-1 in pepper plants. SGT1-silencing compromised HR cell death, blocked H2O2 accumulation, and downregulated HR-associated and hormones-dependent marker genes’ expression triggered by PcINF1/SRC2-1 co-overexpression. The interaction between SRC2-1 and SGT1 was found by the yeast two hybrid system and was further confirmed by bimolecular fluorescence complementation and co-immunoprecipitation analyses. The SGT1/SRC2-1 interaction was enhanced by transient overexpression of PcINF1 and Phytophthora capsici inoculation, and SGT1-silencing attenuated PcINF1/SRC2-1 interaction. Additionally, by modulating subcellular localizations of SRC2-1, SGT1, and the interacting complex of SGT1/SRC2-1, it was revealed that exclusive nuclear targeting of the SGT1/SRC2-1 complex blocks immunity triggered by formation of SGT1/SRC2-1, and a translocation of the SGT1/SRC2-1 complex from the plasma membrane and cytoplasm to the nuclei upon the inoculation of P. capsici. Our data demonstrate that the SGT1/SRC2-1 interaction, and its nucleocytoplasmic partitioning, is involved in pepper’s immunity against P. capsici, thus providing a molecular link between Ca2+ signaling associated SRC2-1 and SGT1-mediated defense signaling. PMID:26898479

A refined method for the calculation of the Non-Methane Volatile Organic Compound emission estimate from Domestic Solvent Usage in Ireland from 1992 to 2014 - A case study for Ireland

NASA Astrophysics Data System (ADS)

Barry, Stephen; O'Regan, Bernadette

2016-08-01

This study describes a new methodology to calculate Non-Methane Volatile Organic Compounds from Domestic Solvent Use including Fungicides over the period 1992-2014. Improved emissions data compiled at a much more refined level can help policy-makers develop more effective policy's to address environmental issues. However, a number of problems were found when member states attempt to use national statistics for Domestic Solvent Use including Fungicides. For instance, EMEP/EEA (2013) provides no guidance regarding which activity data should be used, resulting in emission estimates being potentially inconsistent and un-comparable. Also, previous methods and emission factors described in the EMEP/EEA (2013) guidebook do not exactly match data collected by state agencies. This makes using national statistics difficult. In addition, EMEP/EEA (2013) use broader categories than necessary (e.g. Cosmetics Aerosol/Non Aerosol) to estimate emissions while activity data is available at a more refined level scale (e.g. Personal Cleaning Products, Hair Products, Cosmetics, Deodorants and Perfumes). This can make identifying the drivers of emissions unclear. This study builds upon Tzanidakis et al. (2012) whereby it provides a method for collecting activity data from state statistics, developed country specific emission factors based on a survey of 177 Irish products and importantly, used a new method to account for the volatility of organic compounds found in commonly available domestic solvent containing products. This is the first study to account for volatility based on the characteristics of organic compounds and therefore is considered a more accurate method of accounting for emissions from this emission source. The results of this study can also be used to provide a simple method for other member parties to account for the volatility of organic compounds using sectorial adjustment factors described here. For comparison purposes, emission estimates were calculated using the Tier 1 approach currently used in the emission inventory, using activity data and emission factors unadjusted for volatility and adjusted for volatility. The unadjusted estimate is useful, because it demonstrates the failure to properly account for volatility can produce significantly over-estimated emissions from the Domestic Solvent Usage sector. Unadjusted emissions were found to be 30% lower than the EMEP/EEA (2013) Tier 1 period in 2014. Emissions were found to reduce a further 20.9% when the volatility of the organic compounds was included. This new method shows that member parties may be significantly overestimating emissions from Domestic Solvent Use including pesticides and further work should include refining organic compound content and the sectorial adjustment factor of products.

Thyroid function in mice with compound heterozygous and homozygous disruptions of SRC-1 and TIF-2 coactivators: evidence for haploinsufficiency.

PubMed

Weiss, Roy E; Gehin, Martine; Xu, Jianming; Sadow, Peter M; O'Malley, Bert W; Chambon, Pierre; Refetoff, Samuel

2002-04-01

Steroid receptor coactivator (SRC)-1 and transcriptional intermediary factor (TIF)-2 are homologous nuclear receptor coactivators. We have investigated their possible redundancy as thyroid hormone (TH) coactivators by measuring thyroid function in compound SRC-1 and TIF-2 knock out (KO) mice. Whereas SRC-1 KO (SRC-1(-/-)) mice are resistant to TH and SRC-1(+/-) are not, we now demonstrate that TIF-2 KO (TIF-2(-/-)) mice have normal thyroid function. Yet double heterozygous, SRC-1(+/-)/TIF-2(+/-) mice manifested resistance to TH of a similar degree as that in mice completely deficient in SRC-1. KO of both SRC-1 and TIF-2 resulted in marked increases of serum TH and thyrotropin concentrations. This work demonstrates gene dosage effect in nuclear coactivators manifesting as haploinsufficiency and functional redundancy of SRC-1 and TIF-2.

Design and Analysis of Chronic Aquatic Tests of Toxicity with Daphnia magna.

DTIC Science & Technology

1981-12-01

surface waters. From that need evolved numerous standard toxicity tests. Aquatic toxicologists and biologists developed, refine,, and standard- ized many...experimental categorization summary sheets prepared by Dr. William van der Schalie, which is shown in Table I.I. 7 j-. " .’?, i...partial solution to this dilema can be obtained by studying the effects of the solvent alone. If the solvent by itself produces no toxic responses at

Analysis of biomolecular solvation sites by 3D-RISM theory.

PubMed

Sindhikara, Daniel J; Hirata, Fumio

2013-06-06

We derive, implement, and apply equilibrium solvation site analysis for biomolecules. Our method utilizes 3D-RISM calculations to quickly obtain equilibrium solvent distributions without either necessity of simulation or limits of solvent sampling. Our analysis of these distributions extracts highest likelihood poses of solvent as well as localized entropies, enthalpies, and solvation free energies. We demonstrate our method on a structure of HIV-1 protease where excellent structural and thermodynamic data are available for comparison. Our results, obtained within minutes, show systematic agreement with available experimental data. Further, our results are in good agreement with established simulation-based solvent analysis methods. This method can be used not only for visual analysis of active site solvation but also for virtual screening methods and experimental refinement.

Mobil process converts methanol to high-quality synthetic gasoline

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wood, A.

1978-12-11

If production of gasoline from coal becomes commercially attractive in the United States, a process under development at the Mobil Research and Development Corp. may compete with better known coal liquefaction processes. Mobil process converts methanol to high-octane, unleaded gasoline; methanol can be produced commercially from coal. If gasoline is the desired product, the Mobil process offers strong technical and cost advantages over H-coal, Exxon donor solvent, solvent-refined coal, and Fischer--Tropsch processes. The cost analysis, contained in a report to the Dept. of Energy, concludes that the Mobil process produces more-expensive liquid products than any other liquefaction process except Fischer--Tropsch.more » But Mobil's process produces ready-to-use gasoline, while the others produce oils which require further expensive refining to yield gasoline. Disadvantages and advantages are discussed.« less

Purification of Carbon Nanotubes: Alternative Methods

NASA Technical Reports Server (NTRS)

Files, Bradley; Scott, Carl; Gorelik, Olga; Nikolaev, Pasha; Hulse, Lou; Arepalli, Sivaram

2000-01-01

Traditional carbon nanotube purification process involves nitric acid refluxing and cross flow filtration using surfactant TritonX. This is believed to result in damage to nanotubes and surfactant residue on nanotube surface. Alternative purification procedures involving solvent extraction, thermal zone refining and nitric acid refiuxing are used in the current study. The effect of duration and type of solvent to dissolve impurities including fullerenes and P ACs (polyaromatic compounds) are monitored by nuclear magnetic reasonance, high performance liquid chromatography, and thermogravimetric analysis. Thermal zone refining yielded sample areas rich in nanotubes as seen by scanning electric microscopy. Refluxing in boiling nitric acid seem to improve the nanotube content. Different procedural steps are needed to purify samples produced by laser process compared to arc process. These alternative methods of nanotube purification will be presented along with results from supporting analytical techniques.

Cbl Associates with Pyk2 and Src to Regulate Src Kinase Activity, αvβ3 Integrin-Mediated Signaling, Cell Adhesion, and Osteoclast Motility

PubMed Central

Sanjay, Archana; Houghton, Adam; Neff, Lynn; DiDomenico, Emilia; Bardelay, Chantal; Antoine, Evelyne; Levy, Joan; Gailit, James; Bowtell, David; Horne, William C.; Baron, Roland

2001-01-01

The signaling events downstream of integrins that regulate cell attachment and motility are only partially understood. Using osteoclasts and transfected 293 cells, we find that a molecular complex comprising Src, Pyk2, and Cbl functions to regulate cell adhesion and motility. The activation of integrin αvβ3 induces the [Ca2+]i-dependent phosphorylation of Pyk2 Y402, its association with Src SH2, Src activation, and the Src SH3-dependent recruitment and phosphorylation of c-Cbl. Furthermore, the PTB domain of Cbl is shown to bind to phosphorylated Tyr-416 in the activation loop of Src, the autophosphorylation site of Src, inhibiting Src kinase activity and integrin-mediated adhesion. Finally, we show that deletion of c Src or c-Cbl leads to a decrease in osteoclast migration. Thus, binding of αvβ3 integrin induces the formation of a Pyk2/Src/Cbl complex in which Cbl is a key regulator of Src kinase activity and of cell adhesion and migration. These findings may explain the osteopetrotic phenotype in the Src−/− mice. PMID:11149930

Phosphopeptide occupancy and photoaffinity cross-linking of the v-Src SH2 domain attenuates tyrosine kinase activity.

PubMed

Garcia, P; Shoelson, S E; Drew, J S; Miller, W T

1994-12-02

Phosphorylation of c-Src at carboxyl-terminal Tyr-527 suppresses tyrosine kinase activity and transforming potential, presumably by facilitating the intramolecular interaction of the C terminus of Src with its SH2 domain. In addition, it has been shown previously that occupancy of the c-Src SH2 domain with a phosphopeptide stimulates c-Src kinase catalytic activity. We have performed analogous studies with v-Src, the transforming protein from Rous sarcoma virus, which has extensive homology with c-Src. v-Src lacks an autoregulatory phosphorylation site, and its kinase domain is constitutively active. Phosphopeptides corresponding to the sequences surrounding c-Src Tyr-527 and a Tyr-Glu-Glu-Ile motif from the hamster polyoma virus middle T antigen inhibit tyrosine kinase activity of baculovirus-expressed v-Src 2- and 4-fold, respectively. To determine the mechanism of this regulation, the Tyr-527 phosphopeptide was substituted with the photoactive amino acid p-benzoylphenylalanine at the adjacent positions (N- and C-terminal) to phosphotyrosine. These peptides photoinactivate the v-Src tyrosine kinase 5-fold in a time- and concentration-dependent manner. Furthermore, the peptides cross-link an isolated Src SH2 domain with similar rates and specificity. These data indicate that occupancy of the v-Src SH2 domain induces a conformational change that is transmitted to the kinase domain and attenuates tyrosine kinase activity.

Immunohistochemical localization of steroid receptor coactivators in chondrosarcoma: an in vivo tissue microarray study.

PubMed

Li, Wei; Fu, Jingshu; Bian, Chen; Zhang, Jiqiang; Xie, Zhao

2014-12-01

Chondrosarcoma is the second most common type of primary bone malignancy following up osteosarcoma, characterized by resistance to conventional chemotherapeutic agents and radiation regimens. The p160 family members steroid receptor coactivator-1 and -3 (SRC-1 and SRC-3) have been implied in the regulation of cancer growth, migration, invasion, metastasis and chemotherapeutic resistance; but we still lack detailed information about the levels of SRCs in chondrosarcoma. In this study, expression of SRC-1 and SRC-3 in chondrosarcoma was examined by immunohistochemistry with tissue microarrays; the four score system (0, 1, 2 and 3) was used to evaluate the staining. The results showed that there were no gender-, site- or age-differences regarding the expression of SRC-1 or SRC-3 (p>0.05); organ (bone or cartilage) -differences were only detected for SRC-1 but not SRC-3 (p<0.05). Significant higher levels of SRC-1 and SRC-3 were detected in MDC and PDC when compared to WDC. Our study clearly demonstrated differentiation-dependant expression of SRC-1 and SRC-3 in chondrosarcoma, may be novel targets for the prognosis and/or treatment of chondrosarcoma, would have opened a new avenue and established foundation for studying chondrosarcoma. Copyright © 2014 Elsevier GmbH. All rights reserved.

Cytokinesis Failure Leading to Chromosome Instability in v-Src-Induced Oncogenesis.

PubMed

Nakayama, Yuji; Soeda, Shuhei; Ikeuchi, Masayoshi; Kakae, Keiko; Yamaguchi, Naoto

2017-04-12

v-Src, an oncogene found in Rous sarcoma virus, is a constitutively active variant of c-Src. Activation of Src is observed frequently in colorectal and breast cancers, and is critical in tumor progression through multiple processes. However, in some experimental conditions, v-Src causes growth suppression and apoptosis. In this review, we highlight recent progress in our understanding of cytokinesis failure and the attenuation of the tetraploidy checkpoint in v-Src-expressing cells. v-Src induces cell cycle changes-such as the accumulation of the 4N cell population-and increases the number of binucleated cells, which is accompanied by an excess number of centrosomes. Time-lapse analysis of v-Src-expressing cells showed that cytokinesis failure is caused by cleavage furrow regression. Microscopic analysis revealed that v-Src induces delocalization of cytokinesis regulators including Aurora B and Mklp1. Tetraploid cell formation is one of the causes of chromosome instability; however, tetraploid cells can be eliminated at the tetraploidy checkpoint. Interestingly, v-Src weakens the tetraploidy checkpoint by inhibiting the nuclear exclusion of the transcription coactivator YAP, which is downstream of the Hippo pathway and its nuclear exclusion is critical in the tetraploidy checkpoint. We also discuss the relationship between v-Src-induced chromosome instability and growth suppression in v-Src-induced oncogenesis.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Soeda, Shuhei; Nakayama, Yuji, E-mail: nakayama@mb.kyoto-phu.ac.jp; Department of Biochemistry and Molecular Biology, Kyoto Pharmaceutical University, 5 Nakauchi-cho, Misasagi, Yamashina-ku, Kyoto 607-8414

Src-family tyrosine kinases are aberrantly activated in cancers, and this activation is associated with malignant tumor progression. v-Src, encoded by the v-src transforming gene of the Rous sarcoma virus, is a mutant variant of the cellular proto-oncogene c-Src. Although investigations with temperature sensitive mutants of v-Src have shown that v-Src induces many oncogenic processes, the effects on cell division are unknown. Here, we show that v-Src inhibits cellular proliferation of HCT116, HeLa S3 and NIH3T3 cells. Flow cytometry analysis indicated that inducible expression of v-Src results in an accumulation of 4N cells. Time-lapse analysis revealed that binucleation is induced throughmore » the inhibition of cytokinesis, a final step of cell division. The localization of Mklp1, which is essential for cytokinesis, to the spindle midzone is inhibited in v-Src-expressing cells. Intriguingly, Aurora B, which regulates Mklp1 localization at the midzone, is delocalized from the spindle midzone and the midbody but not from the metaphase chromosomes upon v-Src expression. Mklp2, which is responsible for the relocation of Aurora B from the metaphase chromosomes to the spindle midzone, is also lost from the spindle midzone. These results suggest that v-Src inhibits cytokinesis through the delocalization of Mklp1 and Aurora B from the spindle midzone, resulting in binucleation. -- Highlights: • v-Src inhibits cell proliferation of HCT116, HeLa S3 and NIH3T3 cells. • v-Src induces binucleation together with cytokinesis failure. • v-Src causes delocalization of Mklp1, Aurora B and INCENP from the spindle midzone.« less

Src binds cortactin through an SH2 domain cystine-mediated linkage.

PubMed

Evans, Jason V; Ammer, Amanda G; Jett, John E; Bolcato, Chris A; Breaux, Jason C; Martin, Karen H; Culp, Mark V; Gannett, Peter M; Weed, Scott A

2012-12-15

Tyrosine-kinase-based signal transduction mediated by modular protein domains is critical for cellular function. The Src homology (SH)2 domain is an important conductor of intracellular signaling that binds to phosphorylated tyrosines on acceptor proteins, producing molecular complexes responsible for signal relay. Cortactin is a cytoskeletal protein and tyrosine kinase substrate that regulates actin-based motility through interactions with SH2-domain-containing proteins. The Src kinase SH2 domain mediates cortactin binding and tyrosine phosphorylation, but how Src interacts with cortactin is unknown. Here we demonstrate that Src binds cortactin through cystine bonding between Src C185 in the SH2 domain within the phosphotyrosine binding pocket and cortactin C112/246 in the cortactin repeats domain, independent of tyrosine phosphorylation. Interaction studies show that the presence of reducing agents ablates Src-cortactin binding, eliminates cortactin phosphorylation by Src, and prevents Src SH2 domain binding to cortactin. Tandem MS/MS sequencing demonstrates cystine bond formation between Src C185 and cortactin C112/246. Mutational studies indicate that an intact cystine binding interface is required for Src-mediated cortactin phosphorylation, cell migration, and pre-invadopodia formation. Our results identify a novel phosphotyrosine-independent binding mode between the Src SH2 domain and cortactin. Besides Src, one quarter of all SH2 domains contain cysteines at or near the analogous Src C185 position. This provides a potential alternative mechanism to tyrosine phosphorylation for cysteine-containing SH2 domains to bind cognate ligands that may be widespread in propagating signals regulating diverse cellular functions.

Src binds cortactin through an SH2 domain cystine-mediated linkage

PubMed Central

Evans, Jason V.; Ammer, Amanda G.; Jett, John E.; Bolcato, Chris A.; Breaux, Jason C.; Martin, Karen H.; Culp, Mark V.; Gannett, Peter M.; Weed, Scott A.

2012-01-01

Summary Tyrosine-kinase-based signal transduction mediated by modular protein domains is critical for cellular function. The Src homology (SH)2 domain is an important conductor of intracellular signaling that binds to phosphorylated tyrosines on acceptor proteins, producing molecular complexes responsible for signal relay. Cortactin is a cytoskeletal protein and tyrosine kinase substrate that regulates actin-based motility through interactions with SH2-domain-containing proteins. The Src kinase SH2 domain mediates cortactin binding and tyrosine phosphorylation, but how Src interacts with cortactin is unknown. Here we demonstrate that Src binds cortactin through cystine bonding between Src C185 in the SH2 domain within the phosphotyrosine binding pocket and cortactin C112/246 in the cortactin repeats domain, independent of tyrosine phosphorylation. Interaction studies show that the presence of reducing agents ablates Src-cortactin binding, eliminates cortactin phosphorylation by Src, and prevents Src SH2 domain binding to cortactin. Tandem MS/MS sequencing demonstrates cystine bond formation between Src C185 and cortactin C112/246. Mutational studies indicate that an intact cystine binding interface is required for Src-mediated cortactin phosphorylation, cell migration, and pre-invadopodia formation. Our results identify a novel phosphotyrosine-independent binding mode between the Src SH2 domain and cortactin. Besides Src, one quarter of all SH2 domains contain cysteines at or near the analogous Src C185 position. This provides a potential alternative mechanism to tyrosine phosphorylation for cysteine-containing SH2 domains to bind cognate ligands that may be widespread in propagating signals regulating diverse cellular functions. PMID:23097045

Literature survey of properties of synfuels derived from coal

NASA Technical Reports Server (NTRS)

Flores, F.

1982-01-01

A literature survey of the properties of synfuels for ground-based turbine applications is presented. The four major concepts for converting coal into liquid fuels (solvent extraction, catalytic liquefaction, pyrolysis, and indirect liquefaction), and the most important concepts for coal gasification (fixed bed, fluidized bed, entrained flow, and underground gasification) are described. Upgrading processes for coal derived liquid fuels are also described. Data presented for liquid fuels derived from various processes, including H-coal, synthoil, solvent refined coal, COED, donor solvent, zinc chloride hydrocracking, co-steam, and flash pyrolysis. Typical composition, and property data is also presented for low and medium-BTU gases derived from the various coal gasification processes.

Calcium-permeable α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid Receptors Trigger Neuronal Nitric-oxide Synthase Activation to Promote Nerve Cell Death in an Src Kinase-dependent Fashion*

PubMed Central

Socodato, Renato; Santiago, Felipe N.; Portugal, Camila C.; Domingues, Ana F.; Santiago, Ana R.; Relvas, João B.; Ambrósio, António F.; Paes-de-Carvalho, Roberto

2012-01-01

In the retina information decoding is dependent on excitatory neurotransmission and is critically modulated by AMPA glutamate receptors. The Src-tyrosine kinase has been implicated in modulating neurotransmission in CNS. Thus, our main goal was to correlate AMPA-mediated excitatory neurotransmission with the modulation of Src activity in retinal neurons. Cultured retinal cells were used to access the effects of AMPA stimulation on nitric oxide (NO) production and Src phosphorylation. 4-Amino-5-methylamino-2′,7′-difluorofluorescein diacetate fluorescence mainly determined NO production, and immunocytochemistry and Western blotting evaluated Src activation. AMPA receptors activation rapidly up-regulated Src phosphorylation at tyrosine 416 (stimulatory site) and down-regulated phosphotyrosine 527 (inhibitory site) in retinal cells, an effect mainly mediated by calcium-permeable AMPA receptors. Interestingly, experiments confirmed that neuronal NOS was activated in response to calcium-permeable AMPA receptor stimulation. Moreover, data suggest NO pathway as a key regulatory signaling in AMPA-induced Src activation in neurons but not in glial cells. The NO donor SNAP (S-nitroso-N-acetyl-dl-penicillamine) and a soluble guanylyl cyclase agonist (YC-1) mimicked AMPA effect in Src Tyr-416 phosphorylation, reinforcing that Src activation is indeed modulated by the NO pathway. Gain and loss-of-function data demonstrated that ERK is a downstream target of AMPA-induced Src activation and NO signaling. Furthermore, AMPA stimulated NO production in organotypic retinal cultures and increased Src activity in the in vivo retina. Additionally, AMPA-induced apoptotic retinal cell death was regulated by both NOS and Src activity. Because Src activity is pivotal in several CNS regions, the data presented herein highlight that Src modulation is a critical step in excitatory retinal cell death. PMID:22992730

Src kinase regulation by phosphorylation and dephosphorylation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Roskoski, Robert

2005-05-27

Src and Src-family protein-tyrosine kinases are regulatory proteins that play key roles in cell differentiation, motility, proliferation, and survival. The initially described phosphorylation sites of Src include an activating phosphotyrosine 416 that results from autophosphorylation, and an inhibiting phosphotyrosine 527 that results from phosphorylation by C-terminal Src kinase (Csk) and Csk homologous kinase. Dephosphorylation of phosphotyrosine 527 increases Src kinase activity. Candidate phosphotyrosine 527 phosphatases include cytoplasmic PTP1B, Shp1 and Shp2, and transmembrane enzymes include CD45, PTP{alpha}, PTP{epsilon}, and PTP{lambda}. Dephosphorylation of phosphotyrosine 416 decreases Src kinase activity. Thus far PTP-BL, the mouse homologue of human PTP-BAS, has been shownmore » to dephosphorylate phosphotyrosine 416 in a regulatory fashion. The platelet-derived growth factor receptor protein-tyrosine kinase mediates the phosphorylation of Src Tyr138; this phosphorylation has no direct effect on Src kinase activity. The platelet-derived growth factor receptor and the ErbB2/HER2 growth factor receptor protein-tyrosine kinases mediate the phosphorylation of Src Tyr213 and activation of Src kinase activity. Src kinase is also a substrate for protein-serine/threonine kinases including protein kinase C (Ser12), protein kinase A (Ser17), and CDK1/cdc2 (Thr34, Thr46, and Ser72). Of the three protein-serine/threonine kinases, only phosphorylation by CDK1/cdc2 has been demonstrated to increase Src kinase activity. Although considerable information on the phosphoprotein phosphatases that catalyze the hydrolysis of Src phosphotyrosine 527 is at hand, the nature of the phosphatases that mediate the hydrolysis of phosphotyrosine 138 and 213, and phosphoserine and phosphothreonine residues has not been determined.« less

miR-137 Targets p160 Steroid Receptor Coactivators SRC1, SRC2, and SRC3 and Inhibits Cell Proliferation

PubMed Central

Eedunuri, Vijay Kumar; Rajapakshe, Kimal; Fiskus, Warren; Geng, Chuandong; Chew, Sue Anne; Foley, Christopher; Shah, Shrijal S.; Shou, John; Mohamed, Junaith S.; O'Malley, Bert W.

2015-01-01

The p160 family of steroid receptor coactivators (SRCs) are pleiotropic transcription factor coactivators and “master regulators” of gene expression that promote cancer cell proliferation, survival, metabolism, migration, invasion, and metastasis. Cancers with high p160 SRC expression exhibit poor clinical outcomes and resistance to therapy, highlighting the SRCs as critical oncogenic drivers and, thus, therapeutic targets. microRNAs are important epigenetic regulators of protein expression. To examine the regulation of p160 SRCs by microRNAs, we used and combined 4 prediction algorithms to identify microRNAs that could target SRC1, SRC2, and SRC3 expression. For validation of these predictions, we assessed p160 SRC protein expression and cell viability after transfection of corresponding microRNA mimetics in breast cancer, uveal melanoma, and prostate cancer (PC) cell lines. Transfection of selected microRNA mimetics into breast cancer, uveal melanoma, and PC cells depleted SRC protein expression levels and exerted potent antiproliferative activity in these cell types. In particular, microRNA-137 (miR-137) depleted expression of SRC1, SRC2, and very potently, SRC3. The latter effect can be attributed to the presence of 3 miR-137 recognition sequences within the SRC3 3′-untranslated region. Using reverse phase protein array analysis, we identified a network of proteins, in addition to SRC3, that were modulated by miR-137 in PC cells. We also found that miR-137 and its host gene are epigenetically silenced in human cancer specimens and cell lines. These results support the development and testing of microRNA-based therapies (in particular based on restoring miR-137 levels) for targeting the oncogenic family of p160 SRCs in cancer. PMID:26066330

Calcium-permeable α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors trigger neuronal nitric-oxide synthase activation to promote nerve cell death in an Src kinase-dependent fashion.

PubMed

Socodato, Renato; Santiago, Felipe N; Portugal, Camila C; Domingues, Ana F; Santiago, Ana R; Relvas, João B; Ambrósio, António F; Paes-de-Carvalho, Roberto

2012-11-09

In the retina information decoding is dependent on excitatory neurotransmission and is critically modulated by AMPA glutamate receptors. The Src-tyrosine kinase has been implicated in modulating neurotransmission in CNS. Thus, our main goal was to correlate AMPA-mediated excitatory neurotransmission with the modulation of Src activity in retinal neurons. Cultured retinal cells were used to access the effects of AMPA stimulation on nitric oxide (NO) production and Src phosphorylation. 4-Amino-5-methylamino-2',7'-difluorofluorescein diacetate fluorescence mainly determined NO production, and immunocytochemistry and Western blotting evaluated Src activation. AMPA receptors activation rapidly up-regulated Src phosphorylation at tyrosine 416 (stimulatory site) and down-regulated phosphotyrosine 527 (inhibitory site) in retinal cells, an effect mainly mediated by calcium-permeable AMPA receptors. Interestingly, experiments confirmed that neuronal NOS was activated in response to calcium-permeable AMPA receptor stimulation. Moreover, data suggest NO pathway as a key regulatory signaling in AMPA-induced Src activation in neurons but not in glial cells. The NO donor SNAP (S-nitroso-N-acetyl-DL-penicillamine) and a soluble guanylyl cyclase agonist (YC-1) mimicked AMPA effect in Src Tyr-416 phosphorylation, reinforcing that Src activation is indeed modulated by the NO pathway. Gain and loss-of-function data demonstrated that ERK is a downstream target of AMPA-induced Src activation and NO signaling. Furthermore, AMPA stimulated NO production in organotypic retinal cultures and increased Src activity in the in vivo retina. Additionally, AMPA-induced apoptotic retinal cell death was regulated by both NOS and Src activity. Because Src activity is pivotal in several CNS regions, the data presented herein highlight that Src modulation is a critical step in excitatory retinal cell death.

Differential subcellular membrane recruitment of Src may specify its downstream signalling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Diesbach, Philippe de; Medts, Thierry; Carpentier, Sarah

2008-04-15

Most Src family members are diacylated and constitutively associate with membrane 'lipid rafts' that coordinate signalling. Whether the monoacylated Src, frequently hyperactive in carcinomas, also localizes at 'rafts' remains controversial. Using polarized MDCK cells expressing the thermosensitive v-Src/tsLA31 variant, we here addressed how Src tyrosine-kinase activation may impact on its (i) membrane recruitment, in particular to 'lipid rafts'; (ii) subcellular localization; and (iii) signalling. The kinetics of Src-kinase thermoactivation correlated with its recruitment from the cytosol to sedimentable membranes where Src largely resisted solubilisation by non-ionic detergents at 4 deg. C and floated into sucrose density gradients like caveolin-1 andmore » flotillin-2, i.e. 'lipid rafts'. By immunofluorescence, activated Src showed a dual localization, at apical endosomes/macropinosomes and at the apical plasma membrane. The plasma membrane Src pool did not colocalize with caveolin-1 and flotillin-2, but extensively overlapped GM1 labelling by cholera toxin. Severe ({approx} 70%) cholesterol extraction with methyl-{beta}-cyclodextrin (M{beta}CD) did not abolish 'rafts' floatation, but strongly decreased Src association with floating 'rafts' and abolished its localization at the apical plasma membrane. Src activation independently activated first the MAP-kinase - ERK1/2 pathway, then the PI3-kinase - Akt pathway. MAP-kinase - ERK1/2 activation was insensitive to M{beta}CD, which suppressed Akt phosphorylation and apical endocytosis induced by Src, both depending on the PI3-kinase pathway. We therefore suggest that activated Src is recruited at two membrane compartments, allowing differential signalling, first via ERK1/2 at 'non-raft' domains on endosomes, then via PI3-kinase-Akt on a distinct set of 'rafts' at the apical plasma membrane. Whether this model is applicable to c-Src remains to be examined.« less

Differential Prognostic Implications of Gastric Signet Ring Cell Carcinoma

PubMed Central

Chon, Hong Jae; Hyung, Woo Jin; Kim, Chan; Park, Sohee; Kim, Jie-Hyun; Park, Chan Hyuk; Ahn, Joong Bae; Kim, Hyunki; Chung, Hyun Cheol; Rha, Sun Young; Noh, Sung Hoon; Jeung, Hei-Cheul

2017-01-01

Objective: The aim of this study was to analyze the clinicopathologic characteristics and prognosis of signet ring cell carcinoma (SRC) according to disease status (early vs advanced gastric cancer) in gastric cancer patients. Background: The prognostic implication of gastric SRC remains a subject of debate. Methods: A retrospective analysis was performed using the clinical records of 7667 patients including 1646 SRC patients who underwent radical gastrectomy between 2001 and 2010. A further analysis was also performed after dividing patients into three groups according to histologic subtype: SRC, well-to-moderately differentiated (WMD), and poorly differentiated adenocarcinoma. Results: SRC patients have younger age distribution and female predominance compared with other histologic subtypes. Notably, the distribution of T stage of SRC patients was distinct, located in extremes (T1: 66.2% and T4: 20%). Moreover, the prognosis of SRC in early gastric cancer and advanced gastric cancer was contrasting. In early gastric cancer, SRC demonstrated more favorable prognosis than WMD after adjusting for age, sex, and stage. In contrast, SRC in advanced gastric cancer displayed worse prognosis than WMD. As stage increased, survival outcomes of SRC continued to worsen compared with WMD. Conclusions: Although conferring favorable prognosis in early stage, SRC has worse prognostic impact as disease progresses. The longstanding controversy of SRC on prognosis may result from disease status at presentation, which leads to differing prognosis compared with tubular adenocarinoma. PMID:27232252

Variant estrogen receptor-c-Src molecular interdependence and c-Src structural requirements for endothelial NO synthase activation.

PubMed

Li, Lei; Hisamoto, Koji; Kim, Kyung Hee; Haynes, M Page; Bauer, Philip M; Sanjay, Archana; Collinge, Mark; Baron, Roland; Sessa, William C; Bender, Jeffrey R

2007-10-16

Little is known about the tyrosine kinase c-Src's function in the systemic circulation, in particular its role in arterial responses to hormonal stimuli. In human aortic and venous endothelial cells, c-Src is indispensable for 17beta-estradiol (E2)-stimulated phosphatidylinositol 3-kinase/Akt/endothelial NO synthase (eNOS) pathway activation, a possible mechanism in E2-mediated vascular protection. Here we show that c-Src supports basal and E2-stimulated NO production and is required for E2-induced vasorelaxation in murine aortas. Only membrane c-Src is structurally and functionally involved in E2-induced eNOS activation. Independent of c-Src kinase activity, c-Src is associated with an N-terminally truncated estrogen receptor alpha variant (ER46) and eNOS in the plasma membrane through its "open" (substrate-accessible) conformation. In the presence of E2, c-Src kinase is activated by membrane ER46 and in turn phosphorylates ER46 for subsequent ER46 and c-Src membrane recruitment, the assembly of an eNOS-centered membrane macrocomplex, and membrane-initiated eNOS activation. Overall, these results provide insights into a critical role for the tyrosine kinase c-Src in estrogen-stimulated arterial responses, and in membrane-initiated rapid signal transduction, for which obligate complex assembly and localization require the c-Src substrate-accessible structure.

Focal adhesion kinase-dependent focal adhesion recruitment of SH2 domains directs SRC into focal adhesions to regulate cell adhesion and migration

PubMed Central

Wu, Jui-Chung; Chen, Yu-Chen; Kuo, Chih-Ting; Wenshin Yu, Helen; Chen, Yin-Quan; Chiou, Arthur; Kuo, Jean-Cheng

2015-01-01

Directed cell migration requires dynamical control of the protein complex within focal adhesions (FAs) and this control is regulated by signaling events involving tyrosine phosphorylation. We screened the SH2 domains present in tyrosine-specific kinases and phosphatases found within FAs, including SRC, SHP1 and SHP2, and examined whether these enzymes transiently target FAs via their SH2 domains. We found that the SRC_SH2 domain and the SHP2_N-SH2 domain are associated with FAs, but only the SRC_SH2 domain is able to be regulated by focal adhesion kinase (FAK). The FAK-dependent association of the SRC_SH2 domain is necessary and sufficient for SRC FA targeting. When the targeting of SRC into FAs is inhibited, there is significant suppression of SRC-mediated phosphorylation of paxillin and FAK; this results in an inhibition of FA formation and maturation and a reduction in cell migration. This study reveals an association between FAs and the SRC_SH2 domain as well as between FAs and the SHP2_N-SH2 domains. This supports the hypothesis that the FAK-regulated SRC_SH2 domain plays an important role in directing SRC into FAs and that this SRC-mediated FA signaling drives cell migration. PMID:26681405

Focal adhesion kinase-dependent focal adhesion recruitment of SH2 domains directs SRC into focal adhesions to regulate cell adhesion and migration.

PubMed

Wu, Jui-Chung; Chen, Yu-Chen; Kuo, Chih-Ting; Wenshin Yu, Helen; Chen, Yin-Quan; Chiou, Arthur; Kuo, Jean-Cheng

2015-12-18

Directed cell migration requires dynamical control of the protein complex within focal adhesions (FAs) and this control is regulated by signaling events involving tyrosine phosphorylation. We screened the SH2 domains present in tyrosine-specific kinases and phosphatases found within FAs, including SRC, SHP1 and SHP2, and examined whether these enzymes transiently target FAs via their SH2 domains. We found that the SRC_SH2 domain and the SHP2_N-SH2 domain are associated with FAs, but only the SRC_SH2 domain is able to be regulated by focal adhesion kinase (FAK). The FAK-dependent association of the SRC_SH2 domain is necessary and sufficient for SRC FA targeting. When the targeting of SRC into FAs is inhibited, there is significant suppression of SRC-mediated phosphorylation of paxillin and FAK; this results in an inhibition of FA formation and maturation and a reduction in cell migration. This study reveals an association between FAs and the SRC_SH2 domain as well as between FAs and the SHP2_N-SH2 domains. This supports the hypothesis that the FAK-regulated SRC_SH2 domain plays an important role in directing SRC into FAs and that this SRC-mediated FA signaling drives cell migration.

Endosomal-sorting complexes required for transport (ESCRT) pathway-dependent endosomal traffic regulates the localization of active Src at focal adhesions.

PubMed

Tu, Chun; Ortega-Cava, Cesar F; Winograd, Paul; Stanton, Marissa Jo; Reddi, Alagarsamy Lakku; Dodge, Ingrid; Arya, Ranjana; Dimri, Manjari; Clubb, Robert J; Naramura, Mayumi; Wagner, Kay-Uwe; Band, Vimla; Band, Hamid

2010-09-14

Active Src localization at focal adhesions (FAs) is essential for cell migration. How this pool is linked mechanistically to the large pool of Src at late endosomes (LEs)/lysosomes (LY) is not well understood. Here, we used inducible Tsg101 gene deletion, TSG101 knockdown, and dominant-negative VPS4 expression to demonstrate that the localization of activated cellular Src and viral Src at FAs requires the endosomal-sorting complexes required for transport (ESCRT) pathway. Tsg101 deletion also led to impaired Src-dependent activation of STAT3 and focal adhesion kinase and reduced cell migration. Impairment of the ESCRT pathway or Rab7 function led to the accumulation of active Src at aberrant LE/LY compartments followed by its loss. Analyses using fluorescence recovery after photo-bleaching show that dynamic mobility of Src in endosomes is ESCRT pathway-dependent. These results reveal a critical role for an ESCRT pathway-dependent LE/LY trafficking step in Src function by promoting localization of active Src to FAs.

SRC activates TAZ for intestinal tumorigenesis and regeneration.

PubMed

Byun, Mi Ran; Hwang, Jun-Ha; Kim, A Rum; Kim, Kyung Min; Park, Jung Il; Oh, Ho Taek; Hwang, Eun Sook; Hong, Jeong-Ho

2017-12-01

Proto-oncogene tyrosine-protein kinase Src (cSRC) is involved in colorectal cancer (CRC) development and damage-induced intestinal regeneration, although the cellular mechanisms involved are poorly understood. Here, we report that transcriptional coactivator with PDZ binding domain (TAZ) is activated by cSRC, regulating CRC cell proliferation and tumor formation, where cSRC overexpression increases TAZ expression in CRC cells. In contrast, knockdown of cSRC decreases TAZ expression. Additionally, direct phosphorylation of TAZ at Tyr316 by cSRC stimulates nuclear localization and facilitates transcriptional enhancer factor TEF-3 (TEAD4)-mediated transcription. However, a TAZ phosphorylation mutant significantly decreased cell proliferation, wound healing, colony forming, and tumor formation. In a CRC mouse model, Apc Min/+ , activated SRC expression was associated with increased TAZ expression in polyps and TAZ depletion decreased polyp formation. Moreover, intestinal TAZ knockout mice had intestinal regeneration defects following γ-irradiation. Finally, significant correspondence between SRC activation and TAZ overexpression was observed in CRC patients. These results suggest that TAZ is a critical factor for SRC-mediated intestinal tumor formation and regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

21 CFR 177.1900 - Urea-formaldehyde resins in molded articles.

Code of Federal Regulations, 2010 CFR

2010-04-01

... be mixed with refined wood pulp and the mixture may contain other optional adjuvant substances which... solvent not to exceed 0.5 milligram per square inch of food-contact surface as determined by the methods...

Protective equipment and player characteristics associated with the incidence of sport-related concussion in high school football players: a multifactorial prospective study.

PubMed

McGuine, Timothy A; Hetzel, Scott; McCrea, Michael; Brooks, M Alison

2014-10-01

The incidence of sport-related concussion (SRC) in high school football is well documented. However, limited prospective data are available regarding how player characteristics and protective equipment affect the incidence of SRC. To determine whether the type of protective equipment (helmet and mouth guard) and player characteristics affect the incidence of SRC in high school football players. Cohort study; Level of evidence, 2. Certified athletic trainers (ATs) at each high school recorded the type of helmet worn (brand, model, purchase year, and recondition status) by each player as well as information regarding players' demographics, type of mouth guard used, and history of SRC. The ATs also recorded the incidence and days lost from participation for each SRC. Incidence of SRC was compared for various helmets, type of mouth guard, history of SRC, and player demographics. A total of 2081 players (grades 9-12) enrolled during the 2012 and/or 2013 football seasons (2287 player-seasons) and participated in 134,437 football (practice or competition) exposures. Of these players, 206 (9%) sustained a total of 211 SRCs (1.56/1000 exposures). There was no difference in the incidence of SRC (number of helmets, % SRC [95% CI]) for players wearing Riddell (1171, 9.1% [7.6%-11.0%]), Schutt (680, 8.7% [6.7%-11.1%]), or Xenith (436, 9.2% [6.7%-12.4%]) helmets. Helmet age and recondition status did not affect the incidence of SRC. The rate of SRC (hazard ratio [HR]) was higher in players who wore a custom mouth guard (HR = 1.69 [95% CI, 1.20-2.37], P < .001) than in players who wore a generic mouth guard. The rate of SRC was also higher (HR = 1.96 [95% CI, 1.40-2.73], P < .001) in players who had sustained an SRC within the previous 12 months (15.1% of the 259 players [95% CI, 11.0%-20.1%]) than in players without a previous SRC (8.2% of the 2028 players [95% CI, 7.1%-9.5%]). Incidence of SRC was similar regardless of the helmet brand (manufacturer) worn by high school football players. Players who had sustained an SRC within the previous 12 months were more likely to sustain an SRC than were players without a history of SRC. Sports medicine providers who work with high school football players need to realize that factors other than the type of protective equipment worn affect the risk of SRC in high school players. © 2014 The Author(s).

The new program OPAL for molecular dynamics simulations and energy refinements of biological macromolecules.

PubMed

Luginbühl, P; Güntert, P; Billeter, M; Wüthrich, K

1996-09-01

A new program for molecular dynamics (MD) simulation and energy refinement of biological macromolecules, OPAL, is introduced. Combined with the supporting program TRAJEC for the analysis of MD trajectories, OPAL affords high efficiency and flexibility for work with different force fields, and offers a user-friendly interface and extensive trajectory analysis capabilities. Salient features are computational speeds of up to 1.5 GFlops on vector supercomputers such as the NEC SX-3, ellipsoidal boundaries to reduce the system size for studies in explicit solvents, and natural treatment of the hydrostatic pressure. Practical applications of OPAL are illustrated with MD simulations of pure water, energy minimization of the NMR structure of the mixed disulfide of a mutant E. coli glutaredoxin with glutathione in different solvent models, and MD simulations of a small protein, pheromone Er-2, using either instantaneous or time-averaged NMR restraints, or no restraints.

--No Title--

Science.gov Websites

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Sex Differences in High School Athletes' Knowledge of Sport-Related Concussion Symptoms and Reporting Behaviors.

PubMed

Wallace, Jessica; Covassin, Tracey; Beidler, Erica

2017-07-01

  Recent researchers have reported that athletes' knowledge of sport-related concussion (SRC) has increased but that athletes still lack knowledge of all the signs and symptoms of SRC. Understanding the signs and symptoms of SRC and the dangers of playing while symptomatic are critical to reporting behaviors in high school athletes.   To examine sex differences in knowledge of SRC symptoms and reasons for not reporting a suspected SRC to an authoritative figure in high school athletes.   Cross-sectional study.   Survey.   A total of 288 athletes across 7 sports (198 males [68.8%] and 90 females [31.2%]).   A validated knowledge-of-SRC survey consisted of demographic questions, a list of 21 signs and symptoms of SRC, and reasons why athletes would not report their SRC. The independent variable was sex. Athlete knowledge of SRC symptoms was assessed by having participants identify the signs and symptoms of SRC from a list of 21 symptoms. Knowledge scores were calculated by summing the number of correct answers; scores ranged from 0 to 21, with a score closer to 21 representing greater knowledge. Reporting-behavior questions asked athletes to choose reasons why they decided not to report any possible SRC signs and symptoms to an authoritative figure.   A sex difference in total SRC symptom knowledge was found (F 286 = 4.97, P = .03, d = 0.26). Female high school athletes had more total SRC symptom knowledge (mean ± standard deviation = 15.06 ± 2.63; 95% confidence interval = 14.54, 15.57) than males (14.36 ± 2.76; 95% confidence interval = 13.97, 14.74). Chi-square tests identified significant relationships between sex and 8 different reasons for not reporting an SRC.   High school males and females had similar SRC symptom knowledge; however, female athletes were more likely to report their concussive symptoms to an authoritative figure.

Preparative isolation of huperzines A and B from Huperzia serrata by displacement centrifugal partition chromatography.

PubMed

Toribio, Alix; Delannay, Eldra; Richard, Bernard; Plé, Karen; Zèches-Hanrot, Monique; Nuzillard, Jean-Marc; Renault, Jean-Hugues

2007-01-26

The pH-zone refining centrifugal partition chromatography technique was used to separate the two acetylcholinesterase inhibitors huperzines A and B from a crude alkaloid extract of the club moss Huperzia serrata. Complete co-elution of huperzines A and B was initially observed with the well-known methyl tert-butyl ether-acetonitrile-water (4:1:5, v/v/v) solvent system with triethylamine (8mM) as the displacer and methane sulfonic acid (6mM) as the retainer. An efficient biphasic system was designed on the basis of solvent association that provided selectivity in the elution mode: n-heptane/ethyl acetate/n-propanol/water (5:15:35:45, v/v/v/v). Lowering the bridge solvent content (n-propanol) of this system increased the polarity difference between the two phases thus adapting it to the pH-zone refining mode. Thus, the purification of these compounds was achieved using the biphasic system n-heptane/ethyl acetate/n-propanol/water (10:30:15:45, v/v/v/v) with triethylamine (8mM) as the displacer and methane sulfonic acid (6mM) as the retainer.

Iterative model building, structure refinement and density modification with the PHENIX AutoBuild wizard.

PubMed

Terwilliger, Thomas C; Grosse-Kunstleve, Ralf W; Afonine, Pavel V; Moriarty, Nigel W; Zwart, Peter H; Hung, Li Wei; Read, Randy J; Adams, Paul D

2008-01-01

The PHENIX AutoBuild wizard is a highly automated tool for iterative model building, structure refinement and density modification using RESOLVE model building, RESOLVE statistical density modification and phenix.refine structure refinement. Recent advances in the AutoBuild wizard and phenix.refine include automated detection and application of NCS from models as they are built, extensive model-completion algorithms and automated solvent-molecule picking. Model-completion algorithms in the AutoBuild wizard include loop building, crossovers between chains in different models of a structure and side-chain optimization. The AutoBuild wizard has been applied to a set of 48 structures at resolutions ranging from 1.1 to 3.2 A, resulting in a mean R factor of 0.24 and a mean free R factor of 0.29. The R factor of the final model is dependent on the quality of the starting electron density and is relatively independent of resolution.

Myristoylation of Src kinase mediates Src-induced and high-fat diet-accelerated prostate tumor progression in mice.

PubMed

Kim, Sungjin; Yang, Xiangkun; Li, Qianjin; Wu, Meng; Costyn, Leah; Beharry, Zanna; Bartlett, Michael G; Cai, Houjian

2017-11-10

Exogenous fatty acids provide substrates for energy production and biogenesis of the cytoplasmic membrane, but they also enhance cellular signaling during cancer cell proliferation. However, it remains controversial whether dietary fatty acids are correlated with tumor progression. In this study, we demonstrate that increased Src kinase activity is associated with high-fat diet-accelerated progression of prostate tumors and that Src kinases mediate this pathological process. Moreover, in the in vivo prostate regeneration assay, host SCID mice carrying Src(Y529F)-transduced regeneration tissues were fed a low-fat diet or a high-fat diet and treated with vehicle or dasatinib. The high-fat diet not only accelerated Src-induced prostate tumorigenesis in mice but also compromised the inhibitory effect of the anticancer drug dasatinib on Src kinase oncogenic potential in vivo We further show that myristoylation of Src kinase is essential to facilitate Src-induced and high-fat diet-accelerated tumor progression. Mechanistically, metabolism of exogenous myristic acid increased the biosynthesis of myristoyl CoA and myristoylated Src and promoted Src kinase-mediated oncogenic signaling in human cells. Of the fatty acids tested, only exogenous myristic acid contributed to increased intracellular myristoyl CoA levels. Our results suggest that targeting Src kinase myristoylation, which is required for Src kinase association at the cellular membrane, blocks dietary fat-accelerated tumorigenesis in vivo Our findings uncover the molecular basis of how the metabolism of myristic acid stimulates high-fat diet-mediated prostate tumor progression. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Cyr61 as mediator of Src signaling in triple negative breast cancer cells

PubMed Central

Molinari, Agnese; Wagner, Kay-Uwe; Losada, Jesús Pérez; Ciordia, Sergio; Albar, Juan Pablo; Martín-Pérez, Jorge

2015-01-01

SFKs are involved in tumorigenesis and metastasis. Here we analyzed c-Src contribution to initial steps of metastasis by tetracycline-dependent expression of a specific shRNA-c-Src, which suppressed c-Src mRNA and protein levels in metastatic MDA-MB-231 cells. c-Src suppression did not alter cell proliferation or survival, but it significantly reduced anchorage-independent growth. Concomitantly with diminished tyrosine-phosphorylation/activation of Fak, caveolin-1, paxillin and p130CAS, c-Src depletion also inhibited cellular migration, invasion and transendothelial migration. Quantitative proteomic analyses of the secretome showed that Cyr61 levels, which were detected in the exosomal fraction, were diminished upon shRNA-c-Src expression. In contrast, Cyr61 expression was unaltered inside cells. Cyr61 partially colocalized with cis-Golgi gp74 marker and with exosomal marker CD63, but c-Src depletion did not alter their cellular distribution. In SUM159PT cells, transient c-Src suppression also reduced secreted exosomal Cyr61 levels. Furthermore, conditional expression of a c-Src dominant negative mutant (SrcDN, c-Src-K295M/Y527F) in MDA-MB-231 and in SUM159PT diminished secreted Cyr61 as well. Cyr61 transient suppression in MDA-MB-231 inhibited invasion and transendothelial migration. Finally, in both MDA-MB-231 and SUM159PT, a neutralizing Cyr61 antibody restrained migration. Collectively, these results suggest that c-Src regulates secreted proteins, including the exosomal Cyr61, which are involved in modulating the metastatic potential of triple negative breast cancer cells. PMID:25980494

The Tyrosine Kinase Activity of c-Src Regulates Actin Dynamics and Organization of Podosomes in Osteoclasts

PubMed Central

Destaing, Olivier; Sanjay, Archana; Itzstein, Cecile; Horne, William C.; Toomre, Derek

2008-01-01

Podosomes are dynamic actin-rich structures composed of a dense F-actin core surrounded by a cloud of more diffuse F-actin. Src performs one or more unique functions in osteoclasts (OCLs), and podosome belts and bone resorption are impaired in the absence of Src. Using Src−/− OCLs, we investigated the specific functions of Src in the organization and dynamics of podosomes. We found that podosome number and the podosome-associated actin cloud were decreased in Src−/− OCLs. Videomicroscopy and fluorescence recovery after photobleaching analysis revealed that the life span of Src−/− podosomes was increased fourfold and that the rate of actin flux in the core was decreased by 40%. Thus, Src regulates the formation, structure, life span, and rate of actin polymerization in podosomes and in the actin cloud. Rescue of Src−/− OCLs with Src mutants showed that both the kinase activity and either the SH2 or the SH3 binding domain are required for Src to restore normal podosome organization and dynamics. Moreover, inhibition of Src family kinase activities in Src−/− OCLs by Src inhibitors or by expressing dominant-negative SrcK295M induced the formation of abnormal podosomes. Thus, Src is an essential regulator of podosome structure, dynamics and organization. PMID:17978100

76 FR 21404 - National Park Service Alaska Region's Subsistence Resource Commission (SRC) Program

Federal Register 2010, 2011, 2012, 2013, 2014

2011-04-15

... Resource Commission (SRC) program. SUMMARY: The Gates of the Arctic National Park SRC will meet to develop... to do so. Gates of the Arctic National Park SRC Meeting Date and Location: The Gates of the Arctic... weather or local circumstances. For Further Information on the Gates of the Arctic National Park SRC...

75 FR 51103 - Notice of Public Meetings for the National Park Service (NPS) Alaska Region's Subsistence...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-08-18

... SRC and Wrangell-St. Elias SRC plan to meet to develop and continue work on National Park Service (NPS... SRC Meeting Date and Location: The Lake Clark National Park SRC meeting will be held on Tuesday... Alaska Regional Office, at (907) 644- 3603. Aniakchak National Monument SRC Meeting Date and Location...

Simultaneous inhibition of aryl hydrocarbon receptor (AhR) and Src abolishes androgen receptor signaling.

PubMed

Ghotbaddini, Maryam; Cisse, Keyana; Carey, Alexis; Powell, Joann B

2017-01-01

Altered c-Src activity has been strongly implicated in the development, growth, progression, and metastasis of human cancers including prostate cancer. Src is known to regulate several biological functions of tumor cells, including proliferation. There are several Src inhibitors under evaluation for clinical effectiveness but have shown little activity in monotherapy trials of solid tumors. Combination studies are being explored by in vitro analysis and in clinical trials. Here we investigate the effect of simultaneous inhibition of the aryl hydrocarbon receptor (AhR) and Src on androgen receptor (AR) signaling in prostate cancer cells. AhR has also been reported to interact with the Src signaling pathway during prostate development. c-Src protein kinase is associated with the AhR complex in the cytosol and upon ligand binding to AhR, c-Src is activated and released from the complex. AhR has also been shown to regulate AR signaling which remains functionally important in the development and progression of prostate cancer. We provide evidence that co-inhibition of AhR and Src abolish AR activity. Evaluation of total protein and cellular fractions revealed decreased pAR expression and AR nuclear localization. Assays utilizing an androgen responsive element (ARE) and qRT-PCR analysis of AR genes revealed decreased AR promoter activity and transcriptional activity in the presence of both AhR and Src inhibitors. Furthermore, co-inhibition of AhR and Src reduced the growth of prostate cancer cells compared to individual treatments. Several studies have revealed that AhR and Src individually inhibit cellular proliferation. However, this study is the first to suggest simultaneous inhibition of AhR and Src to inhibit AR signaling and prostate cancer cell growth.

Differential effects of phosphotyrosine phosphatase expression on hormone-dependent and independent pp60c-src activity.

PubMed

Way, B A; Mooney, R A

1994-10-26

pp60c-src kinase activity can be increased by phosphotyrosine dephosphorylation or growth factor-dependent phosphorylation reactions. Expression of the transmembrane phosphotyrosine phosphatase (PTPase) CD45 has been shown to inhibit growth factor receptor signal transduction (Mooney, RA, Freund, GG, Way, BA and Bordwell, KL (1992) J Biol Chem 267, 23443-23446). Here it is shown that PTPase expression decreased platelet-derived growth factor (PDGF)-dependent activation of pp60c-src but failed to increase hormone independent (basal) pp60c-src activity. PDGF-dependent tyrosine phosphorylation of its receptor was reduced by approximately 60% in cells expressing the PTPase. In contrast, a change in phosphotyrosine content of pp60c-src was not detected in response to PDGF or in PTPase+ cells. PDGF increased the intrinsic tyrosine kinase activity of pp60c-src in both control and PTPase+ cells, but the effect was smaller in PTPase+ cells. In an in vitro assay, hormone-stimulated pp60c-src autophosphorylation from PTPase+ cells was decreased 64 +/- 22%, and substrate phosphorylation by pp60c-src was reduced 54 +/- 16% compared to controls. Hormone-independent pp60c-src kinase activity was unchanged by expression of the PTPase. pp60c-src was, however, an in vitro substrate for CD45, being dephosphorylated at both the regulatory (Tyr527) and kinase domain (Tyr416) residues. In addition, in vitro dephosphorylation by CD45 increased pp60c-src activity. These findings suggest that the PDGF receptor was an in vivo substrate of CD45 but pp60c-src was not. The lack of activation of pp60c-src in the presence of expressed PTPase may demonstrate the importance of compartmentalization and/or accessory proteins to PTPase-substrate interactions.

v-Src-induced nuclear localization of YAP is involved in multipolar spindle formation in tetraploid cells.

PubMed

Kakae, Keiko; Ikeuchi, Masayoshi; Kuga, Takahisa; Saito, Youhei; Yamaguchi, Naoto; Nakayama, Yuji

2017-01-01

The protein-tyrosine kinase, c-Src, is involved in a variety of signaling events, including cell division. We have reported that v-Src, which is a mutant variant of the cellular proto-oncogene, c-Src, causes delocalization of Aurora B kinase, resulting in a furrow regression in cytokinesis and the generation of multinucleated cells. However, the effect of v-Src on mitotic spindle formation is unknown. Here we show that v-Src-expressing HCT116 and NIH3T3 cells undergo abnormal cell division, in which cells separate into more than two cells. Upon v-Src expression, the proportion of multinucleated cells is increased in a time-dependent manner. Flow cytometry analysis revealed that v-Src increases the number of cells having a ≥4N DNA content. Microscopic analysis showed that v-Src induces the formation of multipolar spindles with excess centrosomes. These results suggest that v-Src induces multipolar spindle formation by generating multinucleated cells. Tetraploidy activates the tetraploidy checkpoint, leading to a cell cycle arrest of tetraploid cells at the G1 phase, in which the nuclear exclusion of the transcription co-activator YAP plays a critical role. In multinucleated cells that are induced by cytochalasin B and the Plk1 inhibitor, YAP is excluded from the nucleus. However, v-Src prevents this nuclear exclusion of YAP through a decrease in the phosphorylation of YAP at Ser127 in multinucleated cells. Furthermore, v-Src decreases the expression level of p53, which also plays a critical role in the cell cycle arrest of tetraploid cells. These results suggest that v-Src promotes abnormal spindle formation in at least two ways: generation of multinucleated cells and a weakening of the tetraploidy checkpoint. Copyright © 2016 Elsevier Inc. All rights reserved.

Protective Equipment and Player Characteristics Associated With the Incidence of Sport-Related Concussion in High School Football Players

PubMed Central

McGuine, Timothy A.; Hetzel, Scott; McCrea, Michael; Brooks, M. Alison

2015-01-01

Background The incidence of sport-related concussion (SRC) in high school football is well documented. However, limited prospective data are available regarding how player characteristics and protective equipment affect the incidence of SRC. Purpose To determine whether the type of protective equipment (helmet and mouth guard) and player characteristics affect the incidence of SRC in high school football players. Design Cohort study; Level of evidence, 2. Methods Certified athletic trainers (ATs) at each high school recorded the type of helmet worn (brand, model, purchase year, and recondition status) by each player as well as information regarding players’ demographics, type of mouth guard used, and history of SRC. The ATs also recorded the incidence and days lost from participation for each SRC. Incidence of SRC was compared for various helmets, type of mouth guard, history of SRC, and player demographics. Results A total of 2081 players (grades 9–12) enrolled during the 2012 and/or 2013 football seasons (2287 player-seasons) and participated in 134,437 football (practice or competition) exposures. Of these players, 206 (9%) sustained a total of 211 SRCs (1.56/1000 exposures). There was no difference in the incidence of SRC (number of helmets, % SRC [95% CI]) for players wearing Riddell (1171, 9.1% [7.6%–11.0%]), Schutt (680, 8.7% [6.7%–11.1%]), or Xenith (436, 9.2% [6.7%–12.4%]) helmets. Helmet age and recondition status did not affect the incidence of SRC. The rate of SRC (hazard ratio [HR]) was higher in players who wore a custom mouth guard (HR = 1.69 [95% CI, 1.20–2.37], P <.001) than in players who wore a generic mouth guard. The rate of SRC was also higher (HR = 1.96 [95% CI, 1.40–2.73], P <.001) in players who had sustained an SRC within the previous 12 months (15.1% of the 259 players [95% CI, 11.0%–20.1%]) than in players without a previous SRC (8.2% of the 2028 players [95% CI, 7.1%–9.5%]). Conclusion Incidence of SRC was similar regardless of the helmet brand (manufacturer) worn by high school football players. Players who had sustained an SRC within the previous 12 months were more likely to sustain an SRC than were players without a history of SRC. Sports medicine providers who work with high school football players need to realize that factors other than the type of protective equipment worn affect the risk of SRC in high school players. PMID:25060072

c-Src, Insulin-Like Growth Factor I Receptor, G-Protein-Coupled Receptor Kinases and Focal Adhesion Kinase are Enriched Into Prostate Cancer Cell Exosomes.

PubMed

DeRita, Rachel M; Zerlanko, Brad; Singh, Amrita; Lu, Huimin; Iozzo, Renato V; Benovic, Jeffrey L; Languino, Lucia R

2017-01-01

It is well known that Src tyrosine kinase, insulin-like growth factor 1 receptor (IGF-IR), and focal adhesion kinase (FAK) play important roles in prostate cancer (PrCa) development and progression. Src, which signals through FAK in response to integrin activation, has been implicated in many aspects of tumor biology, such as cell proliferation, metastasis, and angiogenesis. Furthermore, Src signaling is known to crosstalk with IGF-IR, which also promotes angiogenesis. In this study, we demonstrate that c-Src, IGF-IR, and FAK are packaged into exosomes (Exo), c-Src in particular being highly enriched in Exo from the androgen receptor (AR)-positive cell line C4-2B and AR-negative cell lines PC3 and DU145. Furthermore, we show that the active phosphorylated form of Src (Src pY416 ) is co-expressed in Exo with phosphorylated FAK (FAK pY861 ), a known target site of Src, which enhances proliferation and migration. We further demonstrate for the first time exosomal enrichment of G-protein-coupled receptor kinase (GRK) 5 and GRK6, both of which regulate Src and IGF-IR signaling and have been implicated in cancer. Finally, Src pY416 and c-Src are both expressed in Exo isolated from the plasma of prostate tumor-bearing TRAMP mice, and those same mice have higher levels of exosomal c-Src than their wild-type counterparts. In summary, we provide new evidence that active signaling molecules relevant to PrCa are enriched in Exo, and this suggests that the Src signaling network may provide useful biomarkers detectable by liquid biopsy, and may contribute to PrCa progression via Exo. J. Cell. Biochem. 118: 66-73, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Heterogeneity of signal transduction by Na-K-ATPase α-isoforms: role of Src interaction.

PubMed

Yu, Hui; Cui, Xiaoyu; Zhang, Jue; Xie, Joe X; Banerjee, Moumita; Pierre, Sandrine V; Xie, Zijian

2018-02-01

Of the four Na-K-ATPase α-isoforms, the ubiquitous α1 Na-K-ATPase possesses both ion transport and Src-dependent signaling functions. Mechanistically, we have identified two putative pairs of domain interactions between α1 Na-K-ATPase and Src that are critical for α1 signaling function. Our subsequent report that α2 Na-K-ATPase lacks these putative Src-binding sites and fails to carry on Src-dependent signaling further supported our proposed model of direct interaction between α1 Na-K-ATPase and Src but fell short of providing evidence for a causative role. This hypothesis was specifically tested here by introducing key residues of the two putative Src-interacting domains present on α1 but not α2 sequence into the α2 polypeptide, generating stable cell lines expressing this mutant, and comparing its signaling properties to those of α2-expressing cells. The mutant α2 was fully functional as a Na-K-ATPase. In contrast to wild-type α2, the mutant gained α1-like signaling function, capable of Src interaction and regulation. Consistently, the expression of mutant α2 redistributed Src into caveolin-1-enriched fractions and allowed ouabain to activate Src-mediated signaling cascades, unlike wild-type α2 cells. Finally, mutant α2 cells exhibited a growth phenotype similar to that of the α1 cells and proliferated much faster than wild-type α2 cells. These findings reveal the structural requirements for the Na-K-ATPase to function as a Src-dependent receptor and provide strong evidence of isoform-specific Src interaction involving the identified key amino acids. The sequences surrounding the putative Src-binding sites in α2 are highly conserved across species, suggesting that the lack of Src binding may play a physiologically important and isoform-specific role.

H-Ras Modulates N-Methyl-d-aspartate Receptor Function via Inhibition of Src Tyrosine Kinase Activity*

PubMed Central

Thornton, Claire; Yaka, Rami; Dinh, Son; Ron, Dorit

2005-01-01

Tyrosine phosphorylation of the NR2A and NR2B subunits of the N-methyl-d-aspartate (NMDA) receptor by Src protein-tyrosine kinases modulates receptor channel activity and is necessary for the induction of long term potentiation (LTP). Deletion of H-Ras increases both NR2 tyrosine phosphorylation and NMDA receptor-mediated hippocampal LTP. Here we investigated whether H-Ras regulates phosphorylation and function of the NMDA receptor via Src family protein-tyrosine kinases. We identified Src as a novel H-Ras binding partner. H-Ras bound to Src but not Fyn both in vitro and in brain via the Src kinase domain. Cotransfection of H-Ras and Src inhibited Src activity and decreased NR2A tyrosine phosphorylation. Treatment of rat brain slices with Tat-H-Ras depleted NR2A from the synaptic membrane, decreased endogenous Src activity and NR2A phosphorylation, and decreased the magnitude of hip-pocampal LTP. No change was observed for NR2B. We suggest that H-Ras negatively regulates Src phosphorylation of NR2A and retention of NR2A into the synaptic membrane leading to inhibition of NMDA receptor function. This mechanism is specific for Src and NR2A and has implications for studies in which regulation of NMDA receptor-mediated LTP is important, such as synaptic plasticity, learning, and memory and addiction. PMID:12695509

Targeting the yin and the yang: combined inhibition of the tyrosine kinase c-Src and the tyrosine phosphatase SHP-2 disrupts pancreatic cancer signaling and biology in vitro and tumor formation in vivo.

PubMed

Gomes, Evan G; Connelly, Sarah F; Summy, Justin M

2013-07-01

Although c-Src (Src) has emerged as a potential pancreatic cancer target in preclinical studies, Src inhibitors have not demonstrated a significant therapeutic benefit in clinical trials. The objective of these studies was to examine the effects of combining Src inhibition with inhibition of the protein tyrosine phosphatase SHP-2 in pancreatic cancer cells in vitro and in vivo. SHP-2 and Src functions were inhibited by siRNA or small molecule inhibitors. The effects of dual Src/SHP-2 functional inhibition were evaluated by Western blot analysis of downstream signaling pathways; cell biology assays to examine caspase activity, viability, adhesion, migration, and invasion in vitro; and an orthotopic nude mouse model to observe pancreatic tumor formation in vivo. Dual targeting of Src and SHP-2 induces an additive or supra-additive loss of phosphorylation of Akt and ERK-1/2 and corresponding increases in expression of apoptotic markers, relative to targeting either protein individually. Combinatorial inhibition of Src and SHP-2 significantly reduces viability, adhesion, migration, and invasion of pancreatic cancer cells in vitro and tumor formation in vivo, relative to individual Src/SHP-2 inhibition. These data suggest that the antitumor effects of Src inhibition in pancreatic cancer may be enhanced through simultaneous inhibition of SHP-2.

SRC-2-mediated coactivation of anti-tumorigenic target genes suppresses MYC-induced liver cancer

PubMed Central

Zhou, Xiaorong; Comerford, Sarah A.; York, Brian; O’Donnell, Kathryn A.

2017-01-01

Hepatocellular carcinoma (HCC) is the fifth most common solid tumor in the world and the third leading cause of cancer-associated deaths. A Sleeping Beauty-mediated transposon mutagenesis screen previously identified mutations that cooperate with MYC to accelerate liver tumorigenesis. This revealed a tumor suppressor role for Steroid Receptor Coactivator 2/Nuclear Receptor Coactivator 2 (Src-2/Ncoa2) in liver cancer. In contrast, SRC-2 promotes survival and metastasis in prostate cancer cells, suggesting a tissue-specific and context-dependent role for SRC-2 in tumorigenesis. To determine if genetic loss of SRC-2 is sufficient to accelerate MYC-mediated liver tumorigenesis, we bred Src-2-/- mice with a MYC-induced liver tumor model and observed a significant increase in liver tumor burden. RNA sequencing of liver tumors and in vivo chromatin immunoprecipitation assays revealed a set of direct target genes that are bound by SRC-2 and exhibit downregulated expression in Src-2-/- liver tumors. We demonstrate that activation of SHP (Small Heterodimer Partner), DKK4 (Dickkopf-4), and CADM4 (Cell Adhesion Molecule 4) by SRC-2 suppresses tumorigenesis in vitro and in vivo. These studies suggest that SRC-2 may exhibit oncogenic or tumor suppressor activity depending on the target genes and nuclear receptors that are expressed in distinct tissues and illuminate the mechanisms of tumor suppression by SRC-2 in liver. PMID:28273073

Acetylation within the N- and C-Terminal Domains of Src Regulates Distinct Roles of STAT3-Mediated Tumorigenesis.

PubMed

Huang, Chao; Zhang, Zhe; Chen, Lihan; Lee, Hank W; Ayrapetov, Marina K; Zhao, Ting C; Hao, Yimei; Gao, Jinsong; Yang, Chunzhang; Mehta, Gautam U; Zhuang, Zhengping; Zhang, Xiaoren; Hu, Guohong; Chin, Y Eugene

2018-06-01

Posttranslational modifications of mammalian c-Src N-terminal and C-terminal domains regulate distinct functions. Myristoylation of G 2 controls its cell membrane association and phosphorylation of Y419/Y527 controls its activation or inactivation, respectively. We provide evidence that Src-cell membrane association-dissociation and catalytic activation-inactivation are both regulated by acetylation. In EGF-treated cells, CREB binding protein (CBP) acetylates an N-terminal lysine cluster (K5, K7, and K9) of c-Src to promote dissociation from the cell membrane. CBP also acetylates the C-terminal K401, K423, and K427 of c-Src to activate intrinsic kinase activity for STAT3 recruitment and activation. N-terminal domain phosphorylation (Y14, Y45, and Y68) of STAT3 by c-Src activates transcriptionally active dimers of STAT3. Moreover, acetyl-Src translocates into nuclei, where it forms the Src-STAT3 enhanceosome for gene regulation and cancer cell proliferation. Thus, c-Src acetylation in the N-terminal and C-terminal domains play distinct roles in Src activity and regulation. Significance: CBP-mediated acetylation of lysine clusters in both the N-terminal and C-terminal regions of c-Src provides additional levels of control over STAT3 transcriptional activity. Cancer Res; 78(11); 2825-38. ©2018 AACR . ©2018 American Association for Cancer Research.

Functional diversity of Csk, Chk, and Src SH2 domains due to a single residue variation.

PubMed

Ayrapetov, Marina K; Nam, Nguyen Hai; Ye, Guofeng; Kumar, Anil; Parang, Keykavous; Sun, Gongqin

2005-07-08

The C-terminal Src kinase (Csk) family of protein tyrosine kinases contains two members: Csk and Csk homologous kinase (Chk). Both phosphorylate and inactivate Src family kinases. Recent reports suggest that the Src homology (SH) 2 domains of Csk and Chk may bind to different phosphoproteins, which provides a basis for different cellular functions for Csk and Chk. To verify and characterize such a functional divergence, we compared the binding properties of the Csk, Chk, and Src SH2 domains and investigated the structural basis for the functional divergence. First, the study demonstrated striking functional differences between the Csk and Chk SH2 domains and revealed functional similarities between the Chk and Src SH2 domains. Second, structural analysis and mutagenic studies revealed that the functional differences among the three SH2 domains were largely controlled by one residue, Glu127 in Csk, Ile167 in Chk, and Lys200 in Src. Mutating these residues in the Csk or Chk SH2 domain to the Src counterpart resulted in dramatic gain of function similar to Src SH2 domain, whereas mutating Lys200 in Src SH2 domain to Glu (the Csk counterpart) resulted in loss of Src SH2 function. Third, a single point mutation of E127K rendered Csk responsive to activation by a Src SH2 domain ligand. Finally, the optimal phosphopeptide sequence for the Chk SH2 domain was determined. These results provide a compelling explanation for the functional differences between two homologous protein tyrosine kinases and reveal a new structure-function relationship for the SH2 domains.

Polder maps: Improving OMIT maps by excluding bulk solvent

DOE PAGES

Liebschner, Dorothee; Afonine, Pavel V.; Moriarty, Nigel W.; ...

2017-02-01

The crystallographic maps that are routinely used during the structure-solution workflow are almost always model-biased because model information is used for their calculation. As these maps are also used to validate the atomic models that result from model building and refinement, this constitutes an immediate problem: anything added to the model will manifest itself in the map and thus hinder the validation. OMIT maps are a common tool to verify the presence of atoms in the model. The simplest way to compute an OMIT map is to exclude the atoms in question from the structure, update the corresponding structure factorsmore » and compute a residual map. It is then expected that if these atoms are present in the crystal structure, the electron density for the omitted atoms will be seen as positive features in this map. This, however, is complicated by the flat bulk-solvent model which is almost universally used in modern crystallographic refinement programs. This model postulates constant electron density at any voxel of the unit-cell volume that is not occupied by the atomic model. Consequently, if the density arising from the omitted atoms is weak then the bulk-solvent model may obscure it further. A possible solution to this problem is to prevent bulk solvent from entering the selected OMIT regions, which may improve the interpretative power of residual maps. This approach is called a polder (OMIT) map. Polder OMIT maps can be particularly useful for displaying weak densities of ligands, solvent molecules, side chains, alternative conformations and residues both in terminal regions and in loops. As a result, the tools described in this manuscript have been implemented and are available in PHENIX.« less

Src inhibitor reduces permeability without disturbing vascularization and prevents bone destruction in steroid-associated osteonecrotic lesions in rabbits.

PubMed

He, Yi-Xin; Liu, Jin; Guo, Baosheng; Wang, Yi-Xiang; Pan, Xiaohua; Li, Defang; Tang, Tao; Chen, Yang; Peng, Songlin; Bian, Zhaoxiang; Liang, Zicai; Zhang, Bao-Ting; Lu, Aiping; Zhang, Ge

2015-03-09

To examine the therapeutic effect of Src inhibitor on the VEGF mediating vascular hyperpermeability and bone destruction within steroid-associated osteonecrotic lesions in rabbits. Rabbits with high risk for progress to destructive repair in steroid-associated osteonecrosis were selected according to our published protocol. The selected rabbits were systemically administrated with either Anti-VEGF antibody (Anti-VEGF Group) or Src inhibitor (Src-Inhibition Group) or VEGF (VEGF-Supplement Group) or a combination of VEGF and Src inhibitor (Supplement &Inhibition Group) or control vehicle (Control Group) for 4 weeks. At 0, 2 and 4 weeks after administration, in vivo dynamic MRI, micro-CT based-angiography, histomorphometry and immunoblotting were employed to evaluate the vascular and skeletal events in different groups. The incidence of the destructive repair in the Anti-VEGF Group, Src-Inhibition Group and Supplement &Inhibition Group was all significantly lower than that in the Control Group. The angiogenesis was promoted in VEGF-Supplement Group, Src-Inhibition Group and Supplement &Inhibition Group, while the hyperpermeability was inhibited in Anti-VEGF Group, Src-Inhibition Group and Supplement &Inhibition Group. The trabecular structure was improved in Src-Inhibition Group and Supplement &Inhibition Group. Src inhibitor could reduce permeability without disturbing vascularization and prevent destructive repair in steroid-associated osteonecrosis.

Src promotes cutaneous wound healing by regulating MMP-2 through the ERK pathway.

PubMed

Wu, Xue; Yang, Longlong; Zheng, Zhao; Li, Zhenzhen; Shi, Jihong; Li, Yan; Han, Shichao; Gao, Jianxin; Tang, Chaowu; Su, Linlin; Hu, Dahai

2016-03-01

Wound healing is a highly orchestrated, multistep process, and delayed wound healing is a significant symptomatic clinical problem. Keratinocyte migration and re-epithelialization play the most important roles in wound healing, as they determine the rate of wound healing. In our previous study, we found that Src, one of the oldest proto‑oncogenes encoding a membrane-associated, non-receptor protein tyrosine kinase, promotes keratinocyte migration. We therefore hypothesized that Src promotes wound healing through enhanced keratinocyte migration. In order to test this hypothesis, vectors for overexpressing Src and small interfering RNAs (siRNAs) for silencing of Src were used in the present study. We found that the overexpression of Src accelerated keratinocyte migration in vitro and promoted wound healing in vivo without exerting a marked effect on cell proliferation. The extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling pathways play important roles in Src-accelerated keratinocyte migration. Further experiments demonstrated that Src induced the protein expression of matrix metalloproteinase-2 (MMP-2) and decreased the protein expression of E-cadherin. We suggest that ERK signaling is involved in the Src-mediated regulation of MMP-2 expression. The present study provided evidence that Src promotes keratinocyte migration and cutaneous wound healing, in which the regulation of MMP-2 through the ERK pathway plays an important role, and thus we also demonstrated a potential therapeutic role for Src in cutaneous wound healing.

Cigarette Smoke Activates the Proto-Oncogene c-Src to Promote Airway Inflammation and Lung Tissue Destruction

PubMed Central

Geraghty, Patrick; Hardigan, Andrew

2014-01-01

The diagnosis of chronic obstructive pulmonary disease (COPD) confers a 2-fold increased lung cancer risk even after adjusting for cigarette smoking, suggesting that common pathways are operative in both diseases. Although the role of the tyrosine kinase c-Src is established in lung cancer, less is known about its impact in other lung diseases, such as COPD. This study examined whether c-Src activation by cigarette smoke contributes to the pathogenesis of COPD. Cigarette smoke increased c-Src activity in human small airway epithelial (SAE) cells from healthy donors and in the lungs of exposed mice. Similarly, higher c-Src activation was measured in SAE cells from patients with COPD compared with healthy control subjects. In SAE cells, c-Src silencing or chemical inhibition prevented epidermal growth factor (EGF) receptor signaling in response to cigarette smoke but not EGF stimulation. Further studies showed that cigarette smoke acted through protein kinase C α to trigger c-Src to phosphorylate EGF receptor and thereby to induce mitogen-activated protein kinase responses in these cells. To further investigate the role of c-Src, A/J mice were orally administered the specific Src inhibitor AZD-0530 while they were exposed to cigarette smoke for 2 months. AZD-0530 treatment blocked c-Src activation, decreased macrophage influx, and prevented airspace enlargement in the lungs of cigarette smoke–exposed mice. Moreover, inhibiting Src deterred the cigarette smoke–mediated induction of matrix metalloproteinase-9 and -12 in alveolar macrophages and lung expression of cathepsin K, IL-17, TNF-α, MCP-1, and KC, all key factors in the pathogenesis of COPD. These results indicate that activation of the proto-oncogene c-Src by cigarette smoke promotes processes linked to the development of COPD. PMID:24111605

Adaptor protein GRB2 promotes Src tyrosine kinase activation and podosomal organization by protein-tyrosine phosphatase ϵ in osteoclasts.

PubMed

Levy-Apter, Einat; Finkelshtein, Eynat; Vemulapalli, Vidyasiri; Li, Shawn S-C; Bedford, Mark T; Elson, Ari

2014-12-26

The non-receptor isoform of protein-tyrosine phosphatase ϵ (cyt-PTPe) supports adhesion of bone-resorbing osteoclasts by activating Src downstream of integrins. Loss of cyt-PTPe reduces Src activity in osteoclasts, reduces resorption of mineralized matrix both in vivo and in cell culture, and induces mild osteopetrosis in young female PTPe KO mice. Activation of Src by cyt-PTPe is dependent upon this phosphatase undergoing phosphorylation at its C-terminal Tyr-638 by partially active Src. To understand how cyt-PTPe activates Src, we screened 73 Src homology 2 (SH2) domains for binding to Tyr(P)-638 of cyt-PTPe. The SH2 domain of GRB2 bound Tyr(P)-638 of cyt-PTPe most prominently, whereas the Src SH2 domain did not bind at all, suggesting that GRB2 may link PTPe with downstream molecules. Further studies indicated that GRB2 is required for activation of Src by cyt-PTPe in osteoclast-like cells (OCLs) in culture. Overexpression of GRB2 in OCLs increased activating phosphorylation of Src at Tyr-416 and of cyt-PTPe at Tyr-638; opposite results were obtained when GRB2 expression was reduced by shRNA or by gene inactivation. Phosphorylation of cyt-PTPe at Tyr-683 and its association with GRB2 are integrin-driven processes in OCLs, and cyt-PTPe undergoes autodephosphorylation at Tyr-683, thus limiting Src activation by integrins. Reduced GRB2 expression also reduced the ability of bone marrow precursors to differentiate into OCLs and reduced the fraction of OCLs in which podosomal adhesion structures assume organization typical of active, resorbing cells. We conclude that GRB2 physically links cyt-PTPe with Src and enables cyt-PTPe to activate Src downstream of activated integrins in OCLs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

A Kinase-Independent Function of c-Src Mediates p130Cas Phosphorylation at the Serine-639 Site in Pressure Overloaded Myocardium.

PubMed

Palanisamy, Arun P; Suryakumar, Geetha; Panneerselvam, Kavin; Willey, Christopher D; Kuppuswamy, Dhandapani

2015-12-01

Early work in pressure overloaded (PO) myocardium shows that integrins mediate focal adhesion complex formation by recruiting the adaptor protein p130Cas (Cas) and nonreceptor tyrosine kinase c-Src. To explore c-Src role in Cas-associated changes during PO, we used a feline right ventricular in vivo PO model and a three-dimensional (3D) collagen-embedded adult cardiomyocyte in vitro model that utilizes a Gly-Arg-Gly-Asp-Ser (RGD) peptide for integrin stimulation. Cas showed slow electrophoretic mobility (band-shifting), recruitment to the cytoskeleton, and tyrosine phosphorylation at 165, 249, and 410 sites in both 48 h PO myocardium and 1 h RGD-stimulated cardiomyocytes. Adenoviral mediated expression of kinase inactive (negative) c-Src mutant with intact scaffold domains (KN-Src) in cardiomyocytes did not block the RGD stimulated changes in Cas. Furthermore, expression of KN-Src or kinase active c-Src mutant with intact scaffold function (A-Src) in two-dimensionally (2D) cultured cardiomyocytes was sufficient to cause Cas band-shifting, although tyrosine phosphorylation required A-Src. These data indicate that c-Src's adaptor function, but not its kinase function, is required for a serine/threonine specific phosphorylation(s) responsible for Cas band-shifting. To explore this possibility, Chinese hamster ovary cells that stably express Cas were infected with either β-gal or KN-Src adenoviruses and used for Cas immunoprecipitation combined with mass spectrometry analysis. In the KN-Src expressing cells, Cas showed phosphorylation at the serine-639 (human numbering) site. A polyclonal antibody raised against phospho-serine-639 detected Cas phosphorylation in 24-48 h PO myocardium. Our studies indicate that c-Src's adaptor function mediates serine-639 phosphorylation of Cas during integrin activation in PO myocardium. © 2015 Wiley Periodicals, Inc.

Role of src-family kinases in hypoxic vasoconstriction of rat pulmonary artery

PubMed Central

Knock, Greg A.; Snetkov, Vladimir A.; Shaifta, Yasin; Drndarski, Svetlana; Ward, Jeremy P.T.; Aaronson, Philip I.

2008-01-01

Aims We investigated the role of src-family kinases (srcFKs) in hypoxic pulmonary vasoconstriction (HPV) and how this relates to Rho-kinase-mediated Ca2+ sensitization and changes in intracellular Ca2+ concentration ([Ca2+]i). Methods and results Intra-pulmonary arteries (IPAs) were obtained from male Wistar rats. HPV was induced in myograph-mounted IPAs. Auto-phosphorylation of srcFKs and phosphorylation of the regulatory subunit of myosin phosphatase (MYPT-1) and myosin light-chain (MLC20) in response to hypoxia were determined by western blotting. Translocation of Rho-kinase and effects of siRNA knockdown of src and fyn were examined in cultured pulmonary artery smooth muscle cells (PASMCs). [Ca2+]i was estimated in Fura-PE3-loaded IPA. HPV was inhibited by two blockers of srcFKs, SU6656 and PP2. Hypoxia enhanced phosphorylation of three srcFK proteins at Tyr-416 (60, 59, and 54 kDa, corresponding to src, fyn, and yes, respectively) and enhanced srcFK-dependent tyrosine phosphorylation of multiple target proteins. Hypoxia caused a complex, time-dependent enhancement of MYPT-1 and MLC20 phosphorylation, both in the absence and presence of pre-constriction. The sustained component of this enhancement was blocked by SU6656 and the Rho-kinase inhibitor Y27632. In PASMCs, hypoxia caused translocation of Rho-kinase from the nucleus to the cytoplasm, and this was prevented by anti-src siRNA and to a lesser extent by anti-fyn siRNA. The biphasic increases in [Ca2+]i that accompany HPV were also inhibited by PP2. Conclusion Hypoxia activates srcFKs and triggers protein tyrosine phosphorylation in IPA. Hypoxia-mediated Rho-kinase activation, Ca2+ sensitization, and [Ca2+]i responses are depressed by srcFK inhibitors and/or siRNA knockdown, suggesting a central role of srcFKs in HPV. PMID:18682436

Divergent modulation of Rho‐kinase and Ca2+ influx pathways by Src family kinases and focal adhesion kinase in airway smooth muscle

PubMed Central

Shaifta, Yasin; Irechukwu, Nneka; Prieto‐Lloret, Jesus; MacKay, Charles E; Marchon, Keisha A; Ward, Jeremy P T

2015-01-01

Background and Purpose The importance of tyrosine kinases in airway smooth muscle (ASM) contraction is not fully understood. The aim of this study was to investigate the role of Src‐family kinases (SrcFK) and focal adhesion kinase (FAK) in GPCR‐mediated ASM contraction and associated signalling events. Experimental Approach Contraction was recorded in intact or α‐toxin permeabilized rat bronchioles. Phosphorylation of SrcFK, FAK, myosin light‐chain‐20 (MLC20) and myosin phosphatase targeting subunit‐1 (MYPT‐1) was evaluated in cultured human ASM cells (hASMC). [Ca2+]i was evaluated in Fura‐2 loaded hASMC. Responses to carbachol (CCh) and bradykinin (BK) and the contribution of SrcFK and FAK to these responses were determined. Key Results Contractile responses in intact bronchioles were inhibited by antagonists of SrcFK, FAK and Rho‐kinase, while after α‐toxin permeabilization, they were sensitive to inhibition of SrcFK and Rho‐kinase, but not FAK. CCh and BK increased phosphorylation of MYPT‐1 and MLC20 and auto‐phosphorylation of SrcFK and FAK. MYPT‐1 phosphorylation was sensitive to inhibition of Rho‐kinase and SrcFK, but not FAK. Contraction induced by SR Ca2+ depletion and equivalent [Ca2+]i responses in hASMC were sensitive to inhibition of both SrcFK and FAK, while depolarization‐induced contraction was sensitive to FAK inhibition only. SrcFK auto‐phosphorylation was partially FAK‐dependent, while FAK auto‐phosphorylation was SrcFK‐independent. Conclusions and Implications SrcFK mediates Ca2+‐sensitization in ASM, while SrcFK and FAK together and individually influence multiple Ca2+ influx pathways. Tyrosine phosphorylation is therefore a key upstream signalling event in ASM contraction and may be a viable target for modulating ASM tone in respiratory disease. PMID:26294392

Mitochondrial events responsible for morphine's cardioprotection against ischemia/reperfusion injury

DOE Office of Scientific and Technical Information (OSTI.GOV)

He, Haiyan; Department of Pharmacology, Tianjin Medical University, Tianjin 300070; Huh, Jin

Morphine may induce cardioprotection by targeting mitochondria, but little is known about the exact mitochondrial events that mediate morphine's protection. We aimed to address the role of the mitochondrial Src tyrosine kinase in morphine's protection. Isolated rat hearts were subjected to 30 min ischemia and 2 h of reperfusion. Morphine was given before the onset of ischemia. Infarct size and troponin I release were measured to evaluate cardiac injury. Oxidative stress was evaluated by measuring mitochondrial protein carbonylation and mitochondrial ROS generation. HL-1 cells were subjected to simulated ischemia/reperfusion and LDH release and mitochondrial membrane potential (ΔΨm) were measured. Morphinemore » reduced infarct size as well as cardiac troponin I release which were aborted by the selective Src tyrosine kinase inhibitors PP2 and Src-I1. Morphine also attenuated LDH release and prevented a loss of ΔΨm at reperfusion in a Src tyrosine kinase dependent manner in HL-1 cells. However, morphine failed to reduce LDH release in HL-1 cells transfected with Src siRNA. Morphine increased mitochondrial Src phosphorylation at reperfusion and this was abrogated by PP2. Morphine attenuated mitochondrial protein carbonylation and mitochondrial superoxide generation at reperfusion through Src tyrosine kinase. The inhibitory effect of morphine on the mitochondrial complex I activity was reversed by PP2. These data suggest that morphine induces cardioprotection by preventing mitochondrial oxidative stress through mitochondrial Src tyrosine kinase. Inhibition of mitochondrial complex I at reperfusion by Src tyrosine kinase may account for the prevention of mitochondrial oxidative stress by morphine. - Highlights: • Morphine induced mito-Src phosphorylation and reduced infarct size in rat hearts. • Morphine failed to reduce I/R-induced LDH release in Src-silencing HL-1 cells. • Morphine prevented mitochondria damage caused by I/R through Src. • Morphine reduced mitochondrial ROS generation by inhibiting complex I via Src.« less

A Protein Scaffold Coordinates SRC-Mediated JNK Activation in Response to Metabolic Stress.

PubMed

Kant, Shashi; Standen, Claire L; Morel, Caroline; Jung, Dae Young; Kim, Jason K; Swat, Wojciech; Flavell, Richard A; Davis, Roger J

2017-09-19

Obesity is a major risk factor for the development of metabolic syndrome and type 2 diabetes. How obesity contributes to metabolic syndrome is unclear. Free fatty acid (FFA) activation of a non-receptor tyrosine kinase (SRC)-dependent cJun NH 2 -terminal kinase (JNK) signaling pathway is implicated in this process. However, the mechanism that mediates SRC-dependent JNK activation is unclear. Here, we identify a role for the scaffold protein JIP1 in SRC-dependent JNK activation. SRC phosphorylation of JIP1 creates phosphotyrosine interaction motifs that bind the SH2 domains of SRC and the guanine nucleotide exchange factor VAV. These interactions are required for SRC-induced activation of VAV and the subsequent engagement of a JIP1-tethered JNK signaling module. The JIP1 scaffold protein, therefore, plays a dual role in FFA signaling by coordinating upstream SRC functions together with downstream effector signaling by the JNK pathway. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Cooperative Atmosphere-Surface Exchange Study-1999.

NASA Astrophysics Data System (ADS)

Moeng, Chin-Hoh; Poulos, Gregory S.; Lemone, Margaret A.

2003-10-01

Surface-station, radiosonde, and Doppler minisodar data from the Cooperative Atmosphere-Surface Exchange Study-1997 (CASES-97) field project, collected in a 60-km-wide array in the lower Walnut River watershed (terrain variation 150 m) southeast of Wichita, Kansas, are used to study the relationship of the change of the 2-m potential temperature 2m with station elevation ze, 2m/ze ,ze to the ambient wind and thermal stratification /z ,z during fair-weather nights. As in many previous studies, predawn 2m varies linearly with ze, and ,ze ,z over a depth h that represents the maximum elevation range of the stations. Departures from the linear 2m-elevation relationship (,ze line) are related to vegetation (cool for vegetation, warm for bare ground), local terrain (drainage flows from nearby hills, although a causal relationship is not established), and the formation of a cold pool at lower elevations on some days.The near-surface flow and its evolution are functions of the Froude number Fr = S/(Nh), where S is the mean wind speed from the surface to h, and N is the corresponding Brunt-Väisälä frequency. The near-surface wind is coupled to the ambient flow for Fr = 3.3, based on where the straight line relating ,ze to ln Fr intersects the ln Fr axis. Under these conditions, 2m is constant horizontally even though ,z > 0, suggesting that near-surface air moves up- and downslope dry adiabatically. However, 2m cools (or warms) everywhere at the same rate. The lowest Froude numbers are associated with drainage flows, while intermediate values characterize regimes with intermediate behavior. The evolution of 2m horizontal variability σ through the night is also a function of the predawn Froude number. For the nights with the lowest Fr, the σ maximum occurs in the last 1-3 h before sunrise. For nights with Fr 3.3 (,ze 0) and for intermediate values, σ peaks 2-3 h after sunset. The standard deviations relative to the ,ze line reach their lowest values in the last hours of darkness. Thus, it is not surprising that the relationships of ,ze to Fr and ,z based on data through the night show more scatter, and ,ze 0.5,z in contrast to the predawn relationship. However, ,ze 0 for ln Fr = 3.7, a value similar to that just before sunrise.A heuristic Lagrangian parcel model is used to explain the horizontal uniformity of time-evolving 2m when the surface flow is coupled with the ambient wind, as well as both the linear variation of 2m with elevation and the time required to reach maximum values of σ under drainage-flow conditions.

Distribution and associations of intraocular pressure in 7- and 12-year-old Chinese children: The Anyang Childhood Eye Study.

PubMed

Li, Shuning; Li, Shi-Ming; Wang, Xiao-Lei; Kang, Meng-Tian; Liu, Luo-Ru; Li, He; Wei, Shi-Fei; Ran, An-Ran; Zhan, Siyan; Thomas, Ravi; Wang, Ningli

2017-01-01

To report the intraocular pressure (IOP) and its association with myopia and other factors in 7 and 12-year-old Chinese children. All children participating in the Anyang Childhood Eye Study underwent non-contact tonometry as well as measurement of central corneal thickness (CCT), axial length, cycloplegic auto-refraction, blood pressure, height and weight. A questionnaire was used to collect other relevant information. Univariable and multivariable analysis were performed to determine the associations of IOP. A total of 2760 7-year-old children (95.4%) and 2198 12-year-old children (97.0%) were included. The mean IOP was 13.5±3.1 mmHg in the younger cohort and 15.8±3.5 mmHg in older children (P<0.0001). On multivariable analysis, higher IOP in the younger cohort was associated with female gender (standardized regression coefficient [SRC], 0.11, P<0.0001), increasing central corneal thickness (SRC, 0.39, P<0.0001), myopia (SRC, 0.05, P = 0.03), deep anterior chamber (SRC, 0.07, P<0.01), smaller waist (SRC, 0.07, P<0.01) and increasing mean arterial pressure (SRC, 0.13, P<0.0001). In the older cohort, higher IOP was again associated with female gender (SRC, 0.16, P<0.0001), increasing central corneal thickness (SRC, 0.43, P<0.0001), deep anterior chamber (SRC, 0.09, P<0.01), higher body mass index (SRC, 0.07, P = 0.04) and with increasing mean arterial pressure (SRC, 0.09, P = 0.01), age at which reading commenced (SRC, 0.10, P<0.01) and birth method (SRC, 0.09, P = 0.01), but not with myopia (SRC, 0.09, P = 0.20). In Chinese children, higher IOP was associated with female gender, older age, thicker central cornea, deeper anterior chamber and higher mean arterial pressure. Higher body mass index, younger age at commencement of reading and being born of a caesarean section was also associated with higher IOP in adolescence.

Short-range order clustering in BCC Fe-Mn alloys induced by severe plastic deformation

NASA Astrophysics Data System (ADS)

Shabashov, V. A.; Kozlov, K. A.; Sagaradze, V. V.; Nikolaev, A. L.; Lyashkov, K. A.; Semyonkin, V. A.; Voronin, V. I.

2018-03-01

The effect of severe plastic deformation, namely, high-pressure torsion (HPT) at different temperatures and ball milling (BM) at different time intervals, has been investigated by means of Mössbauer spectroscopy in Fe100-xMnx (x = 4.1, 6.8, 9) alloys. Deformation affects the short-range clustering (SRC) in BCC lattice. Two processes occur: destruction of SRC by moving dislocations and enhancement of the SRC by migration of non-equilibrium defects. Destruction of SRC prevails during HPT at 80-293 K; whereas enhancement of SRC dominates at 473-573 K. BM starts enhancing the SRC formation at as low as 293 K due to local heating at impacts. The efficiency of HPT in terms of enhancing SRC increases with increasing temperature. The authors suppose that at low temperatures, a significant fraction of vacancies are excluded from enhancing SRC because of formation of mobile bi- and tri-vacancies having low efficiency of enhancing SRC as compared to that of mono vacancies. Milling of BCC Fe100-xMnx alloys stabilises the BCC phase with respect to α → γ transition at subsequent isothermal annealing because of a high degree of work hardening and formation of composition inhomogeneity.

[Factors of work environment and employment of workers in production of fuels and solvents at the oil refinery].

PubMed

Chebotarev, P A; Kharlashova, N V

2012-01-01

Factors of the industrial environment and labor activity of workers of manufacture propellants and solvents at the oil refining enterprise. Working conditions of workers at all installations of manufacture No 1 JSC "Naftan" of Novopolotsk of Byelorussia (production of fuels and solvents). Hygienic evaluation of working conditions of persons working in the production of fuels and solvents at the oil refinery. Sanitary description of the production with hygienic analysis of project design and technological documentation, qualitative and quantitative characteristics of conventional methods in the work environment and working process of employees in the workplace for the main modes of operation of the equipment. The working environment of refineries is influenced by a number of simultaneously acting factors, which have different material nature and characteristics of the action on the human body, the workers in production of fuels and solvents at the refinery, are exposed to a variety of hazardous and dangerous factors of production, a chemical factor is prevalent, of course.

Improved electron transport properties of n-type naphthalenediimide polymers through refined molecular ordering and orientation induced by processing solvents.

PubMed

An, Yujin; Long, Dang Xuan; Kim, Yiho; Noh, Yong-Young; Yang, Changduk

2016-05-14

To determine the role played by the choice of processing solvents in governing the photophysics, microstructure, and charge carrier transport in naphthalenediimide (NDI)-based polymers, we have prepared two new NDI-bithiophene (T2)- and NDI-thienothiophene (TTh)-containing polymers with hybrid siloxane pentyl chains (SiC5) (P(NDI2SiC5-T2) and P(NDI2SiC5-TTh)). Among the various processing solvents studied here, the films prepared using chloroform exhibited far better electron mobilities (0.16 ± 0.1-0.21 ± 0.05 cm(2) V(-1) s(-1)) than the corresponding samples prepared from different solvents, exceeding one order of magnitude higher, indicating the significant influence of the processing solvent on the charge transport. Upon thin-film analysis using atomic force microscopy and grazing incidence X-ray diffraction, we discovered that molecular ordering and orientation are affected by the choice of the processing solvent, which is responsible for the change in the transport characteristics of this class of polymers.

Src- and Fyn-dependent apical membrane trafficking events control endothelial lumen formation during vascular tube morphogenesis.

PubMed

Kim, Dae Joong; Norden, Pieter R; Salvador, Jocelynda; Barry, David M; Bowers, Stephanie L K; Cleaver, Ondine; Davis, George E

2017-01-01

Here we examine the question of how endothelial cells (ECs) develop their apical membrane surface domain during lumen and tube formation. We demonstrate marked apical membrane targeting of activated Src kinases to this apical domain during early and late stages of this process. Immunostaining for phosphotyrosine or phospho-Src reveals apical membrane staining in intracellular vacuoles initially. This is then followed by vacuole to vacuole fusion events to generate an apical luminal membrane, which is similarly decorated with activated phospho-Src kinases. Functional blockade of Src kinases completely blocks EC lumen and tube formation, whether this occurs during vasculogenic tube assembly or angiogenic sprouting events. Multiple Src kinases participate in this apical membrane formation process and siRNA suppression of Src, Fyn and Yes, but not Lyn, blocks EC lumen formation. We also demonstrate strong apical targeting of Src-GFP and Fyn-GFP fusion proteins and increasing their expression enhances lumen formation. Finally, we show that Src- and Fyn-associated vacuoles track and fuse along a subapically polarized microtubule cytoskeleton, which is highly acetylated. These vacuoles generate the apical luminal membrane in a stereotypically polarized, perinuclear position. Overall, our study identifies a critical role for Src kinases in creating and decorating the EC apical membrane surface during early and late stages of lumen and tube formation, a central event in the molecular control of vascular morphogenesis.

Association of spiritual/religious coping with depressive symptoms in high- and low-risk pregnant women.

PubMed

Vitorino, Luciano M; Chiaradia, Raíssa; Low, Gail; Cruz, Jonas Preposi; Pargament, Kenneth I; Lucchetti, Alessandra L G; Lucchetti, Giancarlo

2018-02-01

To investigate the role of spiritual/religious coping (SRC) on depressive symptoms in high- and low-risk pregnant women. Spiritual/religious coping is associated with physical and mental health outcomes. However, only few studies investigated the role of these strategies during pregnancy and whether low- and high-risk pregnant women have different coping mechanisms. This study is a cross-sectional comparative study. This study included a total of 160 pregnant women, 80 with low-risk pregnancy and 80 with high-risk pregnancy. The Beck Depression Inventory, the brief SRC scale and a structured questionnaire on sociodemographic and obstetric aspects were used. General linear model regression analysis was used to identify the factors associated with positive and negative SRC strategies in both groups of pregnant women. Positive SRC use was high, whereas negative SRC use was low in both groups. Although we found no difference in SRC strategies between the two groups, negative SRC was associated with depression in women with high-risk pregnancy, but not in those with low-risk pregnancy. Furthermore, positive SRC was not associated with depressive symptoms in both groups. Results showed that only the negative SRC strategies of Brazilian women with high-risk pregnancies were associated with worsened mental health outcomes. Healthcare professionals, obstetricians and nurse midwives should focus on the use of negative SRC strategies in their pregnant patients. © 2017 John Wiley & Sons Ltd.

Identification of a functional interaction between Kv4.3 channels and c-Src tyrosine kinase.

PubMed

Gomes, Pedro; Saito, Tomoaki; Del Corsso, Cris; Alioua, Abderrahmane; Eghbali, Mansoureh; Toro, Ligia; Stefani, Enrico

2008-10-01

Voltage-gated K(+) (Kv) channels are key determinants of cardiac and neuronal excitability. A substantial body of evidence has accumulated in support of a role for Src family tyrosine kinases in the regulation of Kv channels. In this study, we examined the possibility that c-Src tyrosine kinase participates in the modulation of the transient voltage-dependent K(+) channel Kv4.3. Supporting a mechanistic link between Kv4.3 and c-Src, confocal microscopy analysis of HEK293 cells stably transfected with Kv4.3 showed high degree of co-localization of the two proteins at the plasma membrane. Our results further demonstrate an association between Kv4.3 and c-Src by co-immunoprecipitation and GST pull-down assays, this interaction being mediated by the SH2 and SH3 domains of c-Src. Furthermore, we show that Kv4.3 is tyrosine phosphorylated under basal conditions. The functional relevance of the observed interaction between Kv4.3 and c-Src was established in patch-clamp experiments, where application of the Src inhibitor PP2 caused a decrease in Kv4.3 peak current amplitude, but not the inactive structural analogue PP3. Conversely, intracellular application of recombinant c-Src kinase or the protein tyrosine phosphatase inhibitor bpV(phen) increased Kv4.3 peak current amplitude. In conclusion, our findings provide evidence that c-Src-induced Kv4.3 channel activation involves their association in a macromolecular complex and suggest a role for c-Src-Kv4.3 pathway in regulating cardiac and neuronal excitability.

Steroid Receptor Coactivator 3 Contributes to Host Defense against Enteric Bacteria by Recruiting Neutrophils via Upregulation of CXCL2 Expression.

PubMed

Chen, Wenbo; Lu, Xuqiang; Chen, Yuan; Li, Ming; Mo, Pingli; Tong, Zhangwei; Wang, Wei; Wan, Wei; Su, Guoqiang; Xu, Jianming; Yu, Chundong

2017-02-15

Steroid receptor coactivator 3 (SRC-3) is a transcriptional coactivator that interacts with nuclear receptors and some other transcription factors to enhance their effects on target gene transcription. We reported previously that SRC-3-deficient (SRC-3 -/- ) mice are extremely susceptible to Escherichia coli-induced septic peritonitis as a result of uncontrolled inflammation and a defect in bacterial clearance. In this study, we observed significant upregulation of SRC-3 in colonic epithelial cells in response to Citrobacter rodentium infection. Based on these findings, we hypothesized that SRC-3 is involved in host defense against attaching and effacing bacterial infection. We compared the responses of SRC-3 -/- and wild-type mice to intestinal C. rodentium infection. We found that SRC-3 -/- mice exhibited delayed clearance of C. rodentium and more severe tissue pathology after oral infection with C. rodentium compared with wild-type mice. SRC-3 -/- mice expressed normal antimicrobial peptides in the colons but exhibited delayed recruitment of neutrophils into the colonic mucosa. Accordingly, SRC-3 -/- mice showed a delayed induction of CXCL2 and CXCL5 in colonic epithelial cells, which are responsible for neutrophil recruitment. At the molecular level, we found that SRC-3 can activate the NF-κB signaling pathway to promote CXCL2 expression at the transcriptional level. Collectively, we show that SRC-3 contributes to host defense against enteric bacteria, at least in part via upregulating CXCL2 expression to recruit neutrophils. Copyright © 2017 by The American Association of Immunologists, Inc.

Global Impact of Oncogenic Src on a Phosphotyrosine Proteome

PubMed Central

Luo, Weifeng; Slebos, Robbert J.; Hill, Salisha; Li, Ming; Brábek, Jan; Amanchy, Ramars; Chaerkady, Raghothama; Pandey, Akhilesh; Ham, Amy-Joan L.; Hanks, Steven K.

2008-01-01

Elevated activity of Src, the first characterized protein-tyrosine kinase, is associated with progression of many human cancers, and Src has attracted interest as a therapeutic target. Src is known to act in various receptor signaling systems to impact cell behavior, yet it remains likely that the spectrum of Src protein substrates relevant to cancer is incompletely understood. To better understand the cellular impact of deregulated Src kinase activity, we extensively applied a mass spectrometry shotgun phosphotyrosine (pTyr) proteomics strategy to obtain global pTyr profiles of Src-transformed mouse fibroblasts as well as their nontransformed counterparts. A total of 867 peptides representing 563 distinct pTyr sites on 374 different proteins were identified from the Src-transformed cells, while 514 peptides representing 275 pTyr sites on 167 proteins were identified from nontransformed cells. Distinct characteristics of the two profiles were revealed by spectral counting, indicative of pTyr site relative abundance, and by complementary quantitative analysis using stable isotope labeling with amino acids in cell culture (SILAC). While both pTyr profiles are replete with sites on signaling and adhesion/cytoskeletal regulatory proteins, the Src-transformed profile is more diverse with enrichment in sites on metabolic enzymes and RNA and protein synthesis and processing machinery. Forty-three pTyr sites (32 proteins) are predicted as major biologically relevant Src targets on the basis of frequent identification in both cell populations. This select group, of particular interest as diagnostic biomarkers, includes well-established Src sites on signaling/adhesion/cytoskeletal proteins, but also uncharacterized sites of potential relevance to the transformed cell phenotype. PMID:18563927

Novel Bioluminescent Activatable Reporter for Src Tyrosine Kinase Activity in Living Mice

PubMed Central

Leng, Weibing; Li, Dezhi; Chen, Liang; Xia, Hongwei; Tang, Qiulin; Chen, Baoqin; Gong, Qiyong; Gao, Fabao; Bi, Feng

2016-01-01

Aberrant activation of the Src kinase is implicated in the development of a variety of human malignancies. However, it is almost impossible to monitor Src activity in an in vivo setting with current biochemical techniques. To facilitate the noninvasive investigation of the activity of Src kinase both in vitro and in vivo, we developed a genetically engineered, activatable bioluminescent reporter using split-luciferase complementation. The bioluminescence of this reporter can be used as a surrogate for Src activity in real time. This hybrid luciferase reporter was constructed by sandwiching a Src-dependent conformationally responsive unit (SH2 domain-Srcpep) between the split luciferase fragments. The complementation bioluminescence of this reporter was dependent on the Src activity status. In our study, Src kinase activity in cultured cells and tumor xenografts was monitored quantitatively and dynamically in response to clinical small-molecular kinase inhibitors, dasatinib and saracatinib. This system was also applied for high-throughput screening of Src inhibitors against a kinase inhibitor library in living cells. These results provide unique insights into drug development and pharmacokinetics/phoarmocodynamics of therapeutic drugs targeting Src signaling pathway enabling the optimization of drug administration schedules for maximum benefit. Using both Firefly and Renilla luciferase imaging, we have successfully monitored Src tyrosine kinase activity and Akt serine/threonine kinase activity concurrently in one tumor xenograft. This dual luciferase reporter imaging system will be helpful in exploring the complex signaling networks in vivo. The strategies reported here can also be extended to study and image other important kinases and the cross-talks among them. PMID:26941850

A comparison of choline:urea and choline:oxalic acid deep eutectic solvents at 338 K

NASA Astrophysics Data System (ADS)

Gilmore, Mark; Moura, Leila M.; Turner, Adam H.; Swadźba-Kwaśny, Małgorzata; Callear, Samantha K.; McCune, Jade A.; Scherman, Oren A.; Holbrey, John D.

2018-05-01

1:2 choline chloride:urea and 1:1 choline chloride:oxalic acid deep eutectic solvents are compared at 338 K using liquid-phase neutron diffraction with H/D isotopic substitution to obtain differential neutron scattering cross sections and fitting of models to the experimental data using Empirical Potential Structure Refinement. In comparison to the previously reported study of choline chloride:urea at 303 K, we observed significant weakening and lengthening of choline-OH⋯Cl- and choline-OH⋯hydrogen-bond acceptor correlations.

Single-crystal growth, structure refinement and the properties of Bis(glycine) Strontium Chloride

NASA Astrophysics Data System (ADS)

Balaji, S. R.; Balu, T.; Rajasekaran, T. R.

2018-02-01

Single crystals of Bis (glycine) Strontium Chloride (BGSC) were grown by means of slow evaporation process by using analar grade Glycine and Strontium Chloride Hexahydrate as a parent compound from its aqueous solution at room temperature. The final chemical composition, [{{Sr}}{({{{C}}}2{{{H}}}5{{{NO}}}2)}2{{{Cl}}}2].{{{H}}}4{{{O}}}3+{{{H}}}8{{{O}}}3, formed were metallic light colorless block, about the size of 28 mm × 9 mm × 8 mm. A single-crystal x-ray diffraction study revealed an ordered superstructure with orthorhombic symmetry that could be assigned to the space group Pbcn. The structure in BGSC, revealed in the electron density distribution was analyzed by the direct methods (SHELXS-2014) and refined by least squares full matrix method (SHELXL-2014). The crystal structure, including anisotropic atomic displacement parameters for each atom and isotropic atomic displacement parameters for hydrogen atom, was refined to R1 = 0.0395, wR2 = 0.0776 using 1097 independent reflections. The FTIR spectrum of BGSC confirms the protonation of amino groups and the different molecular groups present in BGSC vibrate in different modes. Reverse Indentation Size Effect (RISE) was revealed in BGSC in the micro-hardness analysis using Vicker’s micro-hardness analysis. DTA and DSC results ruled out the possibility of structural change independent of mass change. The AFM studies shows fine nano size fiber like structure of the grown crystals.

Bayesian refinement of protein structures and ensembles against SAXS data using molecular dynamics

PubMed Central

Shevchuk, Roman; Hub, Jochen S.

2017-01-01

Small-angle X-ray scattering is an increasingly popular technique used to detect protein structures and ensembles in solution. However, the refinement of structures and ensembles against SAXS data is often ambiguous due to the low information content of SAXS data, unknown systematic errors, and unknown scattering contributions from the solvent. We offer a solution to such problems by combining Bayesian inference with all-atom molecular dynamics simulations and explicit-solvent SAXS calculations. The Bayesian formulation correctly weights the SAXS data versus prior physical knowledge, it quantifies the precision or ambiguity of fitted structures and ensembles, and it accounts for unknown systematic errors due to poor buffer matching. The method further provides a probabilistic criterion for identifying the number of states required to explain the SAXS data. The method is validated by refining ensembles of a periplasmic binding protein against calculated SAXS curves. Subsequently, we derive the solution ensembles of the eukaryotic chaperone heat shock protein 90 (Hsp90) against experimental SAXS data. We find that the SAXS data of the apo state of Hsp90 is compatible with a single wide-open conformation, whereas the SAXS data of Hsp90 bound to ATP or to an ATP-analogue strongly suggest heterogenous ensembles of a closed and a wide-open state. PMID:29045407

Iterative model building, structure refinement and density modification with the PHENIX AutoBuild wizard

PubMed Central

Terwilliger, Thomas C.; Grosse-Kunstleve, Ralf W.; Afonine, Pavel V.; Moriarty, Nigel W.; Zwart, Peter H.; Hung, Li-Wei; Read, Randy J.; Adams, Paul D.

2008-01-01

The PHENIX AutoBuild wizard is a highly automated tool for iterative model building, structure refinement and density modification using RESOLVE model building, RESOLVE statistical density modification and phenix.refine structure refinement. Recent advances in the AutoBuild wizard and phenix.refine include automated detection and application of NCS from models as they are built, extensive model-completion algorithms and automated solvent-molecule picking. Model-completion algorithms in the AutoBuild wizard include loop building, crossovers between chains in different models of a structure and side-chain optimization. The AutoBuild wizard has been applied to a set of 48 structures at resolutions ranging from 1.1 to 3.2 Å, resulting in a mean R factor of 0.24 and a mean free R factor of 0.29. The R factor of the final model is dependent on the quality of the starting electron density and is relatively independent of resolution. PMID:18094468

Significance of ERa and c-Src Interaction in the Progression of Hormone Independent Breast Cancer

DTIC Science & Technology

2005-12-01

defects in estrogen signaling [1]. Because of global defects in estrogen signaling observed in these c-Src deficient mice, we have recently generated...1998). Interestingly, the region of the kinase domain of ErbB-2 that correlates with c-Src association, referred to as TK2 (Segatto et al., 1991...ductive organs that are dependent on ERa in c-Src- deficient mice. We show that the loss of the c-Src tyrosine kinase correlates with defects in ductal

Mesoscale Polymer Dissolution Probed by Raman Spectroscopy and Molecular Simulations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chang, Tsun-Mei; Xantheas, Sotiris S.; Vasdekis, Andreas E.

2016-10-13

The diffusion of various solvents into a polystyrene (PS) matrix was probed experimentally by monitoring the temporal profiles of the Raman spectra and theoretically from molecular dynamics (MD) simulations of the binary system. The simulation results assist in providing a fundamental, molecular level connection between the mixing/dissolution processes and the difference = solvent – PS in the values of the Hildebrand parameter () between the two components of the binary systems: solvents having similar values of with PS (small ) exhibit fast diffusion into the polymer matrix, whereas the diffusion slows down considerably when the ’s are different (large ).more » To this end, the Hildebrand parameter was identified as a useful descriptor that governs the process of mixing in polymer – solvent binary systems. The experiments also provide insight into further refinements of the models specific to non-Fickian diffusion phenomena that need to be used in the simulations.« less

EG-1 interacts with c-Src and activates its signaling pathway.

PubMed

Lu, Ming; Zhang, Liping; Sartippour, Maryam R; Norris, Andrew J; Brooks, Mai N

2006-10-01

EG-1 is significantly elevated in breast, colorectal, and prostate cancers. Overexpression of EG-1 stimulates cellular proliferation, and targeted inhibition blocks mouse xenograft tumor growth. To further clarify the function of EG-1, we investigated its role in c-Src activation. We observed that EG-1 overexpression results in activation of c-Src, but found no evidence that EG-1 is a direct Src substrate. EG-1 also binds to other members of the Src family. Furthermore, EG-1 shows interaction with multiple other SH3- and WW-containing molecules involved in various signaling pathways. These observations suggest that EG-1 may be involved in signaling pathways including c-Src activation.

40 CFR 52.2239 - Original Identification of plan section.

Code of Federal Regulations, 2012 CFR

2012-07-01

... Carbon Monoxide Reasonable Further Progress (RFP) curve adopted on May 8, 1985. (ii) Other material. (A...) Permit for battery receiving and breaking operation for Refined Metals Corporation which became effective... Efficiency. (35) Rule 1200-3-18-.46 Test Method for Determination of Solvent...

40 CFR 52.2239 - Original Identification of plan section.

Code of Federal Regulations, 2013 CFR

2013-07-01

... Carbon Monoxide Reasonable Further Progress (RFP) curve adopted on May 8, 1985. (ii) Other material. (A...) Permit for battery receiving and breaking operation for Refined Metals Corporation which became effective... Efficiency. (35) Rule 1200-3-18-.46 Test Method for Determination of Solvent...

AIR TOXICS ASSESSMENT REFINEMENT IN RAPCA'S JURISDICTION - DAYTON, OH AREA

EPA Science Inventory

RAPCA has receive two grants to conduct this project. As part of the original project, RAPCA has improved and expanded their point source inventory by converting the following area sources to point sources: dry cleaners, gasoline throughput processes and halogenated solvent clea...

76 FR 3653 - Alaska Region's Subsistence Resource Commission (SRC) Program; Public Meeting

Federal Register 2010, 2011, 2012, 2013, 2014

2011-01-20

... subsistence management issues. The NPS SRC program is authorized under Title VIII, Section 808 of the Alaska...: 1. Call to order. 2. SRC Roll Call and Confirmation of Quorum. 3. Welcome and Introductions. 4.... c. Resource Management Program Update. 14. Public and other Agency Comments. 15. SRC Work Session...

OH REACTION KINETICS OF GAS-PHASE A- AND G-HEXACHLOROCYCLOHEXANE AND HEXACHLOROBENZENE. (R825377)

EPA Science Inventory

Rate constants for the gas-phase reactions of the hydroxyl
radical (OH) with - and -hexachlorocyclohexane (-
and 78 FR 51207 - Kobuk Valley National Park Subsistence Resource Commission (SRC) and the Denali National Park SRC...

Federal Register 2010, 2011, 2012, 2013, 2014

2013-08-20

... DEPARTMENT OF THE INTERIOR National Park Service [NPS-AKR-DENA-KOVA-DTS-13608; PPAKAKROR4; PPMPRLE1Y.LS0000] Kobuk Valley National Park Subsistence Resource Commission (SRC) and the Denali National Park SRC; Meetings AGENCY: National Park Service, Interior. ACTION: Meeting notice. SUMMARY: As...

76 FR 57763 - Alaska Region's Subsistence Resource Commission (SRC) Program

Federal Register 2010, 2011, 2012, 2013, 2014

2011-09-16

...) program. SUMMARY: The Gates of the Arctic National Park SRC will meet to develop and continue work on NPS... changed based on inclement weather or exceptional circumstances. Gates of the Arctic National Park SRC Meeting Dates and Location: The Gates of the Arctic National Park SRC will meet at Sophie Station Hotel...

Dynamic activation of Src induced by low-power laser irradiation in living cells mediated by reactive oxygen species

NASA Astrophysics Data System (ADS)

Zhang, Juntao; Gao, Xuejuan; Xing, Da; Liu, Lei

2007-11-01

Low-power laser irradiation (LPLI) leads to photochemical reaction and then activates intracellular several signaling pathway. Reactive oxygen species (ROS) are considered to be the primary messengers produced by LPLI. Here, we studied the signaling pathway mediated by ROS upon the stimulation of LPLI. Src tyrosine kinases are well-known targets of ROS and can be activated by oxidative events. Using a Src reporter based on fluorescence resonance energy transfer (FRET) technique, we visualized the dynamic Src activation in Hela cells immediately after LPLI. Moreover, Src activity was enhanced by increasing the duration of LPLI. In addition, our results suggested that ROS were key mediators of Src activation, as ROS scavenger, vitamin C decreased and exogenous H IIO II increased the activity of Src. Meanwhile, Gö6983 loading did not block the effect of LPLI. CCK-8 experiments proved that cell vitality was prominently improved by LPLI with all the doses we applied in our experiments ranging from 3 to 25J/cm2. The results indicated that LPLI/ROS/Src pathway may be involved in the LPLI biostimulation effects.

Disruption of Src Is Associated with Phenotypes Related to Williams-Beuren Syndrome and Altered Cellular Localization of TFII-I1,2

PubMed Central

Ivakine, Evgueni A.; Lam, Emily; Deurloo, Marielle; Dida, Joana; Zirngibl, Ralph A.

2015-01-01

Abstract Src is a nonreceptor protein tyrosine kinase that is expressed widely throughout the central nervous system and is involved in diverse biological functions. Mice homozygous for a spontaneous mutation in Src (Src thl/thl) exhibited hypersociability and hyperactivity along with impairments in visuospatial, amygdala-dependent, and motor learning as well as an increased startle response to loud tones. The phenotype of Src thl/thl mice showed significant overlap with Williams-Beuren syndrome (WBS), a disorder caused by the deletion of several genes, including General Transcription Factor 2-I (GTF2I). Src phosphorylation regulates the movement of GTF2I protein (TFII-I) between the nucleus, where it is a transcriptional activator, and the cytoplasm, where it regulates trafficking of transient receptor potential cation channel, subfamily C, member 3 (TRPC3) subunits to the plasma membrane. Here, we demonstrate altered cellular localization of both TFII-I and TRPC3 in the Src mutants, suggesting that disruption of Src can phenocopy behavioral phenotypes observed in WBS through its regulation of TFII-I. PMID:26464974

A Discovery Strategy for Selective Inhibitors of c-Src in Complex with the Focal Adhesion Kinase SH3/SH2-binding Region.

PubMed

Moroco, Jamie A; Baumgartner, Matthew P; Rust, Heather L; Choi, Hwan Geun; Hur, Wooyoung; Gray, Nathanael S; Camacho, Carlos J; Smithgall, Thomas E

2015-08-01

The c-Src tyrosine kinase co-operates with the focal adhesion kinase to regulate cell adhesion and motility. Focal adhesion kinase engages the regulatory SH3 and SH2 domains of c-Src, resulting in localized kinase activation that contributes to tumor cell metastasis. Using assay conditions where c-Src kinase activity required binding to a tyrosine phosphopeptide based on the focal adhesion kinase SH3-SH2 docking sequence, we screened a kinase-biased library for selective inhibitors of the Src/focal adhesion kinase peptide complex versus c-Src alone. This approach identified an aminopyrimidinyl carbamate compound, WH-4-124-2, with nanomolar inhibitory potency and fivefold selectivity for c-Src when bound to the phospho-focal adhesion kinase peptide. Molecular docking studies indicate that WH-4-124-2 may preferentially inhibit the 'DFG-out' conformation of the kinase active site. These findings suggest that interaction of c-Src with focal adhesion kinase induces a unique kinase domain conformation amenable to selective inhibition. © 2014 John Wiley & Sons A/S.

Association of p60c-src with endosomal membranes in mammalian fibroblasts

PubMed Central

1992-01-01

We have examined the subcellular localization of p60c-src in mammalian fibroblasts. Analysis of indirect immunofluorescence by three- dimensional optical sectioning microscopy revealed a granular cytoplasmic staining that co-localized with the microtubule organizing center. Immunofluorescence experiments with antibodies against a number of membrane markers demonstrated a striking co-localization between p60c-src and the cation-dependent mannose-6-phosphate receptor (CI- MPR), a marker that identifies endosomes. Both p60c-src and the CI-MPR were found to cluster at the spindle poles throughout mitosis. In addition, treatment of interphase and mitotic cells with brefeldin A resulted in a clustering of p60c-src and CI-MPR at a peri-centriolar position. Biochemical fractionation of cellular membranes showed that a major proportion of p60c-src co-enriched with endocytic membranes. Treatment of membranes containing HRP to alter their apparent density also altered the density of p60c-src-containing membranes. Similar density shift experiments with total cellular membranes revealed that the majority of membrane-associated p60c-src in the cell is associated with endosomes, while very little is associated with plasma membranes. These results support a role for p60c-src in the regulation of endosomal membranes and protein trafficking. PMID:1378446

Coactivator SRC-2–dependent metabolic reprogramming mediates prostate cancer survival and metastasis

PubMed Central

Dasgupta, Subhamoy; Putluri, Nagireddy; Long, Weiwen; Zhang, Bin; Wang, Jianghua; Kaushik, Akash K.; Arnold, James M.; Bhowmik, Salil K.; Stashi, Erin; Brennan, Christine A.; Rajapakshe, Kimal; Coarfa, Cristian; Mitsiades, Nicholas; Ittmann, Michael M.; Chinnaiyan, Arul M.; Sreekumar, Arun; O’Malley, Bert W.

2015-01-01

Metabolic pathway reprogramming is a hallmark of cancer cell growth and survival and supports the anabolic and energetic demands of these rapidly dividing cells. The underlying regulators of the tumor metabolic program are not completely understood; however, these factors have potential as cancer therapy targets. Here, we determined that upregulation of the oncogenic transcriptional coregulator steroid receptor coactivator 2 (SRC-2), also known as NCOA2, drives glutamine-dependent de novo lipogenesis, which supports tumor cell survival and eventual metastasis. SRC-2 was highly elevated in a variety of tumors, especially in prostate cancer, in which SRC-2 was amplified and overexpressed in 37% of the metastatic tumors evaluated. In prostate cancer cells, SRC-2 stimulated reductive carboxylation of α-ketoglutarate to generate citrate via retrograde TCA cycling, promoting lipogenesis and reprogramming of glutamine metabolism. Glutamine-mediated nutrient signaling activated SRC-2 via mTORC1-dependent phosphorylation, which then triggered downstream transcriptional responses by coactivating SREBP-1, which subsequently enhanced lipogenic enzyme expression. Metabolic profiling of human prostate tumors identified a massive increase in the SRC-2–driven metabolic signature in metastatic tumors compared with that seen in localized tumors, further implicating SRC-2 as a prominent metabolic coordinator of cancer metastasis. Moreover, SRC-2 inhibition in murine models severely attenuated the survival, growth, and metastasis of prostate cancer. Together, these results suggest that the SRC-2 pathway has potential as a therapeutic target for prostate cancer. PMID:25664849

Identification of the SRC pyrimidine-binding protein (SPy) as hnRNP K: implications in the regulation of SRC1A transcription

PubMed Central

Ritchie, Shawn A.; Pasha, Mohammed K.; Batten, Danielle J. P.; Sharma, Rajendra K.; Olson, Douglas J. H.; Ross, Andrew R. S.; Bonham, Keith

2003-01-01

The human SRC gene encodes pp60c–src, a non-receptor tyrosine kinase involved in numerous signaling pathways. Activation or overexpression of c-Src has also been linked to a number of important human cancers. Transcription of the SRC gene is complex and regulated by two closely linked but highly dissimilar promoters, each associated with its own distinct non-coding exon. In many tissues SRC expression is regulated by the housekeeping-like SRC1A promoter. In addition to other regulatory elements, three substantial polypurine:polypyrimidine (TC) tracts within this promoter are required for full transcriptional activity. Previously, we described an unusual factor called SRC pyrimidine-binding protein (SPy) that could bind to two of these TC tracts in their double-stranded form, but was also capable of interacting with higher affinity to all three pyrimidine tracts in their single-stranded form. Mutations in the TC tracts, which abolished the ability of SPy to interact with its double-stranded DNA target, significantly reduced SRC1A promoter activity, especially in concert with mutations in critical Sp1 binding sites. Here we expand upon our characterization of this interesting factor and describe the purification of SPy from human SW620 colon cancer cells using a DNA affinity-based approach. Subsequent in-gel tryptic digestion of purified SPy followed by MALDI-TOF mass spectrometric analysis identified SPy as heterogeneous nuclear ribonucleoprotein K (hnRNP K), a known nucleic-acid binding protein implicated in various aspects of gene expression including transcription. These data provide new insights into the double- and single-stranded DNA-binding specificity, as well as functional properties of hnRNP K, and suggest that hnRNP K is a critical component of SRC1A transcriptional processes. PMID:12595559

A Kinase-Independent Function of c-Src Mediates p130Cas Phosphorylation at the Serine-639 Site in Pressure Overloaded Myocardium

PubMed Central

Palanisamy, Arun P.; Suryakumar, Geetha; Panneerselvam, Kavin; Willey, Christopher D.; Kuppuswamy, Dhandapani

2017-01-01

Early work in pressure overloaded (PO) myocardium shows that integrins mediate focal adhesion complex formation by recruiting the adaptor protein p130Cas (Cas) and nonreceptor tyrosine kinase c-Src. To explore c-Src role in Cas-associated changes during PO, we used a feline right ventricular in vivo PO model and a three-dimensional (3D) collagen-embedded adult cardiomyocyte in vitro model that utilizes a Gly-Arg-Gly-Asp-Ser (RGD) peptide for integrin stimulation. Cas showed slow electrophoretic mobility (band-shifting), recruitment to the cytoskeleton, and tyrosine phosphorylation at 165, 249, and 410 sites in both 48 h PO myocardium and 1 h RGD-stimulated cardiomyocytes. Adenoviral mediated expression of kinase inactive (negative) c-Src mutant with intact scaffold domains (KN-Src) in cardiomyocytes did not block the RGD stimulated changes in Cas. Furthermore, expression of KN-Src or kinase active c-Src mutant with intact scaffold function (A-Src) in two-dimensionally (2D) cultured cardiomyocytes was sufficient to cause Cas band-shifting, although tyrosine phosphorylation required A-Src. These data indicate that c-Src’s adaptor function, but not its kinase function, is required for a serine/threonine specific phosphorylation(s) responsible for Cas band-shifting. To explore this possibility, Chinese hamster ovary cells that stably express Cas were infected with either β-gal or KN-Src adenoviruses and used for Cas immunoprecipitation combined with mass spectrometry analysis. In the KN-Src expressing cells, Cas showed phosphorylation at the serine-639 (human numbering) site. A polyclonal antibody raised against phospho-serine-639 detected Cas phosphorylation in 24–48 h PO myocardium. Our studies indicate that c-Src’s adaptor function mediates serine-639 phosphorylation of Cas during integrin activation in PO myocardium. PMID:25976166

Antiangiogenic and Antitumor Effects of Src Inhibition in Ovarian Carcinoma

PubMed Central

Han, Liz Y.; Landen, Charles N.; Trevino, Jose G.; Halder, Jyotsnabaran; Lin, Yvonne G.; Kamat, Aparna A.; Kim, Tae-Jin; Merritt, William M.; Coleman, Robert L.; Gershenson, David M.; Shakespeare, William C.; Wang, Yihan; Sundaramoorth, Raji; Metcalf, Chester A.; Dalgarno, David C.; Sawyer, Tomi K.; Gallick, Gary E.; Sood, Anil K.

2011-01-01

Src, a nonreceptor tyrosine kinase, is a key mediator for multiple signaling pathways that regulate critical cellular functions and is often aberrantly activated in a number of solid tumors, including ovarian carcinoma. The purpose of this study was to determine the role of activated Src inhibition on tumor growth in an orthotopic murine model of ovarian carcinoma. In vitro studies on HeyA8 and SKOV3ip1 cell lines revealed that Src inhibition by the Src-selective inhibitor, AP23846, occurred within 1 hour and responded in a dose-dependent manner. Furthermore, Src inhibition enhanced the cytotoxicity of docetaxel in both chemosensitive and chemoresistant ovarian cancer cell lines, HeyA8 and HeyA8-MDR, respectively. In vivo, Src inhibition by AP23994, an orally bioavailable analogue of AP23846, significantly decreased tumor burden in HeyA8 (P = 0.02), SKOV3ip1 (P = 0.01), as well as HeyA8-MDR (P < 0.03) relative to the untreated controls. However, the greatest effect on tumor reduction was observed in combination therapy with docetaxel (P < 0.001, P = 0.002, and P = 0.01, for the above models, respectively). Proliferating cell nuclear antigen staining showed that Src inhibition alone (P = 0.02) and in combination with docetaxel (P = 0.007) significantly reduced tumor proliferation. In addition, Src inhibition alone and in combination with docetaxel significantly down-regulated tumoral production of vascular endothelial growth factor and interleukin 8, whereas combination therapy decreased the microvessel density (P = 0.02) and significantly affected vascular permeability (P < 0.05). In summary, Src inhibition with AP23994 has potent antiangiogenic effects and significantly reduces tumor burden in preclinical ovarian cancer models. Thus, Src inhibition may be an attractive therapeutic approach for patients with ovarian carcinoma. PMID:16951177

v-Src oncogene product increases sphingosine kinase 1 expression through mRNA stabilization: alteration of AU-rich element-binding proteins.

PubMed

Sobue, S; Murakami, M; Banno, Y; Ito, H; Kimura, A; Gao, S; Furuhata, A; Takagi, A; Kojima, T; Suzuki, M; Nozawa, Y; Murate, T

2008-10-09

Sphingosine kinase 1 (SPHK1) is overexpressed in solid tumors and leukemia. However, the mechanism of SPHK1 overexpression by oncogenes has not been defined. We found that v-Src-transformed NIH3T3 cells showed a high SPHK1 mRNA, SPHK1 protein and SPHK enzyme activity. siRNA of SPHK1 inhibited the growth of v-Src-NIH3T3, suggesting the involvement of SPHK1 in v-Src-induced oncogenesis. v-Src-NIH3T3 showed activations of protein kinase C-alpha, signal transducers and activators of transcription 3 and c-Jun NH(2)-terminal kinase. Their inhibition suppressed SPHK1 expression in v-Src-NIH3T3, whereas their overexpression increased SPHK1 mRNA in NIH3T3. Unexpectedly, the nuclear run-on assay and the promoter analysis using 5'-promoter region of mouse SPHK1 did not show any significant difference between mock- and v-Src-NIH3T3. Furthermore, the half-life of SPHK1 mRNA in mock-NIH3T3 was nearly 15 min, whereas that of v-Src-NIH3T3 was much longer. Examination of two AU-rich region-binding proteins, AUF1 and HuR, that regulate mRNA decay reciprocally, showed decreased total AUF1 protein associated with increased tyrosine-phosphorylated form and increased serine-phosphorylated HuR protein in v-Src-NIH3T3. Modulation of AUF1 and HuR by their overexpression or siRNA revealed that SPHK1 mRNA in v-Src- and mock-NIH3T3 was regulated reciprocally by these factors. Our results showed, for the first time, a novel mechanism of v-Src-induced SPHK1 overexpression.

Platelet-derived growth factor-dependent association of the GTPase-activating protein of Ras and Src.

PubMed Central

Schlesinger, T K; Demali, K A; Johnson, G L; Kazlauskas, A

1999-01-01

Here we report that the platelet-derived growth factor beta receptor (betaPDGFR) is not the only tyrosine kinase able to associate with the GTPase-activating protein of Ras (RasGAP). The interaction of non-betaPDGFR kinase(s) with RasGAP was dependent on stimulation with platelet-derived growth factor (PDGF) and seemed to require tyrosine phosphorylation of RasGAP. Because the tyrosine phosphorylation site of RasGAP is in a sequence context that is favoured by the Src homology 2 ('SH2') domain of Src family members, we tested the possibility that Src was the kinase that associated with RasGAP. Indeed, Src interacted with phosphorylated RasGAP fusion proteins; immunodepletion of Src markedly decreased the recovery of the RasGAP-associated kinase activity. Thus PDGF-dependent tyrosine phosphorylation of RasGAP results in the formation of a complex between RasGAP and Src. To begin to address the relevance of these observations, we focused on the consequences of the interaction of Src and RasGAP. We found that a receptor mutant that did not activate Src was unable to efficiently mediate the tyrosine phosphorylation of phospholipase Cgamma (PLCgamma). Taken together, these observations support the following hypothesis. When RasGAP is recruited to the betaPDGFR, it is phosphorylated and associates with Src. Once bound to RasGAP, Src is no longer able to promote the phosphorylation of PLCgamma. This hypothesis offers a mechanistic explanation for our previously published findings that the recruitment of RasGAP to the betaPDGFR attenuates the tyrosine phosphorylation of PLCgamma. Finally, these findings suggest a novel way in which RasGAP negatively regulates signal relay by the betaPDGFR. PMID:10567236

DOE Office of Scientific and Technical Information (OSTI.GOV)

Los Alamos National Laboratory, Mailstop M888, Los Alamos, NM 87545, USA; Lawrence Berkeley National Laboratory, One Cyclotron Road, Building 64R0121, Berkeley, CA 94720, USA; Department of Haematology, University of Cambridge, Cambridge CB2 0XY, England

The PHENIX AutoBuild Wizard is a highly automated tool for iterative model-building, structure refinement and density modification using RESOLVE or TEXTAL model-building, RESOLVE statistical density modification, and phenix.refine structure refinement. Recent advances in the AutoBuild Wizard and phenix.refine include automated detection and application of NCS from models as they are built, extensive model completion algorithms, and automated solvent molecule picking. Model completion algorithms in the AutoBuild Wizard include loop-building, crossovers between chains in different models of a structure, and side-chain optimization. The AutoBuild Wizard has been applied to a set of 48 structures at resolutions ranging from 1.1 {angstrom} tomore » 3.2 {angstrom}, resulting in a mean R-factor of 0.24 and a mean free R factor of 0.29. The R-factor of the final model is dependent on the quality of the starting electron density, and relatively independent of resolution.« less

Separation of chlorogenic acid and concentration of trace caffeic acid from natural products by pH-zone-refining countercurrent chromatography.

PubMed

Lu, Yuanyuan; Dong, Genlai; Gu, Yanxiang; Ito, Yoichiro; Wei, Yun

2013-07-01

Chlorogenic acid and caffeic acid were selected as test samples for separation by the pH-zone-refining countercurrent chromatography (CCC). The separation of these test samples was performed with a two-phase solvent system composed of methyl-tert-butyl-ether/acetonitrile/water at a volume ratio of 4:1:5 v/v/v where trifluoroacetic acid (TFA; 8 mM) was added to the organic stationary phase as a retainer and NH4 OH (10 mM) to the aqueous mobile phase as an eluter. Chlorogenic acid was successfully separated from Flaveria bidentis (L.) Kuntze (F. bidentis) and Lonicerae Flos by pH-zone-refining CCC, a slightly polar two-phase solvent system composed of methyl-tert-butyl-ether/acetonitrile/n-butanol/water at a volume ratio of 4:1:1:5 v/v/v/v was selected where TFA (3 mM) was added to the organic stationary phase as a retainer and NH4 OH (3 mM) to the aqueous mobile phase as an eluter. A 16.2 mg amount of chlorogenic acid with the purity of 92% from 1.4 g of F. bidentis, and 134 mg of chlorogenic acid at the purity of 99% from 1.3 g of crude extract of Lonicerae Flos have been obtained. These results suggest that pH-zone-refining CCC is suitable for the isolation of the chlorogenic acid from the crude extracts of F. bidentis and Lonicerae Flos. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Differentiation-induced Colocalization of the KH-type Splicing Regulatory Protein with Polypyrimidine Tract Binding Protein and the c-src Pre-mRNA

PubMed Central

Hall, Megan P.; Huang, Sui; Black, Douglas L.

2004-01-01

We have examined the subcellular localization of the KH-type splicing regulatory protein (KSRP). KSRP is a multidomain RNA-binding protein implicated in a variety of cellular processes, including splicing in the nucleus and mRNA localization in the cytoplasm. We find that KSRP is primarily nuclear with a localization pattern that most closely resembles that of polypyrimidine tract binding protein (PTB). Colocalization experiments of KSRP with PTB in a mouse neuroblastoma cell line determined that both proteins are present in the perinucleolar compartment (PNC), as well as in other nuclear enrichments. In contrast, HeLa cells do not show prominent KSRP staining in the PNC, even though PTB labeling identified the PNC in these cells. Because both PTB and KSRP interact with the c-src transcript to affect N1 exon splicing, we examined the localization of the c-src pre-mRNA by fluorescence in situ hybridization. The src transcript is present in specific foci within the nucleus that are presumably sites of src transcription but are not generally perinucleolar. In normally cultured neuroblastoma cells, these src RNA foci contain PTB, but little KSRP. However, upon induced neuronal differentiation of these cells, KSRP occurs in the same foci with src RNA. PTB localization remains unaffected. This differentiation-induced localization of KSRP with src RNA correlates with an increase in src exon N1 inclusion. These results indicate that PTB and KSRP do indeed interact with the c-src transcript in vivo, and that these associations change with the differentiated state of the cell. PMID:14657238

Inhibition of src family kinases by a combinatorial action of 5'-AMP and small heat shock proteins, identified from the adult heart.

PubMed

Kasi, V S; Kuppuswamy, D

1999-10-01

Src family kinases are implicated in cellular proliferation and transformation. Terminally differentiated myocytes have lost the ability to proliferate, indicating the existence of a down-regulatory mechanism(s) for these mitogenic kinases. Here we show that feline cardiomyocyte lysate contains thermostable components that inhibit c-Src kinase in vitro. This inhibitory activity, present predominantly in heart tissue, involves two components acting combinatorially. After purification by sequential chromatography, one component was identified by mass and nuclear magnetic resonance spectroscopies as 5'-AMP, while the other was identified by peptide sequencing as a small heat shock protein (sHSP). 5'-AMP and to a lesser extent 5'-ADP inhibit c-Src when combined with either HSP-27 or HSP-32. Other HSPs, including alphaB-crystallin, HSP-70, and HSP-90, did not exhibit this effect. The inhibition, observed preferentially on Src family kinases and independent of the Src tyrosine phosphorylation state, occurs via a direct interaction of the c-Src catalytic domain with the inhibitory components. Our study indicates that sHSPs increase the affinity of 5'-AMP for the c-Src ATP binding site, thereby facilitating the inhibition. In vivo, elevation of ATP levels in the cardiomyocytes results in the tyrosine phosphorylation of cellular proteins including c-Src at the activatory site, and this effect is blocked when the 5'-AMP concentration is raised. Thus, this study reveals a novel role for sHSPs and 5'-AMP in the regulation of Src family kinases, presumably for the maintenance of the terminally differentiated state.

Structure-based design of an osteoclast-selective, nonpeptide Src homology 2 inhibitor with in vivo antiresorptive activity

PubMed Central

Shakespeare, William; Yang, Michael; Bohacek, Regine; Cerasoli, Franklin; Stebbins, Karin; Sundaramoorthi, Raji; Azimioara, Mihai; Vu, Chi; Pradeepan, Selvi; Metcalf, Chester; Haraldson, Chad; Merry, Taylor; Dalgarno, David; Narula, Surinder; Hatada, Marcos; Lu, Xiaode; van Schravendijk, Marie Rose; Adams, Susan; Violette, Shelia; Smith, Jeremy; Guan, Wei; Bartlett, Catherine; Herson, Jay; Iuliucci, John; Weigele, Manfred; Sawyer, Tomi

2000-01-01

Targeted disruption of the pp60src (Src) gene has implicated this tyrosine kinase in osteoclast-mediated bone resorption and as a therapeutic target for the treatment of osteoporosis and other bone-related diseases. Herein we describe the discovery of a nonpeptide inhibitor (AP22408) of Src that demonstrates in vivo antiresorptive activity. Based on a cocrystal structure of the noncatalytic Src homology 2 (SH2) domain of Src complexed with citrate [in the phosphotyrosine (pTyr) binding pocket], we designed 3′,4′-diphosphonophenylalanine (Dpp) as a pTyr mimic. In addition to its design to bind Src SH2, the Dpp moiety exhibits bone-targeting properties that confer osteoclast selectivity, hence minimizing possible undesired effects on other cells that have Src-dependent activities. The chemical structure AP22408 also illustrates a bicyclic template to replace the post-pTyr sequence of cognate Src SH2 phosphopeptides such as Ac-pTyr-Glu-Glu-Ile (1). An x-ray structure of AP22408 complexed with Lck (S164C) SH2 confirmed molecular interactions of both the Dpp and bicyclic template of AP22408 as predicted from molecular modeling. Relative to the cognate phosphopeptide, AP22408 exhibits significantly increased Src SH2 binding affinity (IC50 = 0.30 μM for AP22408 and 5.5 μM for 1). Furthermore, AP22408 inhibits rabbit osteoclast-mediated resorption of dentine in a cellular assay, exhibits bone-targeting properties based on a hydroxyapatite adsorption assay, and demonstrates in vivo antiresorptive activity in a parathyroid hormone-induced rat model. PMID:10944210

Src promotes delta opioid receptor (DOR) desensitization by interfering with receptor recycling.

PubMed

Archer-Lahlou, Elodie; Audet, Nicolas; Amraei, Mohammad Gholi; Huard, Karine; Paquin-Gobeil, Mélanie; Pineyro, Graciela

2009-01-01

Abstract An important limitation in the clinical use of opiates is progressive loss of analgesic efficacy over time. Development of analgesic tolerance is tightly linked to receptor desensitization. In the case of delta opioid receptors (DOR), desensitization is especially swift because receptors are rapidly internalized and are poorly recycled to the membrane. In the present study, we investigated whether Src activity contributed to this sorting pattern and to functional desensitization of DORs. A first series of experiments demonstrated that agonist binding activates Src and destabilizes a constitutive complex formed by the spontaneous association of DORs with the kinase. Src contribution to DOR desensitization was then established by showing that pre-treatment with Src inhibitor PP2 (20 microM; 1 hr) or transfection of a dominant negative Src mutant preserved DOR signalling following sustained exposure to an agonist. This protection was afforded without interfering with endocytosis, but suboptimal internalization interfered with PP2 ability to preserve DOR signalling, suggesting a post-endocytic site of action for the kinase. This assumption was confirmed by demonstrating that Src inhibition by PP2 or its silencing by siRNA increased membrane recovery of internalized DORs and was further corroborated by showing that inhibition of recycling by monensin or dominant negative Rab11 (Rab11S25N) abolished the ability of Src blockers to prevent desensitization. Finally, Src inhibitors accelerated recovery of DOR-Galphal3 coupling after desensitization. Taken together, these results indicate that Src dynamically regulates DOR recycling and by doing so contributes to desensitization of these receptors.

The Endoplasmic Reticulum Is a Reservoir for WAVE/SCAR Regulatory Complex Signaling in the Arabidopsis Leaf1[W][OA

PubMed Central

Zhang, Chunhua; Mallery, Eileen; Reagan, Sara; Boyko, Vitaly P.; Kotchoni, Simeon O.; Szymanski, Daniel B.

2013-01-01

During plant cell morphogenesis, signal transduction and cytoskeletal dynamics interact to locally organize the cytoplasm and define the geometry of cell expansion. The WAVE/SCAR (for WASP family verprolin homologous/suppressor of cyclic AMP receptor) regulatory complex (W/SRC) is an evolutionarily conserved heteromeric protein complex. Within the plant kingdom W/SRC is a broadly used effector that converts Rho-of-Plants (ROP)/Rac small GTPase signals into Actin-Related Protein2/3 and actin-dependent growth responses. Although the components and biochemistry of the W/SRC pathway are well understood, a basic understanding of how cells partition W/SRC into active and inactive pools is lacking. In this paper, we report that the endoplasmic reticulum (ER) is an important organelle for W/SRC regulation. We determined that a large intracellular pool of the core W/SRC subunit NAP1, like the known positive regulator of W/SRC, the DOCK family guanine nucleotide-exchange factor SPIKE1 (SPK1), localizes to the surface of the ER. The ER-associated NAP1 is inactive because it displays little colocalization with the actin network, and ER localization requires neither activating signals from SPK1 nor a physical association with its W/SRC-binding partner, SRA1. Our results indicate that in Arabidopsis (Arabidopsis thaliana) leaf pavement cells and trichomes, the ER is a reservoir for W/SRC signaling and may have a key role in the early steps of W/SRC assembly and/or activation. PMID:23613272

A Ser75-to-Asp phospho-mimicking mutation in Src accelerates ageing-related loss of retinal ganglion cells in mice.

PubMed

Kashiwagi, Kenji; Ito, Sadahiro; Maeda, Shuichiro; Kato, Goro

2017-12-01

Src knockout mice show no detectable abnormalities in central nervous system (CNS) post-mitotic neurons, likely reflecting functional compensation by other Src family kinases. Cdk1- or Cdk5-dependent Ser75 phosphorylation in the amino-terminal Unique domain of Src, which shares no homology with other Src family kinases, regulates the stability of active Src. To clarify the roles of Src Ser75 phosphorylation in CNS neurons, we established two types of mutant mice with mutations in Src: phospho-mimicking Ser75Asp (SD) and non-phosphorylatable Ser75Ala (SA). In ageing SD/SD mice, retinal ganglion cell (RGC) number in whole retinas was significantly lower than that in young SD/SD mice in the absence of inflammation and elevated intraocular pressure, resembling the pathogenesis of progressive optic neuropathy. By contrast, SA/SA mice and wild-type (WT) mice exhibited no age-related RGC loss. The age-related retinal RGC number reduction was greater in the peripheral rather than the mid-peripheral region of the retina in SD/SD mice. Furthermore, Rho-associated kinase activity in whole retinas of ageing SD/SD mice was significantly higher than that in young SD/SD mice. These results suggest that Src regulates RGC survival during ageing in a manner that depends on Ser75 phosphorylation.

Pharmacological inhibition of Src kinase protects against acute kidney injury in a murine model of renal ischemia/reperfusion

PubMed Central

Zhou, Xiaoxu; Liu, Lirong; Masucci, Monica V.; Tang, Jinhua; Li, Xuezhu; Liu, Na; Bayliss, George; Zhao, Ting C.; Zhuang, Shougang

2017-01-01

Activation of Src kinase has been implicated in the pathogenesis of acute brain, liver, and lung injury. However, the role of Src in acute kidney injury (AKI) remains unestablished. To address this, we evaluated the effects of Src inhibition on renal dysfunction and pathological changes in a murine model of AKI induced by ischemia/reperfusion (I/R). I/R injury to the kidney resulted in increased Src phosphorylation at tyrosine 416 (activation). Administration of PP1, a highly selective Src inhibitor, blocked Src phosphorylation, improved renal function and ameliorated renal pathological damage. PP1 treatment also suppressed renal expression of neutrophil gelatinase-associated lipocalin and reduced apoptosis in the injured kidney. Moreover, Src inhibition prevented downregulation of several adherens and tight junction proteins, including E-cadherin, ZO-1, and claudins-1/−4 in the kidney after I/R injury as well as in cultured renal proximal tubular cells following oxidative stress. Finally, PP1 inhibited I/R–induced renal expression of matrix metalloproteinase-2 and -9, phosphorylation of extracellular signal–regulated kinases1/2, signal transducer and activator of transcription-3, and nuclear factor-κB, and the infiltration of macrophages into the kidney. These data indicate that Src is a pivotal mediator of renal epithelial injury and that its inhibition may have a therapeutic potential to treat AKI. PMID:28415724

Clinicopathological Characteristics and Prognostic Value of Signet Ring Cells in Gastric Carcinoma: A Meta-Analysis.

PubMed

Nie, Run-Cong; Yuan, Shu-Qiang; Li, Yuan-Fang; Chen, Yong-Ming; Chen, Xiao-Jiang; Zhu, Bao-Yan; Xu, Li-Pu; Zhou, Zhi-Wei; Chen, Shi; Chen, Ying-Bo

2017-01-01

Background and Objectives: Previous studies of the prognostic value of the signet ring cell (SRC) type have yielded inconsistent results. Therefore, the aim of the present meta-analysis is to explore the clinicopathological characteristics and prognostic value of SRCs. Methods: Relevant articles that compared SRC and non-SRC type in PubMed and Web of Science were comprehensively searched. Then, a meta-analysis was performed. Results: A total of 19 studies including 35947 cases were analyzed. Compared with non-SRC patients, SRC patients tended to be younger (WMD: -3.88, P=0.001) and predominantly female (OR: 1.60, P<0.001). Additionally, SRC patients exhibited less upper third tumor location (OR: 0.62, P<0.001) and less frequent hematogenous metastasis (OR: 0.41, P<0.001). There was no difference in overall survival (OS) between SRC and non-SRC patients in the total population (HR: 1.02, P=0.830). Early gastric cancer with SRCs was associated with better OS (HR: 0.57, P=0.002), while advanced gastric cancer with non-SRCs was associated with a worse prognosis (HR: 1.17, P<0.001). Conclusions: This meta-analysis revealed that SRC tends to affect young females and tends to be located in the middle and lower third of the stomach. Early SRCs are associated with better prognoses, while advanced SRCs are associated with worse prognoses.

Lyn tyrosine kinase promotes silencing of ATM-dependent checkpoint signaling during recovery from DNA double-strand breaks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fukumoto, Yasunori, E-mail: fukumoto@faculty.chiba-u.jp; Kuki, Kazumasa; Morii, Mariko

2014-09-26

Highlights: • Inhibition of Src family kinases decreased γ-H2AX signal. • Inhibition of Src family increased ATM-dependent phosphorylation of Chk2 and Kap1. • shRNA-mediated knockdown of Lyn increased phosphorylation of Kap1 by ATM. • Ectopic expression of Src family kinase suppressed ATM-mediated Kap1 phosphorylation. • Src is involved in upstream signaling for inactivation of ATM signaling. - Abstract: DNA damage activates the DNA damage checkpoint and the DNA repair machinery. After initial activation of DNA damage responses, cells recover to their original states through completion of DNA repair and termination of checkpoint signaling. Currently, little is known about the processmore » by which cells recover from the DNA damage checkpoint, a process called checkpoint recovery. Here, we show that Src family kinases promote inactivation of ataxia telangiectasia mutated (ATM)-dependent checkpoint signaling during recovery from DNA double-strand breaks. Inhibition of Src activity increased ATM-dependent phosphorylation of Chk2 and Kap1. Src inhibition increased ATM signaling both in G2 phase and during asynchronous growth. shRNA knockdown of Lyn increased ATM signaling. Src-dependent nuclear tyrosine phosphorylation suppressed ATM-mediated Kap1 phosphorylation. These results suggest that Src family kinases are involved in upstream signaling that leads to inactivation of the ATM-dependent DNA damage checkpoint.« less

40 CFR Appendix V to Part 86 - The Standard Road Cycle (SRC)

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 19 2010-07-01 2010-07-01 false The Standard Road Cycle (SRC) V... Appendix V to Part 86—The Standard Road Cycle (SRC) 1. The standard road cycle (SRC) is a mileage accumulation cycle that may be used for any vehicle which is covered by the applicability provisions of § 86...

N-terminal deletions in Rous sarcoma virus p60src: effects on tyrosine kinase and biological activities and on recombination in tissue culture with the cellular src gene.

PubMed Central

Cross, F R; Garber, E A; Hanafusa, H

1985-01-01

We have constructed deletions within the region of cloned Rous sarcoma virus DNA coding for the N-terminal 30 kilodaltons of p60src. Infectious virus was recovered after transfection. Deletions of amino acids 15 to 149, 15 to 169, or 149 to 169 attenuated but did not abolish transforming activity, as assayed by focus formation and anchorage-independent growth. These deletions also had only slight effects on the tyrosine kinase activity of the mutant src protein. Deletion of amino acids 169 to 264 or 15 to 264 completely abolished transforming activity, and src kinase activity was reduced at least 10-fold. However, these mutant viruses generated low levels of transforming virus by recombination with the cellular src gene. The results suggest that as well as previously identified functional domains for p60src myristylation and membrane binding (amino acids 1 to 14) and tyrosine kinase activity (amino acids 250 to 526), additional N-terminal sequences (particularly amino acids 82 to 169) can influence the transforming activity of the src protein. Images PMID:2426576

c-Src activity is differentially required by cancer cell motility modes.

PubMed

Logue, Jeremy S; Cartagena-Rivera, Alexander X; Chadwick, Richard S

2018-04-01

Cancer cell migration requires that cells respond and adapt to their surroundings. In the absence of extracellular matrix cues, cancer cells will undergo a mesenchymal to ameboid transition, whereas a highly confining space will trigger a switch to "leader bleb-based" migration. To identify oncogenic signaling pathways mediating these transitions, we undertook a targeted screen using clinically useful inhibitors. Elevated Src activity was found to change actin and focal adhesion dynamics, whereas inhibiting Src triggered focal adhesion disassembly and blebbing. On non-adherent substrates and in collagen matrices, amoeboid-like, blebbing cells having high Src activity formed protrusions of the plasma membrane. To evaluate the role of Src in confined cells, we use a novel approach that places cells under a slab of polydimethylsiloxane (PDMS), which is held at a defined height. Using this method, we find that leader bleb-based migration is resistant to Src inhibition. High Src activity was found to markedly change the architecture of cortical actomyosin, reduce cell mechanical properties, and the percentage of cells that undergo leader bleb-based migration. Thus, Src is a signal transducer that can potently influence transitions between migration modes with implications for the rational development of metastasis inhibitors.

Inhibition of SRC-3 enhances sensitivity of human cancer cells to histone deacetylase inhibitors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zou, Zhengzhi, E-mail: zouzhengzhi@m.scnu.edu.cn; Luo, Xiaoyong; Nie, Peipei

SRC-3 is widely expressed in multiple tumor types and involved in cancer cell proliferation and apoptosis. Histone deacetylase (HDAC) inhibitors are promising antitumor drugs. However, the poor efficacy of HDAC inhibitors in solid tumors has restricted its further clinical application. Here, we reported the novel finding that depletion of SRC-3 enhanced sensitivity of breast and lung cancer cells to HDAC inhibitors (SAHA and romidepsin). In contrast, overexpression of SRC-3 decreased SAHA-induced cancer cell apoptosis. Furthermore, we found that SRC-3 inhibitor bufalin increased cancer cell apoptosis induced by HDAC inhibitors. The combination of bufalin and SAHA was particular efficient in attenuatingmore » AKT activation and reducing Bcl-2 levels. Taken together, these accumulating data might guide development of new breast and lung cancer therapies. - Highlights: • Depletion of SRC-3 enhanced sensitivity of breast and lung cancer cells to HDAC inhibitors. • Overexpression of SRC-3 enhanced cancer cell resistance to HDAC inhibitors. • SRC-3 inhibitor bufalin increased cancer cell apoptosis induced by HDAC inhibitors. • Bufalin synergized with HDAC inhibitor attenuated AKT activation and reduced Bcl-2 levels in human cancer cell.« less

Prognostic Significance of Signet Ring Gastric Cancer

PubMed Central

Taghavi, Sharven; Jayarajan, Senthil N.; Davey, Adam; Willis, Alliric I.

2012-01-01

Purpose Studies in Asia have questioned the dictum that signet ring cell carcinoma (SRC) has a worse prognosis than other forms of gastric cancer. Our study determined differences in presentation and outcomes between SRC and gastric adenocarcinoma (AC) in the United States. Patients and Methods The National Cancer Institute Surveillance, Epidemiology, and End Results database was reviewed for SRC and AC from 2004 to 2007. Results We reviewed 10,246 cases of patients with gastric cancer, including 2,666 of SRC and 7,580 of AC. SRC presented in younger patients (61.9 v 68.7 years; P < .001) and less often in men (52.7% v 68.7%; P < .001). SRC patients were more frequently black (11.3% v 10.9%), Asian (16.4% v 13.2%), American Indian/Alaska Native (0.9% v 0.8%), or Hispanic (23.3% v 14.0%; P < .001). SRC was more likely to be stage T3-4 (45.8% v 33.3%), have lymph node spread (59.7% v 51.8%), and distant metastases (40.2% v 37.6%; P < .001). SRC was more likely to be found in the lower (30.7% v 24.2%) and middle stomach (30.6% v 20.7%; P < .001). Median survival was not different between the two (AC, 14.0 months v SRC, 13.0 months; P = .073). Multivariable analyses demonstrated SRC was not associated with mortality (hazard ratio [HR], 1.05; 95% CI, 0.96 to 1.11; P = .150). Mortality was associated with age (HR, 1.01; 95% CI, 1.01 to 1.02; P < .001), black race (HR, 1.10; 95% CI, 1.01 to 1.20; P = .026), and tumor grade. Variables associated with lower mortality risk included Asian race (HR, 0.83; 95% CI, 0.77 to 0.91; P < .001) and surgery (HR, 0.37; 95% CI, 0.34 to 0.39; P < .001). Conclusion In the United States, SRC significantly differs from AC in extent of disease at presentation. However, when adjusted for stage, SRC does not portend a worse prognosis. PMID:22927530

Lemongrass essential oil and citral inhibit Src/Stat3 activity and suppress the proliferation/survival of small-cell lung cancer cells, alone or in combination with chemotherapeutic agents.

PubMed

Maruoka, Takayuki; Kitanaka, Akira; Kubota, Yoshitsugu; Yamaoka, Genji; Kameda, Tomohiro; Imataki, Osamu; Dobashi, Hiroaki; Bandoh, Shuji; Kadowaki, Norimitsu; Tanaka, Terukazu

2018-03-13

Small-cell lung cancer (SCLC) is intractable due to its high propensity for relapse. Novel agents are thus needed for SCLC treatment. Lemongrass essential oil (LG-EO) and its major constituent, citral, have been reported to inhibit the proliferation and survival of several types of cancer cells. However, the precise mechanisms through which LG-EO and citral exert their effects on SCLC cells have not been fully elucidated. SCLC cells express Src and have high levels of Src-tyrosine kinase (Src-TK) activity. In most SCLC cell lines, constitutive phosphorylation of Stat3(Y705), which is essential for its activation, has been detected. Src-TK can phosphorylate Stat3(Y705), and activated Stat3 promotes the expression of the anti-apoptotic factors Bcl-xL and Mcl-1. In the present study, LG-EO and citral prevented Src-TK from phosphorylating Stat3(Y705), resulting in decreased Bcl-xL and Mcl-1 expression, in turn suppressing the proliferation/survival of SCLC cells. To confirm these findings, the wild-type-src gene was transfected into the LU135 SCLC cell line (LU135‑wt-src), in which Src and activated phospho-Stat3(Y705) were overexpressed. The suppression of cell proliferation and the induction of apoptosis by treatment with LG-EO or citral were significantly attenuated in the LU135-wt-src cells compared with the control LU135-mock cells. The signal transducer and activator of transcription 3 (Stat3) signaling pathway is also associated with intrinsic drug resistance. LU135-wt-src cells were significantly resistant to conventional chemotherapeutic agents compared with LU135-mock cells. The combined effects of citral and each conventional chemotherapeutic agent on SCLC cells were also evaluated. The combination treatment exerted additive or more prominent effects on LU135-wt-src, LU165 and MN1112 cells, which are relatively chemoresistant SCLC cells. These findings suggest that either LG-EO or citral, alone or in combination with chemotherapeutic agents, may be a novel therapeutic option for SCLC patients.

The role of Src kinase in the biology and pathogenesis of Acanthamoeba castellanii

PubMed Central

2012-01-01

Background Acanthamoeba species are the causative agents of fatal granulomatous encephalitis in humans. Haematogenous spread is thought to be a primary step, followed by blood–brain barrier penetration, in the transmission of Acanthmaoeba into the central nervous system, but the associated molecular mechanisms remain unclear. Here, we evaluated the role of Src, a non-receptor protein tyrosine kinase in the biology and pathogenesis of Acanthamoeba. Methods Amoebistatic and amoebicidal assays were performed by incubating amoeba in the presence of Src kinase-selective inhibitor, PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine) and its inactive analog, PP3 (4-amino-7-phenylpyrazolo[3,4-d]pyrimidine). Using this inhibitor, the role of Src kinase in A. castellanii interactions with Escherichia coli was determined. Zymographic assays were performed to study effects of Src kinase on extracellular proteolytic activities of A. castellanii. The human brain microvascular endothelial cells were used to determine the effects of Src kinase on A. castellanii adhesion to and cytotoxicity of host cells. Results Inhibition of Src kinase using a specific inhibitor, PP2 (4-amino-5-(4 chlorophenyl)-7-(t-butyl)pyrazolo [3,4-d] pyrimidine) but not its inactive analog, PP3 (4-amino-7-phenylpyrazolo[3,4-d] pyrimidine), had detrimental effects on the growth of A. castellanii (keratitis isolate, belonging to the T4 genotype). Interestingly, inhibition of Src kinase hampered the phagocytic ability of A. castellanii, as measured by the uptake of non-invasive bacteria, but, on the contrary, invasion by pathogenic bacteria was enhanced. Zymographic assays revealed that inhibition of Src kinases reduced extracellular protease activities of A. castellanii. Src kinase inhibition had no significant effect on A. castellanii binding to and cytotoxicity of primary human brain microvascular endothelial cells, which constitute the blood–brain barrier. Conclusions For the first time, these findings demonstrated that Src kinase is involved in A. castellanii proliferation, protease secretions and phagocytic properties. Conversely, invasion of Acanthamoeba by pathogenic bacteria was stimulated by Src kinase inhibition. PMID:22676352

Method for reclaiming waste lubricating oils

DOEpatents

Whisman, Marvin L.; Goetzinger, John W.; Cotton, Faye O.

1978-01-01

A method for purifying and reclaiming used lubricating oils containing additives such as detergents, antioxidants, corrosion inhibitors, extreme pressure agents and the like and other solid and liquid contaminants by preferably first vacuum distilling the used oil to remove water and low-boiling contaminants, and treating the dried oil with a solvent mixture of butanol, isopropanol and methylethyl ketone which causes the separation of a layer of sludge containing contaminants, unspent additives and oxidation products. After solvent recovery, the desludged oil is then subjected to conventional lubricating oil refining steps such as distillation followed by decolorization and deodorization.

Evidence for in vivo phosphorylation of the Grb2 SH2-domain binding site on focal adhesion kinase by Src-family protein-tyrosine kinases.

PubMed

Schlaepfer, D D; Hunter, T

1996-10-01

Focal adhesion kinase (FAK) is a nonreceptor protein-tyrosine kinase (PTK) that associates with integrin receptors and participates in extracellular matrix-mediated signal transduction events. We showed previously that the c-Src nonreceptor PTK and the Grb2 SH2/SH3 adaptor protein bound directly to FAK after fibronectin stimulation (D. D. Schlaepfer, S.K. Hanks, T. Hunter, and P. van der Geer, Nature [London] 372:786-791, 1994). Here, we present evidence that c-Src association with FAK is required for Grb2 binding to FAK. Using a tryptic phosphopeptide mapping approach, the in vivo phosphorylation of the Grb2 binding site on FAK (Tyr-925) was detected after fibronectin stimulation of NIH 3T3 cells and was constitutively phosphorylated in v-Src-transformed NIH 3T3 cells. In vitro, c-Src phosphorylated FAK Tyr-925 in a glutathione S-transferase-FAK C-terminal domain fusion protein, whereas FAK did not. Using epitope-tagged FAK constructs, transiently expressed in human 293 cells, we determined the effect of site-directed mutations on c-Src and Grb2 binding to FAK. Mutation of FAK Tyr-925 disrupted Grb2 binding, whereas mutation of the c-Src binding site on FAK (Tyr-397) disrupted both c-Src and Grb2 binding to FAK in vivo. These results support a model whereby Src-family PTKs are recruited to FAK and focal adhesions following integrin-induced autophosphorylation and exposure of FAK Tyr-397. Src-family binding and phosphorylation of FAK at Tyr-925 creates a Grb2 SH2-domain binding site and provides a link to the activation of the Ras signal transduction pathway. In Src-transformed cells, this pathway may be constitutively activated as a result of FAK Tyr-925 phosphorylation in the absence of integrin stimulation.

Focal complex formation in adult cardiomyocytes is accompanied by the activation of beta3 integrin and c-Src.

PubMed

Willey, Christopher D; Balasubramanian, Sundaravadivel; Rodríguez Rosas, María C; Ross, Robert S; Kuppuswamy, Dhandapani

2003-06-01

In pressure-overloaded myocardium, our recent study demonstrated cytoskeletal assembly of c-Src and other signaling proteins which was partially mimicked in vitro using adult feline cardiomyocytes embedded in three-dimensional (3D) collagen matrix and stimulated with an integrin-binding Arg-Gly-Asp (RGD) peptide. In the present study, we improved this model further to activate c-Src and obtain a full assembly of the focal adhesion complex (FAC), and characterized c-Src localization and integrin subtype(s) involved. RGD dose response experiments revealed that c-Src activation occurs subsequent to its cytoskeletal recruitment and is accompanied by p130Cas cytoskeletal binding and focal adhesion kinase (FAK) Tyr925 phosphorylation. When cardiomyocytes expressing hexahistidine-tagged c-Src via adenoviral gene delivery were used for RGD stimulation, the expressed c-Src exhibited relocation: (i) biochemical analysis revealed c-Src movement from the detergent-soluble to the -insoluble cytoskeletal fraction and (ii) confocal microscopic analysis showed c-Src movement from a nuclear/perinuclear to a sarcolemmal region. RGD treatment also caused sarcolemmal co-localization of FAK and vinculin. Characterization of integrin subtypes revealed that beta3, but not beta1, integrin plays a predominant role: (i) expression of cytoplasmic domain of beta1A integrin did not affect the RGD-stimulated FAC formation and (ii) both pressure-overloaded myocardium and RGD-stimulated cardiomyocytes exhibited phosphorylation of beta3 integrin at Tyr773/785 sites but not beta1 integrin at Thr788/789 sites. Together these data indicate that RGD treatment in cardiomyocytes causes beta3 integrin activation and c-Src sarcolemmal localization, that subsequent c-Src activation is accompanied by p130Cas binding and FAK Tyr925 phosphorylation, and that these events might be crucial for growth and remodeling of hypertrophying adult cardiomyocytes.

v-src induction of the TIS10/PGS2 prostaglandin synthase gene is mediated by an ATF/CRE transcription response element.

PubMed

Xie, W; Fletcher, B S; Andersen, R D; Herschman, H R

1994-10-01

We recently reported the cloning of a mitogen-inducible prostaglandin synthase gene, TIS10/PGS2. In addition to growth factors and tumor promoters, the v-src oncogene induces TIS10/PGS2 expression in 3T3 cells. Deletion analysis, using luciferase reporters, identifies a region between -80 and -40 nucleotides 5' of the TIS10/PGS2 transcription start site that mediates pp60v-src induction in 3T3 cells. This region contains the sequence CGTCACGTG, which includes overlapping ATF/CRE (CGTCA) and E-box (CACGTG) sequences. Gel shift-oligonucleotide competition experiments with nuclear extracts from cells stably transfected with a temperature-sensitive v-src gene demonstrate that the CGTCACGTG sequence can bind proteins at both the ATF/CRE and E-box sequences. Dominant-negative CREB and Myc proteins that bind DNA, but do not transactivate, block v-src induction of a luciferase reporter driven by the first 80 nucleotides of the TIS10/PGS2 promoter. Mutational analysis distinguishes which TIS10/PGS2 cis-acting element mediates pp60v-src induction. E-box mutation has no effect on the fold induction in response to pp60v-src. In contrast, ATF/CRE mutation attenuates the pp60v-src response. Antibody supershift and methylation interference experiments demonstrate that CREB and at least one other ATF transcription factor in these extracts bind to the TIS10/PGS2 ATF/CRE element. Expression of a dominant-negative ras gene also blocks TIS10/PGS2 induction by v-src. Our data suggest that Ras mediates pp60v-src activation of an ATF transcription factor, leading to induced TIS10/PGS2 expression via the ATF/CRE element of the TIS10/PGS2 promoter. This is the first description of v-src activation of gene expression via an ATF/CRE element.

Tuning Structural Properties of Biocompatible Block Copolymer Micelles by Varying Solvent Composition

NASA Astrophysics Data System (ADS)

Cooksey, Tyler; Singh, Avantika; Mai Le, Kim; Wang, Shu; Kelley, Elizabeth; He, Lilin; Vajjala Kesava, Sameer; Gomez, Enrique; Kidd, Bryce; Madsen, Louis; Robertson, Megan

The self-assembly of block copolymers into micelles when introduced to selective solvents enables a wide array of applications, ranging from drug delivery to personal care products to nanoreactors. In order to probe the assembly and dynamics of micellar systems, the structural properties and solvent uptake of biocompatible poly(ethylene oxide-b- ɛ-caprolactone) (PEO-PCL) diblock copolymers in deuterated water (D2O) / tetrahydrofuran (THFd8) mixtures were investigated using small-angle neutron scattering in combination with nuclear magnetic resonance. PEO-PCL block copolymers, of varying molecular weight yet constant block ratio, formed spherical micelles through a wide range of solvent compositions. Varying the composition from 10 to 60 % by volume THFd8\\ in D2O / THFd8 mixtures was a means of varying the core-corona interfacial tension in the micelle system. An increase in THFd8 content in the bulk solvent increased the solvent uptake within the micelle core, which was comparable for the two series, irrespective of the polymer molecular weight. Differences in the behaviors of the micelle size parameters as the solvent composition varied originated from the differing trends in aggregation number for the two micelle series. Incorporation of the known unimer content determined from NMR allowed refinement of extracted micelle parameters.

Inhibition of Src Family Kinases by a Combinatorial Action of 5′-AMP and Small Heat Shock Proteins, Identified from the Adult Heart

PubMed Central

Kasi, Vijaykumar S.; Kuppuswamy, Dhandapani

1999-01-01

Src family kinases are implicated in cellular proliferation and transformation. Terminally differentiated myocytes have lost the ability to proliferate, indicating the existence of a down-regulatory mechanism(s) for these mitogenic kinases. Here we show that feline cardiomyocyte lysate contains thermostable components that inhibit c-Src kinase in vitro. This inhibitory activity, present predominantly in heart tissue, involves two components acting combinatorially. After purification by sequential chromatography, one component was identified by mass and nuclear magnetic resonance spectroscopies as 5′-AMP, while the other was identified by peptide sequencing as a small heat shock protein (sHSP). 5′-AMP and to a lesser extent 5′-ADP inhibit c-Src when combined with either HSP-27 or HSP-32. Other HSPs, including αB-crystallin, HSP-70, and HSP-90, did not exhibit this effect. The inhibition, observed preferentially on Src family kinases and independent of the Src tyrosine phosphorylation state, occurs via a direct interaction of the c-Src catalytic domain with the inhibitory components. Our study indicates that sHSPs increase the affinity of 5′-AMP for the c-Src ATP binding site, thereby facilitating the inhibition. In vivo, elevation of ATP levels in the cardiomyocytes results in the tyrosine phosphorylation of cellular proteins including c-Src at the activatory site, and this effect is blocked when the 5′-AMP concentration is raised. Thus, this study reveals a novel role for sHSPs and 5′-AMP in the regulation of Src family kinases, presumably for the maintenance of the terminally differentiated state. PMID:10490624

Discovery of Diffuse Hard X-Ray Emission from the Vicinity of PSR J1648-4611 with Suzaku

NASA Astrophysics Data System (ADS)

Sakai, Michito; Matsumoto, Hironori; Haba, Yoshito; Kanou, Yasufumi; Miyamoto, Youhei

2013-06-01

We observed the pulsar PSR J1648-4611 with Suzaku. Two X-ray sources, Suzaku J1648-4610 (Src A) and Suzaku J1648-4615 (Src B), were found in the field of view. Src A is coincident with the pulsar PSR J1648-4611, which was also detected by the Fermi Gamma-ray Space Telescope. A hard-band image indicates that Src A is spatially extended. We found point sources in the vicinity of Src A by using a Chandra image of the same region, but the point sources have soft X-ray emission, and cannot explain the hard X-ray emission of Src A. The hard-band spectrum of Src A can be reproduced by a power-law model with a photon index of 2.0+0.9-0.7. The X-ray flux in the 2-10 keV band is 1.4 × 10-13 erg cm-2 s-1. The diffuse emission suggests a pulsar wind nebula around PSR J1648&"8211;4611, but the luminosity of Src A is much larger than that expected from the spin-down luminosity of the pulsar. Parts of the very-high-energy γ-ray emission of HESS J1646-458 may be powered by this pulsar wind nebula driven by PSR J1648-4611. Src B has soft emission, and its X-ray spectrum can be described by a power-law model with a photon index of 3.0+1.4-0.8. The X-ray flux in the 0.4-10 keV band is 6.4 × 10-14 erg s-1 cm-2. No counterpart for Src B has been found in the literature.

Activation of Src kinase by protein-tyrosine phosphatase-PEST in osteoclasts: comparative analysis of the effects of bisphosphonate and protein-tyrosine phosphatase inhibitor on Src activation in vitro.

PubMed

Chellaiah, Meenakshi A; Schaller, Michael D

2009-08-01

PTP-PEST is involved in the regulation of sealing ring formation in osteoclasts. In this article, we have shown a regulatory role for PTP-PEST on dephosphorylation of c-Src at Y527 and phosphorylation at Y418 in the catalytic site. Activation of Src in osteoclasts by over-expression of PTP-PEST resulted in the phosphorylation of cortactin at Y421 and WASP at Y294. Also enhanced as a result, is the interaction of Src, cortactin, and Arp2 with WASP. Moreover, the number of osteoclasts displaying sealing ring and bone resorbing activity was increased in response to PTP-PEST over-expression as compared with control osteoclasts. Cells expressing constitutively active-Src (527YDeltaF) simulate the effects mediated by PTP-PEST. Treatment of osteoclasts with a bisphosphonate alendronate or a potent PTP inhibitor PAO decreased the activity and phosphorylation of Src at Y418 due to reduced dephosphorylation state at Y527. Therefore, Src-mediated phosphorylation of cortactin and WASP as well as the formation of WASP.cortactin.Arp2 complex and sealing ring were reduced in these osteoclasts. Similar effects were observed in osteoclasts treated with an Src inhibitor PP2. We have shown that bisphosphonates could modulate the function of osteoclasts by inhibiting downstream signaling mediated by PTP-PEST/Src, in addition to its effect on the inhibition of the post-translational modification of small GTP-binding proteins such as Rab, Rho, and Rac as shown by others. The promising effects of the inhibitors PP2 and PAO on osteoclast function suggest a therapeutic approach for patients with bone metastases and osteoporosis as an alternative to bisphosphonates.

The Chromatin Assembly Factor Complex 1 (CAF1) and 5-Azacytidine (5-AzaC) Affect Cell Motility in Src-transformed Human Epithelial Cells.

PubMed

Endo, Akinori; Ly, Tony; Pippa, Raffaella; Bensaddek, Dalila; Nicolas, Armel; Lamond, Angus I

2017-01-06

Tumor invasion into surrounding stromal tissue is a hallmark of high grade, metastatic cancers. Oncogenic transformation of human epithelial cells in culture can be triggered by activation of v-Src kinase, resulting in increased cell motility, invasiveness, and tumorigenicity and provides a valuable model for studying how changes in gene expression cause cancer phenotypes. Here, we show that epithelial cells transformed by activated Src show increased levels of DNA methylation and that the methylation inhibitor 5-azacytidine (5-AzaC) potently blocks the increased cell motility and invasiveness induced by Src activation. A proteomic screen for chromatin regulators acting downstream of activated Src identified the replication-dependent histone chaperone CAF1 as an important factor for Src-mediated increased cell motility and invasion. We show that Src causes a 5-AzaC-sensitive decrease in both mRNA and protein levels of the p150 (CHAF1A) and p60 (CHAF1B), subunits of CAF1. Depletion of CAF1 in untransformed epithelial cells using siRNA was sufficient to recapitulate the increased motility and invasive phenotypes characteristic of transformed cells without activation of Src. Maintaining high levels of CAF1 by exogenous expression suppressed the increased cell motility and invasiveness phenotypes when Src was activated. These data identify a critical role of CAF1 in the dysregulation of cell invasion and motility phenotypes seen in transformed cells and also highlight an important role for epigenetic remodeling through DNA methylation for Src-mediated induction of cancer phenotypes. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Scavenger Receptor C Mediates Phagocytosis of White Spot Syndrome Virus and Restricts Virus Proliferation in Shrimp

PubMed Central

Yang, Ming-Chong; Shi, Xiu-Zhen; Yang, Hui-Ting; Sun, Jie-Jie; Xu, Ling; Wang, Xian-Wei; Zhao, Xiao-Fan

2016-01-01

Scavenger receptors are an important class of pattern recognition receptors that play several important roles in host defense against pathogens. The class C scavenger receptors (SRCs) have only been identified in a few invertebrates, and their role in the immune response against viruses is seldom studied. In this study, we firstly identified an SRC from kuruma shrimp, Marsupenaeus japonicus, designated MjSRC, which was significantly upregulated after white spot syndrome virus (WSSV) challenge at the mRNA and protein levels in hemocytes. The quantity of WSSV increased in shrimp after knockdown of MjSRC, compared with the controls. Furthermore, overexpression of MjSRC led to enhanced WSSV elimination via phagocytosis by hemocytes. Pull-down and co-immunoprecipitation assays demonstrated the interaction between MjSRC and the WSSV envelope protein. Electron microscopy observation indicated that the colloidal gold-labeled extracellular domain of MjSRC was located on the outer surface of WSSV. MjSRC formed a trimer and was internalized into the cytoplasm after WSSV challenge, and the internalization was strongly inhibited after knockdown of Mjβ-arrestin2. Further studies found that Mjβ-arrestin2 interacted with the intracellular domain of MjSRC and induced the internalization of WSSV in a clathrin-dependent manner. WSSV were co-localized with lysosomes in hemocytes and the WSSV quantity in shrimp increased after injection of lysosome inhibitor, chloroquine. Collectively, this study demonstrated that MjSRC recognized WSSV via its extracellular domain and invoked hemocyte phagocytosis to restrict WSSV systemic infection. This is the first study to report an SRC as a pattern recognition receptor promoting phagocytosis of a virus. PMID:28027319

Multiple Kernel Sparse Representation based Orthogonal Discriminative Projection and Its Cost-Sensitive Extension.

PubMed

Zhang, Guoqing; Sun, Huaijiang; Xia, Guiyu; Sun, Quansen

2016-07-07

Sparse representation based classification (SRC) has been developed and shown great potential for real-world application. Based on SRC, Yang et al. [10] devised a SRC steered discriminative projection (SRC-DP) method. However, as a linear algorithm, SRC-DP cannot handle the data with highly nonlinear distribution. Kernel sparse representation-based classifier (KSRC) is a non-linear extension of SRC and can remedy the drawback of SRC. KSRC requires the use of a predetermined kernel function and selection of the kernel function and its parameters is difficult. Recently, multiple kernel learning for SRC (MKL-SRC) [22] has been proposed to learn a kernel from a set of base kernels. However, MKL-SRC only considers the within-class reconstruction residual while ignoring the between-class relationship, when learning the kernel weights. In this paper, we propose a novel multiple kernel sparse representation-based classifier (MKSRC), and then we use it as a criterion to design a multiple kernel sparse representation based orthogonal discriminative projection method (MK-SR-ODP). The proposed algorithm aims at learning a projection matrix and a corresponding kernel from the given base kernels such that in the low dimension subspace the between-class reconstruction residual is maximized and the within-class reconstruction residual is minimized. Furthermore, to achieve a minimum overall loss by performing recognition in the learned low-dimensional subspace, we introduce cost information into the dimensionality reduction method. The solutions for the proposed method can be efficiently found based on trace ratio optimization method [33]. Extensive experimental results demonstrate the superiority of the proposed algorithm when compared with the state-of-the-art methods.

Mouse fibroblasts homozygous for c-Src oncogene disruption shows dramatic suppression of expression of the gene encoding osteopontin, and adhesive phosphoprotein implicated in bone differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chackalaparampil, I.; Mukherjee, B.B.; Peri, A.

1994-09-01

Osteopetrosis, affecting mice and humans alike, arises from reduced or impaired bone resorption, causing abnormally dense bone formation. Normal bone differentiation requires continuous resorption and remodeling by osteoclasts which are derived from monocyte/macrophage lineage in the bone marrow. It has been reported that targeted homozygous disruption of c-src proto-oncogene in mice results in the development of osteopetrosis due to impaired bone-resorbing function of osteoclast cells. However, the molecular mechanism(s) which leads to osteoclast dysfunction in c-src deficient (src{sup -/-}) mice remains unclear. Here, we report that in embryonic fibroblasts derived from homozygous Src{sup -/-} mice, the expression of the genemore » coding for osteopontin (OP), a phosphorylated glycoprotein involved in bone differentiation, is drastically repressed. OP gene expression is not, however, affected in the heterozygous (Src{sup +/-}) mutant cells of identical origin, or in the c-src expression and OP production. Moreover, OP expression in c-src-deficient cells could be rescued upon treatment with 12-0-tetradecanoyl phorbol-13-myristate-acetate or okadaic acid. These observations indicate that OP expression is regulated via an src-mediated protein kinase C signaling pathway. Since it is known that OP mediates osteoclast adherence to the bone matrix, a key event in bone differentiation, our data is most significant in that they strongly suggest that drastic inhibition of synthesis of OP prevents osteoclasts in Src{sup -/-} mice from anchoring to the bone matrix. Consequently, this disruption of osteoclast adherence impairs their ability to form bone-resorbing ruffled border, causing osteopetrosis.« less

A Multifactorial Approach to Sport-Related Concussion Prevention and Education: Application of the Socioecological Framework.

PubMed

Register-Mihalik, Johna; Baugh, Christine; Kroshus, Emily; Y Kerr, Zachary; Valovich McLeod, Tamara C

2017-03-01

To offer an overview of sport-related concussion (SRC) prevention and education strategies in the context of the socioecological framework (SEF). Athletic trainers (ATs) will understand the many factors that interact to influence SRC prevention and the implications of these interactions for effective SRC education. Concussion is a complex injury that is challenging to identify and manage, particularly when athletes fail to disclose symptoms to their health care providers. Education is 1 strategy for increasing disclosure. However, limited information addresses how ATs can integrate the many factors that may influence the effectiveness of SRC education into their specific settings. Public health models provide an example through the SEF, which highlights the interplay among various levels of society and sport that can facilitate SRC prevention strategies, including education. For ATs to develop appropriate SRC prevention strategies, a framework for application is needed. A growing body of information concerning SRC prevention indicates that knowledge alone is insufficient to change concussion-related behaviors. The SEF allows this information to be considered at levels such as policy and societal, community, interpersonal (relationships), and intrapersonal (athlete). The use of such a framework will facilitate more comprehensive SRC prevention efforts that can be applied in all athletic training practice settings. Clinical Applications: Athletic trainers can use this information as they plan SRC prevention strategies in their specific settings. This approach will aid in addressing the layers of complexity that exist when developing a concussion-management policy and plan.

Short-term residential care for dementia patients: predictors for utilization and expected quality from a family caregiver's point of view.

PubMed

Donath, Carolin; Winkler, Angelika; Grässel, Elmar

2009-08-01

Short-term residential care (SRC) has proved to be effective in reducing the burden on family caregivers of dementia patients. Nevertheless, little is known about the factors which influence its usage or the expectations of family caregivers regarding quality. In this paper we address the following questions: (i) which variables of the care situation, the caregivers and their attitudes act as predictors for the utilization of SRC facilities? (ii) What are the views of caregivers about the quality of SRC? The cross-sectional study was carried out as an anonymous written survey of family caregivers of dementia patients in four regions of Germany. With a 20% response it was possible to analyze the quantitative and qualitative data from 404 and 254 family caregivers respectively. Predictors for utilization were evaluated using binary logistic regression analysis. The answers to questions of quality were evaluated using qualitative content analysis. Significant predictors for the utilization of SRC are the assessment of the helpfulness of SRC and the caregiver's knowledge of the accessibility of SRC facilities. Family caregivers who had already used SRC most frequently expressed the wish for "good care" in SRC facilities, followed by a program of suitable activities for dementia patients. In order to increase the rate of utilization, family caregivers must be convinced of the relevant advantages of using SRC facilities. The staff should be trained in caring for dementia patients and appropriate activities should be available.

Steroid receptor coactivator-3 regulates glucose metabolism in bladder cancer cells through coactivation of hypoxia inducible factor 1α.

PubMed

Zhao, Wei; Chang, Cunjie; Cui, Yangyan; Zhao, Xiaozhi; Yang, Jun; Shen, Lan; Zhou, Ji; Hou, Zhibo; Zhang, Zhen; Ye, Changxiao; Hasenmayer, Donald; Perkins, Robert; Huang, Xiaojing; Yao, Xin; Yu, Like; Huang, Ruimin; Zhang, Dianzheng; Guo, Hongqian; Yan, Jun

2014-04-18

Cancer cell proliferation is a metabolically demanding process, requiring high glycolysis, which is known as "Warburg effect," to support anabolic growth. Steroid receptor coactivator-3 (SRC-3), a steroid receptor coactivator, is overexpressed and/or amplified in multiple cancer types, including non-steroid targeted cancers, such as urinary bladder cancer (UBC). However, whether SRC-3 regulates the metabolic reprogramming for cancer cell growth is unknown. Here, we reported that overexpression of SRC-3 accelerated UBC cell growth, accompanied by the increased expression of genes involved in glycolysis. Knockdown of SRC-3 reduced the UBC cell glycolytic rate under hypoxia, decreased tumor growth in nude mice, with reduction of proliferating cell nuclear antigen and lactate dehydrogenase expression levels. We further revealed that SRC-3 could interact with hypoxia inducible factor 1α (HIF1α), which is a key transcription factor required for glycolysis, and coactivate its transcriptional activity. SRC-3 was recruited to the promoters of HIF1α-target genes, such as glut1 and pgk1. The positive correlation of expression levels between SRC-3 and Glut1 proteins was demonstrated in human UBC patient samples. Inhibition of glycolysis through targeting HK2 or LDHA decelerated SRC-3 overexpression-induced cell growth. In summary, overexpression of SRC-3 promoted glycolysis in bladder cancer cells through HIF1α to facilitate tumorigenesis, which may be an intriguing drug target for bladder cancer therapy.

Rhynchophylline Ameliorates Endothelial Dysfunction via Src-PI3K/Akt-eNOS Cascade in the Cultured Intrarenal Arteries of Spontaneous Hypertensive Rats

PubMed Central

Hao, Hui-Feng; Liu, Li-Mei; Pan, Chun-Shui; Wang, Chuan-She; Gao, Yuan-Sheng; Fan, Jing-Yu; Han, Jing-Yan

2017-01-01

Objectives: To examine the protective effect of Rhynchophylline (Rhy) on vascular endothelial function in spontaneous hypertensive rats (SHRs) and the underlying mechanism. Methods: Intrarenal arteries of SHRs and Wistar rats were suspended in myograph for force measurement. Expression and phosphorylation of endothelial nitric oxide (NO) synthase (eNOS), Akt, and Src kinase (Src) were examined by Western blotting. NO production was assayed by ELISA. Results: Rhy time- and concentration-dependently improved endothelium-dependent relaxation in the renal arteries from SHRs, but had no effect on endothelium-independent relaxation in SHR renal arteries. Wortmannin (an inhibitor of phosphatidylinositol 3-kinase) or PP2 (an inhibitor of Src) inhibited the improvement of relaxation in response to acetylcholine by 12 h-incubation with 300 μM Rhy. Western blot analysis revealed that Rhy elevated phosphorylations of eNOS, Akt, and Src in SHR renal arteries. Moreover, wortmannin reversed the increased phosphorylations of Akt and eNOS induced by Rhy, but did not affect the phosphorylation of Src. Furthermore, the enhanced phosphorylations of eNOS, Akt, and Src were blunted by PP2. Importantly, Rhy increased NO production and this effect was blocked by inhibition of Src or PI3K/Akt. Conclusion: The present study provides evidences for the first time that Rhy ameliorates endothelial dysfunction in SHRs through the activation of Src-PI3K/Akt-eNOS signaling pathway. PMID:29187825

Histochemical assessment for osteoblastic activity coupled with dysfunctional osteoclasts in c-src deficient mice.

PubMed

Toray, Hisashi; Hasegawa, Tomoka; Sakagami, Naoko; Tsuchiya, Erika; Kudo, Ai; Zhao, Shen; Moritani, Yasuhito; Abe, Miki; Yoshida, Taiji; Yamamoto, Tomomaya; Yamamoto, Tsuneyuki; Oda, Kimimitsu; Udagawa, Nobuyuki; Luiz de Freitas, Paulo Henrique; Li, Minqi

2017-01-01

Since osteoblastic activities are believed to be coupled with osteoclasts, we have attempted to histologically verify which of the distinct cellular circumstances, the presence of osteoclasts themselves or bone resorption by osteoclasts, is essential for coupled osteoblastic activity, by examining c-fos -/- or c-src -/- mice. Osteopetrotic c-fos deficient (c-fos -/- ) mice have no osteoclasts, while c-src deficient (c-src -/- ) mice, another osteopetrotic model, develop dysfunctional osteoclasts due to a lack of ruffled borders. c-fos -/- mice possessed no tartrate-resistant acid phosphatase (TRAPase)-reactive osteoclasts, and showed very weak tissue nonspecific alkaline phosphatase (TNALPase)-reactive mature osteoblasts. In contrast, c-src -/- mice had many TNALPase-positive osteoblasts and TRAPase-reactive osteoclasts. Interestingly, the parallel layers of TRAPase-reactive/osteopontin-positive cement lines were observed in the superficial region of c-src -/- bone matrix. This indicates the possibility that in c-src -/- mice, osteoblasts were activated to deposit new bone matrices on the surfaces that osteoclasts previously passed along, even without bone resorption. Transmission electron microscopy demonstrated cell-to-cell contacts between mature osteoblasts and neighboring ruffled border-less osteoclasts, and osteoid including many mineralized nodules in c-src -/- mice. Thus, it seems likely that osteoblastic activities would be maintained in the presence of osteoclasts, even if they are dysfunctional.

v-src induces clonal sarcomas and rapid metastasis following transduction with a replication-defective retrovirus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoker, A.W.; Sieweke, M.H.

1989-12-01

v-src is an effective carcinogen when expressed from Rous sarcoma virus (RSV) in vivo. Whereas RSV tumors require sustained oncogene expression, their growth is largely a balance between viral recruitment of tissues and host immune destruction of infected cells. The authors have therefore examined the tumorigenic potential of v-src in the absence of viral recruitment and viral antigen expression. v-src was introduced with high efficiency into chicken wing web tissues using replication-defective (rd) retroviral vectors. Clonal sarcomas were induced rapidly, and furthermore, v-src potentiated metastatic progression in {approx} 0.1%-1% of tumor clones with unexpectedly short latency. rd vectors proved effectivemore » not only in transducing v-src into tissues but also as insertional markers of tumor clonality. The rd vector present in most primary and metastatic tumors was a highly truncated form of RSV derived by viral transmission of spliced v-src mRNA; this vector should thus avoid viral recruitment and host anti-viral immune reaction through its complete lack of viral structural genes. Under such conditions v-src maintains strong carcinogenicity in vivo when restricted to clonal tumor growth and can confer rapid metastatic potential on a discrete subset of tumor clones.« less

Two Sides of the Same Coin: The Positive and Negative Impact of Spiritual Religious Coping on Quality of Life and Depression in Dialysis Patients.

PubMed

Vitorino, Luciano Magalhães; Soares, Renata de Castro E Santos; Santos, Ana Eliza Oliveira; Lucchetti, Alessandra Lamas Granero; Cruz, Jonas Preposi; Cortez, Paulo José Oliveira; Lucchetti, Giancarlo

2017-08-01

Studies have shown that spiritual/religious beliefs are associated with mental health and health-related quality of life (HRQoL). However, few studies evaluated how spiritual/religious coping (SRC) could affect hemodialysis patients. The present study investigated the role of SRC behaviors on HRQoL and depressive symptoms in hemodialysis patients. This was cross-sectional study with 184 patients. Patients completed the Beck Depression Inventory, Brief SRC Scale, Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36), and a Sociodemographic and Health Characterization Questionnaire. From 218 patients, 184 (84.4%) were included (53.8% male with a median age of 55.9 years). Negative SRC, but not positive SRC, was associated with depressive symptoms. Positive SRC presented significant effects in SF-36 pain and physical and social functioning. On the other hand, negative SRC exhibited significant effects in SF-36 role emotional, energy/fatigue, pain, and physical functioning. SRC influences the mental health and HRQoL in Brazilian hemodialysis patients in two distinct ways. If used positively, it may have positive outcomes. However, if used negatively, it may lead to dysfunctional consequences such as greater depressive symptomatology and affect HRQoL. Health professionals must be aware of these "two sides of the same coin."

Overview of refinement procedures within REFMAC5: utilizing data from different sources.

PubMed

Kovalevskiy, Oleg; Nicholls, Robert A; Long, Fei; Carlon, Azzurra; Murshudov, Garib N

2018-03-01

Refinement is a process that involves bringing into agreement the structural model, available prior knowledge and experimental data. To achieve this, the refinement procedure optimizes a posterior conditional probability distribution of model parameters, including atomic coordinates, atomic displacement parameters (B factors), scale factors, parameters of the solvent model and twin fractions in the case of twinned crystals, given observed data such as observed amplitudes or intensities of structure factors. A library of chemical restraints is typically used to ensure consistency between the model and the prior knowledge of stereochemistry. If the observation-to-parameter ratio is small, for example when diffraction data only extend to low resolution, the Bayesian framework implemented in REFMAC5 uses external restraints to inject additional information extracted from structures of homologous proteins, prior knowledge about secondary-structure formation and even data obtained using different experimental methods, for example NMR. The refinement procedure also generates the `best' weighted electron-density maps, which are useful for further model (re)building. Here, the refinement of macromolecular structures using REFMAC5 and related tools distributed as part of the CCP4 suite is discussed.

SLAP displays tumour suppressor functions in colorectal cancer via destabilization of the SRC substrate EPHA2

NASA Astrophysics Data System (ADS)

Naudin, Cécile; Sirvent, Audrey; Leroy, Cédric; Larive, Romain; Simon, Valérie; Pannequin, Julie; Bourgaux, Jean-François; Pierre, Josiane; Robert, Bruno; Hollande, Frédéric; Roche, Serge

2014-01-01

The adaptor SLAP is a negative regulator of receptor signalling in immune cells but its role in human cancer is ill defined. Here we report that SLAP is abundantly expressed in healthy epithelial intestine but strongly downregulated in 50% of colorectal cancer. SLAP overexpression suppresses cell tumorigenicity and invasiveness while SLAP silencing enhances these transforming properties. Mechanistically, SLAP controls SRC/EPHA2/AKT signalling via destabilization of the SRC substrate and receptor tyrosine kinase EPHA2. This activity is independent from CBL but requires SLAP SH3 interaction with the ubiquitination factor UBE4A and SLAP SH2 interaction with pTyr594-EPHA2. SRC phosphorylates EPHA2 on Tyr594, thus creating a feedback loop that promotes EPHA2 destruction and thereby self-regulates its transforming potential. SLAP silencing enhances SRC oncogenicity and sensitizes colorectal tumour cells to SRC inhibitors. Collectively, these data establish a tumour-suppressive role for SLAP in colorectal cancer and a mechanism of SRC oncogenic induction through stabilization of its cognate substrates.

Lipid binding by the Unique and SH3 domains of c-Src suggests a new regulatory mechanism

PubMed Central

Pérez, Yolanda; Maffei, Mariano; Igea, Ana; Amata, Irene; Gairí, Margarida; Nebreda, Angel R.; Bernadó, Pau; Pons, Miquel

2013-01-01

c-Src is a non-receptor tyrosine kinase involved in numerous signal transduction pathways. The kinase, SH3 and SH2 domains of c-Src are attached to the membrane-anchoring SH4 domain through the flexible Unique domain. Here we show intra- and intermolecular interactions involving the Unique and SH3 domains suggesting the presence of a previously unrecognized additional regulation layer in c-Src. We have characterized lipid binding by the Unique and SH3 domains, their intramolecular interaction and its allosteric modulation by a SH3-binding peptide or by Calcium-loaded calmodulin binding to the Unique domain. We also show reduced lipid binding following phosphorylation at conserved sites of the Unique domain. Finally, we show that injection of full-length c-Src with mutations that abolish lipid binding by the Unique domain causes a strong in vivo phenotype distinct from that of wild-type c-Src in a Xenopus oocyte model system, confirming the functional role of the Unique domain in c-Src regulation. PMID:23416516

An efficient and target-oriented sample enrichment method for preparative separation of minor alkaloids by pH-zone-refining counter-current chromatography.

PubMed

Feng, Rui-Hong; Hou, Jin-Jun; Zhang, Yi-Bei; Pan, Hui-Qin; Yang, Wenzhi; Qi, Peng; Yao, Shuai; Cai, Lu-Ying; Yang, Min; Jiang, Bao-Hong; Liu, Xuan; Wu, Wan-Ying; Guo, De-An

2015-08-28

An efficient and target-oriented sample enrichment method was established to increase the content of the minor alkaloids in crude extract by using the corresponding two-phase solvent system applied in pH-zone-refining counter-current chromatography. The enrichment and separation of seven minor indole alkaloids from Uncaria rhynchophylla (Miq.) Miq. ex Havil(UR) were selected as an example to show the advantage of this method. An optimized two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (3:7:1:9, v/v) was used in this study, where triethylamine (TEA) as the retainer and hydrochloric acid (HCl) as the eluter were added at the equimolar of 10mM. Crude alkaloids of UR dissolved in the corresponding upper phase (containing 10mM TEA) were extracted twice with lower phase (containing 10mM TEA) and lower phase (containing 10mM HCl), respectively, the second lower phase extract was subjected to pH-zone-refining CCC separation after alkalization and desalination. Finally, from 10g of crude alkaloids, 4g of refined alkaloids was obtained and the total content of seven target indole alkaloids was increased from 4.64% to 15.78%. Seven indole alkaloids, including 54mg isocorynoxeine, 21mg corynoxeine, 46mg isorhynchophylline, 35mg rhynchophylline, 65mg hirsutine, 51mg hirsuteine and 27mg geissoschizine methylether were all simultaneously separated from 2.5g of refined alkaloids, with the purity of 86.4%, 97.5%, 90.3%, 92.1%, 98.5%, 92.3%, and 92.8%, respectively. The total content and purities of the seven minor indole alkaloids were tested by HPLC and their chemical structures were elucidated by ESI-HRMS and (1)H NMR. Copyright © 2015 Elsevier B.V. All rights reserved.

The imaging performance of the SRC on Mars Express

USGS Publications Warehouse

Oberst, J.; Schwarz, G.; Behnke, T.; Hoffmann, H.; Matz, K.-D.; Flohrer, J.; Hirsch, H.; Roatsch, T.; Scholten, F.; Hauber, E.; Brinkmann, B.; Jaumann, R.; Williams, D.; Kirk, R.; Duxbury, T.; Leu, C.; Neukum, G.

2008-01-01

The Mars Express spacecraft carries the pushbroom scanner high-resolution stereo camera (HRSC) and its added imaging subsystem super resolution channel (SRC). The SRC is equipped with its own optical system and a 1024??1024 framing sensor. SRC produces snapshots with 2.3 m ground pixel size from the nominal spacecraft pericenter height of 250 km, which are typically embedded in the central part of the large HRSC scenes. The salient features of the SRC are its light-weight optics, a reliable CCD detector, and high-speed read-out electronics. The quality and effective visibility of details in the SRC images unfortunately falls short of what has been expected. In cases where thermal balance cannot be reached, artifacts, such as blurring and "ghost features" are observed in the images. In addition, images show large numbers of blemish pixels and are plagued by electronic noise. As a consequence, we have developed various image improving algorithms, which are discussed in this paper. While results are encouraging, further studies of image restoration by dedicated processing appear worthwhile. The SRC has obtained more than 6940 images at the time of writing (1 September 2007), which often show fascinating details in surface morphology. SRC images are highly useful for a variety of applications in planetary geology, for studies of the Mars atmosphere, and for astrometric observations of the Martian satellites. This paper will give a full account of the design philosophy, technical concept, calibration, operation, integration with HRSC, and performance, as well as science accomplishments of the SRC. ?? 2007 Elsevier Ltd. All rights reserved.

Combinative application of pH-zone-refining and conventional high-speed counter-current chromatography for preparative separation of caged polyprenylated xanthones from gamboge.

PubMed

Xu, Min; Fu, Wenwei; Zhang, Baojun; Tan, Hongsheng; Xiu, Yanfeng; Xu, Hongxi

2016-02-01

An efficient method for the preparative separation of four structurally similar caged xanthones from the crude extracts of gamboge was established, which involves the combination of pH-zone-refining counter-current chromatography and conventional high-speed counter-current chromatography for the first time. pH-zone-refining counter-current chromatography was performed with the solvent system composed of n-hexane/ethyl acetate/methanol/water (7:3:8:2, v/v/v/v), where 0.1% trifluoroacetic acid was added to the upper organic stationary phase as a retainer and 0.03% triethylamine was added to the aqueous mobile phase as an eluter. From 3.157 g of the crude extract, 1.134 g of gambogic acid, 180.5 mg of gambogenic acid and 572.9 mg of a mixture of two other caged polyprenylated xanthones were obtained. The mixture was further separated by conventional high-speed counter-current chromatography with a solvent system composed of n-hexane/ethyl acetate/methanol/water (5:5:10:5, v/v/v/v) and n-hexane/methyl tert-butyl ether/acetonitrile/water (8:2:6:4,v/v/v/v), yielding 11.6 mg of isogambogenic acid and 10.4 mg of β-morellic acid from 218.0 mg of the mixture, respectively. The purities of all four of the compounds were over 95%, as determined by high-performance liquid chromatography, and the chemical structures of the four compounds were confirmed by electrospray ionization mass spectrometry and NMR spectroscopy. The combinative application of pH-zone-refining counter-current chromatography and conventional high-speed counter-current chromatography shows great advantages in isolating and enriching the caged polyprenylated xanthones. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Homogeneous catalytic hydrogenations of complex carbonaceous substrates. [16 references

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cox, J L; Wilcox, W A; Roberts, G L

1976-11-05

Results of homogeneous catalytic hydrogenation of complex unsaturated substrates including coal and coal-derived materials are reported, with organic soluble molecular complexes as catalysts. Among the substrates used were Hvab coal, solvent-refined coal, and COED pyrolysate. The hydrogenations were carried out in an autoclave. The results are summarized in tables.

Interactions between relay helix and Src homology 1 domain helix (SH1) drive the converter domain rotation during the recovery stroke of myosin II

PubMed Central

Baumketner, Andrij

2012-01-01

Myosin motor protein exists in two alternative conformations, pre-recovery state M* and post-recovery state M**, upon ATP binding. The details of the M*-to-M** transition, known as the recovery stroke to reflect its role as the functional opposite of the force-generating power stroke, remain elusive. The defining feature of the post-recovery state is a kink in the relay helix, a key part of the protein involved in force generation. In this paper we determine the interactions that are responsible for the appearance of the kink. We design a series of computational models that contain three other segments, relay loop, converter domain and Src homology 1 domain helix (SH1), with which relay helix interacts, and determine their structure in accurate replica exchange molecular dynamics simulations in explicit solvent. By conducting an exhaustive combinatorial search among different models we find that: 1) the converter domain must be attached to the relay helix during the transition, so it does not interfere with other parts of the protein, 2) the structure of the relay helix is controlled by SH1 helix. The kink is strongly coupled to the position of SH1 helix. It arises as a result of direct interactions between SH1 and the relay helix and leads to a rotation of the C-terminal part of the relay helix which is subsequently transmitted to the converter domain. PMID:22411190

A novel disulfide bond in the SH2 Domain of the C-terminal Src kinase controls catalytic activity.

PubMed

Mills, Jamie E; Whitford, Paul C; Shaffer, Jennifer; Onuchic, Jose N; Adams, Joseph A; Jennings, Patricia A

2007-02-02

The SH2 domain of the C-terminal Src kinase [Csk] contains a unique disulfide bond that is not present in other known SH2 domains. To investigate whether this unusual disulfide bond serves a novel function, the effects of disulfide bond formation on catalytic activity of the full-length protein and on the structure of the SH2 domain were investigated. The kinase activity of full-length Csk decreases by an order of magnitude upon formation of the disulfide bond in the distal SH2 domain. NMR spectra of the fully oxidized and fully reduced SH2 domains exhibit similar chemical shift patterns and are indicative of similar, well-defined tertiary structures. The solvent-accessible disulfide bond in the isolated SH2 domain is highly stable and far from the small lobe of the kinase domain. However, reduction of this bond results in chemical shift changes of resonances that map to a cluster of residues that extend from the disulfide bond across the molecule to a surface that is in direct contact with the small lobe of the kinase domain in the intact molecule. Normal mode analyses and molecular dynamics calculations suggest that disulfide bond formation has large effects on residues within the kinase domain, most notably within the active-site cleft. Overall, the data indicate that reversible cross-linking of two cysteine residues in the SH2 domain greatly impacts catalytic function and interdomain communication in Csk.

Src kinases in chondrosarcoma chemoresistance and migration: dasatinib sensitises to doxorubicin in TP53 mutant cells

PubMed Central

van Oosterwijk, J G; van Ruler, M A J H; Briaire-de Bruijn, I H; Herpers, B; Gelderblom, H; van de Water, B; Bovée, J V M G

2013-01-01

Background: Chondrosarcomas are malignant cartilage-forming tumours of bone. Because of their resistance to conventional chemotherapy and radiotherapy, currently no treatment strategies exist for unresectable and metastatic chondrosarcoma. Previously, PI3K/AKT/GSK3β and Src kinase pathways were shown to be activated in chondrosarcoma cell lines. Our aim was to investigate the role of these kinases in chemoresistance and migration in chondrosarcoma in relation to TP53 mutation status. Methods: We used five conventional and three dedifferentiated chondrosarcoma cell lines and investigated the effect of PI3K/AKT/GSK3β pathway inhibition (enzastaurin) and Src pathway inhibition (dasatinib) in chemoresistance using WST assay and live cell imaging with AnnexinV staining. Immunohistochemistry on tissue microarrays (TMAs) containing 157 cartilaginous tumours was performed for Src family members. Migration assays were performed with the RTCA xCelligence System. Results: Src inhibition was found to overcome chemoresistance, to induce apoptosis and to inhibit migration. Cell lines with TP53 mutations responded better to combination therapy than wild-type cell lines (P=0.002). Tissue microarray immunohistochemistry confirmed active Src (pSrc) signalling, with Fyn being most abundantly expressed (76.1%). Conclusion: These results strongly indicate Src family kinases, in particular Fyn, as a potential target for the treatment of inoperable and metastatic chondrosarcomas, and to sensitise for doxorubicin especially in the presence of TP53 mutations. PMID:23922104

An epitope localized in c-Src negative regulatory domain is a potential marker in early stage of colonic neoplasms.

PubMed

Sakai, T; Kawakatsu, H; Fujita, M; Yano, J; Owada, M K

1998-02-01

In previous work, we established a new monoclonal antibody that specifically recognizes the active form of c-Src tyrosine kinase (Kawakatsu et al, 1996). To determine whether c-Src is active in colorectal tumorigenesis, we examined the expression of an active form of c-Src in human normal mucosa, hyperplastic polyps, adenomas, and adenocarcinomas. The tissue distribution of the active form of c-Src was studied by immunohistochemistry using this antibody, termed Clone 28. Among 66 cases of adenoma tested, 61 (92%) showed positive staining (adenoma with mild atypia, 3 of 3; adenoma with moderate atypia, 38 of 42; adenoma with severe atypia, 20 of 21). In contrast to the frequent and intense staining in adenomas, adenocarcinoma showed weak staining with less frequency in 4 of 16 (25%) cases. The number of specimens with positive staining in well- and moderately differentiated adenocarcinomas was limited to an early stage. The active form of c-Src mainly localized to the nuclear membrane and the perinuclear region. These results provide the first direct evidence that the activation of c-Src appears to be an early event in colonic carcinogenesis in situ. The findings of the present study thus allow us to propose a molecular mechanism involving c-Src activation in the process of malignant transformation of the human colonic neoplastic cells.

Interfacing with USSTRATCOM and UTTR during Stardust Earth Return

NASA Technical Reports Server (NTRS)

Jefferson, David C.; Baird, Darren T.; Cangahuala, Laureano A.; Lewis, George D.

2006-01-01

The Stardust Sample Return Capsule separated from the main spacecraft four hours prior to atmospheric entry. Between this time and the time at which the SRC touched down at the Utah Test and Training Range, two organizations external to JPL were involved in tracking the Sample Return Capsule. Orbit determination for the Stardust spacecraft during deep space cruise, the encounters of asteroid Annefrank and comet Wild 2, and the final approach to Earth used X-band radio metric Doppler and range data obtained through the Deep Space Network. The SRC lacked the electronics needed for coherently transponded radio metric tracking, so the DSN was not able to track the SRC after it separated from the main spacecraft. Although the expected delivery accuracy at atmospheric entry was well within the capability needed to target the SRC to the desired ground location, it was still desirable to obtain direct knowledge of the SRC trajectory in case of anomalies. For this reason U.S. Strategic Command was engaged to track the SRC between separation and atmospheric entry. Once the SRC entered the atmosphere, ground sensors at UTTR were tasked to acquire the descending SRC and maintain track during the descent in order to determine the landing location, to which the ground recovery team was then directed. This paper discusses organizational interfaces, data products, and delivery schedules, and the actual tracking operations are described.

Process for preparing lubricating oil from used waste lubricating oil

DOEpatents

Whisman, Marvin L.; Reynolds, James W.; Goetzinger, John W.; Cotton, Faye O.

1978-01-01

A re-refining process is described by which high-quality finished lubricating oils are prepared from used waste lubricating and crankcase oils. The used oils are stripped of water and low-boiling contaminants by vacuum distillation and then dissolved in a solvent of 1-butanol, 2-propanol and methylethyl ketone, which precipitates a sludge containing most of the solid and liquid contaminants, unspent additives, and oxidation products present in the used oil. After separating the purified oil-solvent mixture from the sludge and recovering the solvent for recycling, the purified oil is preferably fractional vacuum-distilled, forming lubricating oil distillate fractions which are then decolorized and deodorized to prepare blending stocks. The blending stocks are blended to obtain a lubricating oil base of appropriate viscosity before being mixed with an appropriate additive package to form the finished lubricating oil product.

PLC-γ directly binds activated c-Src, which is necessary for carbachol-mediated inhibition of NHE3 activity in Caco-2/BBe cells

PubMed Central

Lee, Luke J.; Kovbasnjuk, Olga; Li, Xuhang; Donowitz, Mark

2013-01-01

Elevated levels of intracellular Ca2+ ([Ca2+]i) inhibit Na+/H+ exchanger 3 (NHE3) activity in the intact intestine. We previously demonstrated that PLC-γ directly binds NHE3, an interaction that is necessary for [Ca2+]i inhibition of NHE3 activity, and that PLC-γ Src homology 2 (SH2) domains may scaffold Ca2+ signaling proteins necessary for regulation of NHE3 activity. [Ca2+]i regulation of NHE3 activity is also c-Src dependent; however, the mechanism by which c-Src is involved is undetermined. We hypothesized that the SH2 domains of PLC-γ might link c-Src to NHE3-containing complexes to mediate [Ca2+]i inhibition of NHE3 activity. In Caco-2/BBe cells, carbachol (CCh) decreased NHE3 activity by ∼40%, an effect abolished with the c-Src inhibitor PP2. CCh treatment increased the amount of active c-Src as early as 1 min through increased Y416 phosphorylation. Coimmunoprecipitation demonstrated that c-Src associated with PLC-γ, but not NHE3, under basal conditions, an interaction that increased rapidly after CCh treatment and occurred before the dissociation of PLC-γ and NHE3 that occurred 10 min after CCh treatment. Finally, direct binding to c-Src only occurred through the PLC-γ SH2 domains, an interaction that was prevented by blocking the PLC-γ SH2 domain. This study demonstrated that c-Src 1) activity is necessary for [Ca2+]i inhibition of NHE3 activity, 2) activation occurs rapidly (∼1 min) after CCh treatment, 3) directly binds PLC-γ SH2 domains and associates dynamically with PLC-γ under elevated [Ca2+]i conditions, and 4) does not directly bind NHE3. Under elevated [Ca2+]i conditions, PLC-γ scaffolds c-Src into NHE3-containing multiprotein complexes before dissociation of PLC-γ from NHE3 and subsequent endocytosis of NHE3. PMID:23703528

Targeting Drug-Sensitive and -Resistant Strains of Mycobacterium tuberculosis by Inhibition of Src Family Kinases Lowers Disease Burden and Pathology.

PubMed

Chandra, Pallavi; Rajmani, R S; Verma, Garima; Bhavesh, Neel Sarovar; Kumar, Dhiraj

2016-01-01

In view of emerging drug resistance among bacterial pathogens, including Mycobacterium tuberculosis, the development of novel therapeutic strategies is increasingly being sought. A recent paradigm in antituberculosis (anti-TB) drug development is to target the host molecules that are crucial for intracellular survival of the pathogen. We previously showed the importance of Src tyrosine kinases in mycobacterial pathogenesis. Here, we report that inhibition of Src significantly reduced survival of H37Rv as well as multidrug-resistant (MDR) and extremely drug-resistant (XDR) strains of M. tuberculosis in THP-1 macrophages. Src inhibition was also effective in controlling M. tuberculosis infection in guinea pigs. In guinea pigs, reduced M. tuberculosis burden due to Src inhibition also led to a marked decline in the disease pathology. In agreement with the theoretical framework of host-directed approaches against the pathogen, Src inhibition was equally effective against an XDR strain in controlling infection in guinea pigs. We propose that Src inhibitors could be developed into effective host-directed anti-TB drugs, which could be indiscriminately used against both drug-sensitive and drug-resistant strains of M. tuberculosis. IMPORTANCE The existing treatment regimen for tuberculosis (TB) suffers from deficiencies like high doses of antibiotics, long treatment duration, and inability to kill persistent populations in an efficient manner. Together, these contribute to the emergence of drug-resistant tuberculosis. Recently, several host factors were identified which help intracellular survival of Mycobacterium tuberculosis within the macrophage. These factors serve as attractive targets for developing alternate therapeutic strategies against M. tuberculosis. This strategy promises to be effective against drug-resistant strains. The approach also has potential to considerably lower the risk of emergence of new drug-resistant strains. We explored tyrosine kinase Src as a host factor exploited by virulent M. tuberculosis for intracellular survival. We show that Src inhibition can effectively control tuberculosis in infected guinea pigs. Moreover, Src inhibition ameliorated TB-associated pathology in guinea pigs. Thus, Src inhibitors have strong potential to be developed as possible anti-TB drugs.

Subject-specific increases in serum S-100B distinguish sports-related concussion from sports-related exertion.

PubMed

Kiechle, Karin; Bazarian, Jeffrey J; Merchant-Borna, Kian; Stoecklein, Veit; Rozen, Eric; Blyth, Brian; Huang, Jason H; Dayawansa, Samantha; Kanz, Karl; Biberthaler, Peter

2014-01-01

The on-field diagnosis of sports-related concussion (SRC) is complicated by the lack of an accurate and objective marker of brain injury. To compare subject-specific changes in the astroglial protein, S100B, before and after SRC among collegiate and semi-professional contact sport athletes, and compare these changes to differences in S100B before and after non-contact exertion. Longitudinal cohort study. From 2009-2011, we performed a prospective study of athletes from Munich, Germany, and Rochester, New York, USA. Serum S100B was measured in all SRC athletes at pre-season baseline, within 3 hours of injury, and at days 2, 3 and 7 post-SRC. Among a subset of athletes, S100B was measured after non-contact exertion but before injury. All samples were collected identically and analyzed using an automated electrochemiluminescent assay to quantify serum S100B levels. Forty-six athletes (30 Munich, 16 Rochester) underwent baseline testing. Thirty underwent additional post-exertion S100B testing. Twenty-two athletes (16 Rochester, 6 Munich) sustained a SRC, and 17 had S100B testing within 3 hours post-injury. The mean 3-hour post-SRC S100B was significantly higher than pre-season baseline (0.099±0.008 µg/L vs. 0.058±0.006 µg/L, p = 0.0002). Mean post-exertion S100B was not significantly different than the preseason baseline. S100B levels at post-injury days 2, 3 and 7 were significantly lower than the 3-hour level, and not different than baseline. Both the absolute change and proportional increase in S100B 3-hour post-injury were accurate discriminators of SRC from non-contact exertion without SRC (AUC 0.772 and 0.904, respectively). A 3-hour post-concussion S100B >0.122 µg/L and a proportional S100B increase of >45.9% over baseline were both 96.7% specific for SRC. Relative and absolute increases in serum S100B can accurately distinguish SRC from sports-related exertion, and may be a useful adjunct to the diagnosis of SRC.

PLC-γ directly binds activated c-Src, which is necessary for carbachol-mediated inhibition of NHE3 activity in Caco-2/BBe cells.

PubMed

Zachos, Nicholas C; Lee, Luke J; Kovbasnjuk, Olga; Li, Xuhang; Donowitz, Mark

2013-08-01

Elevated levels of intracellular Ca(2+) ([Ca(2+)]i) inhibit Na(+)/H(+) exchanger 3 (NHE3) activity in the intact intestine. We previously demonstrated that PLC-γ directly binds NHE3, an interaction that is necessary for [Ca(2+)]i inhibition of NHE3 activity, and that PLC-γ Src homology 2 (SH2) domains may scaffold Ca(2+) signaling proteins necessary for regulation of NHE3 activity. [Ca(2+)]i regulation of NHE3 activity is also c-Src dependent; however, the mechanism by which c-Src is involved is undetermined. We hypothesized that the SH2 domains of PLC-γ might link c-Src to NHE3-containing complexes to mediate [Ca(2+)]i inhibition of NHE3 activity. In Caco-2/BBe cells, carbachol (CCh) decreased NHE3 activity by ∼40%, an effect abolished with the c-Src inhibitor PP2. CCh treatment increased the amount of active c-Src as early as 1 min through increased Y(416) phosphorylation. Coimmunoprecipitation demonstrated that c-Src associated with PLC-γ, but not NHE3, under basal conditions, an interaction that increased rapidly after CCh treatment and occurred before the dissociation of PLC-γ and NHE3 that occurred 10 min after CCh treatment. Finally, direct binding to c-Src only occurred through the PLC-γ SH2 domains, an interaction that was prevented by blocking the PLC-γ SH2 domain. This study demonstrated that c-Src 1) activity is necessary for [Ca(2+)]i inhibition of NHE3 activity, 2) activation occurs rapidly (∼1 min) after CCh treatment, 3) directly binds PLC-γ SH2 domains and associates dynamically with PLC-γ under elevated [Ca(2+)]i conditions, and 4) does not directly bind NHE3. Under elevated [Ca(2+)]i conditions, PLC-γ scaffolds c-Src into NHE3-containing multiprotein complexes before dissociation of PLC-γ from NHE3 and subsequent endocytosis of NHE3.

The American College of Surgeons National Surgical Quality Improvement Program Surgical Risk Calculator Does Not Accurately Predict Risk of 30-Day Complications Among Patients Undergoing Microvascular Head and Neck Reconstruction.

PubMed

Arce, Kevin; Moore, Eric J; Lohse, Christine M; Reiland, Matthew D; Yetzer, Jacob G; Ettinger, Kyle S

2016-09-01

The American College of Surgeons (ACS) National Surgical Quality Improvement Program (NSQIP) Surgical Risk Calculator (SRC) is a novel universal risk calculator designed to aid in risk stratification of patients undergoing various types of major surgery. The purpose of this study was to assess the validity of the ACS NSQIP SRC in predicting postoperative complications in patients undergoing microvascular head and neck reconstruction. A retrospective cohort study of patients undergoing head and neck microvascular reconstruction with fibular free flaps at a single institution was completed. The NSQIP SRC was used to compute complication risk estimates and length of stay (LOS) estimates for all patients under study. Associations between complication risk estimates generated by the SRC and actual rates of observed complications were evaluated using logistic regression models. Logistic regression models also were used to evaluate the SRC estimates for LOS duration compared with the actual observed LOS after surgery. Of 153 patients under study, 46 (30%) developed a postoperative complication corresponding to those defined by NSQIP SRC. Thirty-eight patients (25%) developed a postoperative complication categorized as severe in the parameters of the NSQIP SRC. None of the SRC complication estimates showed a statistically relevant association with the corresponding observed rates of complications. The mean LOS predicted by the SRC was 8.0 days (median, 7.5 days; interquartile range [IQR], 6.5 to 9; range, 5.0 to 18.5 days). The mean observed LOS for the study group was 9.6 days (median, 7.0 days; IQR, 6 to 9; range, 5 to 67 days). Lin's (Biometrics 45:255, 1989) concordance correlation coefficient to measure agreement between observed and predicted LOS was 0.10, indicating only slight agreement between the 2 values. The ACS NSQIP SRC is not a useful risk-stratifying metric for patients undergoing major head and neck reconstruction with microvascular fibular free flaps. The SRC also does not accurately predict hospital LOS for this same patient cohort. Copyright © 2016 American Association of Oral and Maxillofacial Surgeons. Published by Elsevier Inc. All rights reserved.

What is the definition of sports-related concussion: a systematic review.

PubMed

McCrory, Paul; Feddermann-Demont, Nina; Dvořák, Jiří; Cassidy, J David; McIntosh, Andrew; Vos, Pieter E; Echemendia, Ruben J; Meeuwisse, Willem; Tarnutzer, Alexander A

2017-06-01

Various definitions for concussion have been proposed, each having its strengths and weaknesses. We reviewed and compared current definitions and identified criteria necessary for an operational definition of sports-related concussion (SRC) in preparation of the 5th Concussion Consensus Conference (Berlin, Germany). We also assessed the role of biomechanical studies in informing an operational definition of SRC. This is a systematic literature review. Data sources include MEDLINE, Embase, Cumulative Index to Nursing and Allied Health Literature, Cochrane Central Register of Clinical Trials and SPORT Discus (accessed 14 September 2016). Eligibility criteria were studies reporting (clinical) criteria for diagnosing SRC and studies containing SRC impact data. Out of 1601 articles screened, 36 studies were included (2.2%), 14 reported on criteria for SRC definitions and 22 on biomechanical aspects of concussions. Six different operational definitions focusing on clinical findings and their dynamics were identified. Biomechanical studies were obtained almost exclusively on American football players. Angular and linear head accelerations linked to clinically confirmed concussions demonstrated considerable individual variation. SRC is a traumatic brain injury that is defined as a complex pathophysiological process affecting the brain, induced by biomechanical forces with several common features that help define its nature. Limitations identified include that the current criteria for diagnosing SRC are clinically oriented and that there is no gold/standard to assess their diagnostic properties. A future, more valid definition of SRC would better identify concussed players by demonstrating high predictive positive/negative values. Currently, the use of helmet-based systems to study the biomechanics of SRC is limited to few collision sports. New approaches need to be developed to provide objective markers for SRC. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Src is activated by the nuclear receptor peroxisome proliferator-activated receptor β/δ in ultraviolet radiation-induced skin cancer.

PubMed

Montagner, Alexandra; Delgado, Maria B; Tallichet-Blanc, Corinne; Chan, Jeremy S K; Sng, Ming K; Mottaz, Hélén; Degueurce, Gwendoline; Lippi, Yannick; Moret, Catherine; Baruchet, Michael; Antsiferova, Maria; Werner, Sabine; Hohl, Daniel; Saati, Talal Al; Farmer, Pierre J; Tan, Nguan S; Michalik, Liliane; Wahli, Walter

2014-01-01

Although non-melanoma skin cancer (NMSC) is the most common human cancer and its incidence continues to rise worldwide, the mechanisms underlying its development remain incompletely understood. Here, we unveil a cascade of events involving peroxisome proliferator-activated receptor (PPAR) β/δ and the oncogene Src, which promotes the development of ultraviolet (UV)-induced skin cancer in mice. UV-induced PPARβ/δ activity, which directly stimulated Src expression, increased Src kinase activity and enhanced the EGFR/Erk1/2 signalling pathway, resulting in increased epithelial-to-mesenchymal transition (EMT) marker expression. Consistent with these observations, PPARβ/δ-null mice developed fewer and smaller skin tumours, and a PPARβ/δ antagonist prevented UV-dependent Src stimulation. Furthermore, the expression of PPARβ/δ positively correlated with the expression of SRC and EMT markers in human skin squamous cell carcinoma (SCC), and critically, linear models applied to several human epithelial cancers revealed an interaction between PPARβ/δ and SRC and TGFβ1 transcriptional levels. Taken together, these observations motivate the future evaluation of PPARβ/δ modulators to attenuate the development of several epithelial cancers.

Transmembrane adaptor protein PAG1 is a novel tumor suppressor in neuroblastoma

PubMed Central

Agarwal, Saurabh; Ghosh, Rajib; Chen, Zaowen; Lakoma, Anna; Gunaratne, Preethi H.; Kim, Eugene S.; Shohet, Jason M.

2016-01-01

(NB) is the most common extracranial pediatric solid tumor with high mortality rates. The tyrosine kinase c-Src has been known to play an important role in differentiation of NB cells, but the mechanism of c-Src regulation has not been defined. Here, we characterize PAG1 (Cbp, Csk binding protein), a central inhibitor of c-Src and other Src family kinases, as a novel tumor suppressor in NB. Clinical cohort analysis demonstrate that low expression of PAG1 is a significant prognostic factor for high stage disease, increased relapse, and worse overall survival for children with NB. PAG1 knockdown in NB cells promotes proliferation and anchorage-independent colony formation with increased activation of AKT and ERK downstream of c-Src, while PAG1 overexpression significantly rescues these effects. In vivo, PAG1 overexpression significantly inhibits NB tumorigenicity in an orthotopic xenograft model. Our results establish PAG1 as a potent tumor suppressor in NB by inhibiting c-Src and downstream effector pathways. Thus, reactivation of PAG1 and inhibition of c-Src kinase activity represents an important novel therapeutic approach for high-risk NB. PMID:26993602

An adaptor role for cytoplasmic Sam68 in modulating Src activity during cell polarization.

PubMed

Huot, Marc-Etienne; Brown, Claire M; Lamarche-Vane, Nathalie; Richard, Stéphane

2009-04-01

The Src-associated substrate during mitosis with a molecular mass of 68 kDa (Sam68) is predominantly nuclear and is known to associate with proteins containing the Src homology 3 (SH3) and SH2 domains. Although Sam68 is a Src substrate, little is known about the signaling pathway that link them. Src is known to be activated transiently after cell spreading, where it modulates the activity of small Rho GTPases. Herein we report that Sam68-deficient cells exhibit loss of cell polarity and cell migration. Interestingly, Sam68-deficient cells exhibited sustained Src activity after cell attachment, resulting in the constitutive tyrosine phosphorylation and activation of p190RhoGAP and its association with p120rasGAP. Consistently, we observed that Sam68-deficient cells exhibited deregulated RhoA and Rac1 activity. By using total internal reflection fluorescence microscopy, we observed Sam68 near the plasma membrane after cell attachment coinciding with phosphorylation of its C-terminal tyrosines and association with Csk. These findings show that Sam68 localizes near the plasma membrane during cell attachment and serves as an adaptor protein to modulate Src activity for proper signaling to small Rho GTPases.

SRC family kinases as novel therapeutic targets to treat breast cancer brain metastases.

PubMed

Zhang, Siyuan; Huang, Wen-Chien; Zhang, Lin; Zhang, Chenyu; Lowery, Frank J; Ding, Zhaoxi; Guo, Hua; Wang, Hai; Huang, Suyun; Sahin, Aysegul A; Aldape, Kenneth D; Steeg, Patricia S; Yu, Dihua

2013-09-15

Despite better control of early-stage disease and improved overall survival of patients with breast cancer, the incidence of life-threatening brain metastases continues to increase in some of these patients. Unfortunately, other than palliative treatments there is no effective therapy for this condition. In this study, we reveal a critical role for Src activation in promoting brain metastasis in a preclinical model of breast cancer and we show how Src-targeting combinatorial regimens can treat HER2(+) brain metastases in this model. We found that Src was hyperactivated in brain-seeking breast cancer cells derived from human cell lines or from patients' brain metastases. Mechanistically, Src activation promoted tumor cell extravasation into the brain parenchyma via permeabilization of the blood-brain barrier. When combined with the EGFR/HER2 dual-targeting drug lapatinib, an Src-targeting combinatorial regimen prevented outgrowth of disseminated breast cancer cells through the induction of cell-cycle arrest. More importantly, this combinatorial regimen inhibited the outgrowth of established experimental brain metastases, prolonging the survival of metastases-bearing mice. Our results provide a rationale for clinical evaluation of Src-targeting regimens to treat patients with breast cancer suffering from brain metastasis. ©2013 AACR.

SRC: A Model of Industry-University Cooperation.

ERIC Educational Resources Information Center

Cavin, Ralph K., III; Phillips, D. Howard

1988-01-01

Describes the Semiconductor Research Corporation (SRC), a non-profit research cooperative designed to conduct research in the field of integrated circuits, principally in U.S. universities, with membership restricted to U.S.-owned companies. Analyzes SRC's impact on the U.S. educational system. (TW)

Role of SRC-3delta4 in the Progression and Metastasis of Castration-Resistant Prostate Cancer

DTIC Science & Technology

2014-12-01

tyrosine phosphorylation of SRC-3∆4, which was inhibited by the treatment with EGFR inhibitor AG1478. Mutation of Y1159 to phenylalanine (Y1159F...Y1159 to phenylalanine (Y1159F) greatly reduced SRC-3∆4/AR interaction that is stimulated by EGF. Figure 7 Overexpression of SRC-3∆4 promoted...adhesion turnover and matrix metalloproteinase expression. Cancer research 68, 5460-5468. 6. Chung, A.C., Zhou, S., Liao, L ., Tien, J.C., Greenberg

Src is a major signaling component for CTGF induction by TGF-β1 in osteoblasts

PubMed Central

X, Zhang; JA, Arnott; S, Rehman; WG, DeLong; A, Sanjay; FF, Safadi; SN, Popoff

2010-01-01

Connective tissue growth factor (CTGF/CCN2) is induced by transforming growth factor beta 1(TGF-β1) where it acts as a downstream mediator of TGF-β1 induced matrix production in osteoblasts. We have shown the requirement of Src, Erk and Smad signaling for CTGF induction by TGF-β1 in osteoblasts, however the potential interaction among these signaling pathways remains undetermined. In this study we demonstrate that TGF-β1 activates Src kinase in ROS17/2.8 cells and that treatment with the Src family kinase inhibitor PP2 prevents Src activation and CTGF induction by TGF-β1. Additionally, inhibiting Src activation prevented Erk activation, Smad 2 & 3 activation and nuclear translocation by TGF-β1, demonstrating that Src is an essential upstream signaling partner of both Erk and Smads in osteoblasts. MAPKs such as Erk can modulate the Smad pathway through directly mediating the phosphorylation of Smads or indirectly through activation/inactivation of required nuclear co-activators that mediate Smad DNA binding. When we treated cells with the Erk inhibitor, PD98059 it inhibited TGF-β1-induced CTGF protein expression but had no effect on Src activation, Smad activation or Smad nuclear translocation. However PD98059 impaired transcriptional complex formation on the Smad binding element (SBE) on the CTGF promoter, demonstrating that Erk activation was required for SBE transactivation. This data demonstrates that Src is an essential upstream signaling transducer of Erk and Smad signaling with respect to TGF-β1 in osteoblasts and that Smads and Erk function independently but are both essential for forming a transcriptionally active complex on the CTGF promoter in osteoblasts. PMID:20432467

Phosphorylation of TOPK at Y74, Y272 by Src increases the stability of TOPK and promotes tumorigenesis of colon

PubMed Central

Wang, Zhe; Yan, Wei; Sun, Huimin; Xue, Peipei; Fan, Xiaoming; Zeng, Xiaoyu; Chen, Juan; Shao, Chen; Zhu, Feng

2016-01-01

T-LAK cell-originated protein kinase (TOPK), a serine/threonine protein kinase, is highly expressed in a variety of tumors and associated with a poor prognosis of human malignancies. However, the activation mechanism of TOPK is still unrevealed. Herein, first we found that Src directly bound with and phosphorylated TOPK at Y74 and Y272 in vitro. Anti-phospho-TOPK at Y74 was prepared, the endogenous phosphorylation of TOPK at Y74 was detected in colon cancer cells, and the phosphorylation was inhibited in cells expressing low levels of Src. Subsequently, we stably transfected Y74 and Y272 double mutated TOPK (TOPK-FF) into JB6 or SW480 cells, and observed that both the anchorage-independent growth ability and tumorigenesis of TOPK-FF cells were suppressed compared with those of wild type TOPK (TOPK-WT) ex vivo and in vivo. The phosphorylation level of TOPK substrate, Histone H3 at Ser10 also decreased dramatically ex vivo or in vivo. Moreover, we showed that Src could inhibit the ubiquitination of TOPK. Transiently expressed TOPK-WT was more stable than TOPK-FF in pause and chase experiment. Endogenous TOPK was more stable in Src wild type (Src+/+) MEFs than in Src knockout (Src−/−). Taken together, our results indicate that Src is a novel upstream kinase of TOPK. The phosphorylation of TOPK at Y74 and Y272 by Src increases the stability and activity of TOPK, and promotes the tumorigenesis of colon cancer. It may provide opportunities for TOPK based prognosis and targeted therapy for colon cancer patients. PMID:27016416

Explanatory characteristics for nutrient concentrations and loads in the Sava River Catchment and cross-regionally

NASA Astrophysics Data System (ADS)

Levi, L.; Cvetkovic, V.; Destouni, G.

2015-12-01

This study compiles estimates of waterborne nutrient concentrations and loads in the Sava River Catchment (SRC). Based on this compilation, we investigate hotspots of nutrient inputs and retention along the river, as well as concentration and load correlations with river discharge and various human drivers of excess nutrient inputs to the SRC. For cross-regional assessment and possible generalization, we also compare corresponding results between the SRC and the Baltic Sea Drainage Basin (BSDB). In the SRC, one small incremental subcatchment, which is located just downstream of Zagreb and has the highest population density among the SRC subcatchments, is identified as a major hotspot for net loading (input minus retention) of both total nitrogen (TN) and total phosphorus (TP) to the river and through it to downstream areas of the SRC. The other SRC subcatchments exhibit relatively similar characteristics with smaller net nutrient loading. The annual loads of both TN and TP along the Sava River exhibit dominant temporal variability with considerably higher correlation with annual river discharge (R2 = 0.51 and 0.28, respectively) than that of annual average nutrient concentrations (R2 = 0.0 versus discharge for both TN and TP). Nutrient concentrations exhibit instead dominant spatial variability with relatively high correlation with population density among the SRC subcatchments (R2=0.43-0.64). These SRC correlation characteristics compare well with corresponding ones for the BSDB, even though the two regions are quite different in their hydroclimatic, agricultural and wastewater treatment conditions. Such cross-regional consistency in dominant variability type and explanatory catchment characteristics may be a useful generalization basis, worthy of further investigation, for at least first-order estimation of nutrient concentration and load conditions in less data-rich regions.

Atypical protein kinase C activity is required for extracellular matrix degradation and invasion by Src-transformed cells.

PubMed

Rodriguez, Elena M; Dunham, Elizabeth E; Martin, G Steven

2009-10-01

Atypical protein kinase C (aPKC) isoforms have been shown to mediate Src-dependent signaling in response to growth factor stimulation. To determine if aPKC activity contributes to the transformed phenotype of cells expressing oncogenic Src, we have examined the activity and function of aPKCs in 3T3 cells expressing viral Src (v-Src). aPKC activity and tyrosine phosphorylation were found to be elevated in some but not all clones of mouse fibroblasts expressing v-Src. aPKC activity was inhibited either by addition of a membrane-permeable pseudosubstrate, by expression of a dominant-negative aPKC, or by RNAi-mediated knockdown of specific aPKC isoforms. aPKC activity contributes to morphological transformation and stress fiber disruption, and is required for migration of Src-transformed cells and for their ability to polarize at the edge of a monolayer. The lambda isoform of aPKC is specifically required for invasion through extracellular matrix in Boyden chamber assays and for degradation of the extracellular matrix in in situ zymography assays. Tyrosine phosphorylation of aPKClambda is required for its ability to promote cell invasion. The defect in invasion upon aPKC inhibition appears to result from a defect in the assembly and/or function of podosomes, invasive adhesions on the ventral surface of the cell that are sites of protease secretion. aPKC was also found to localize to podosomes of v-Src transformed cells, suggesting a direct role for aPKC in podosome assembly and/or function. We conclude that basal or elevated aPKC activity is required for the ability of Src-transformed cells to degrade and invade the extracellular matrix. Copyright 2009 Wiley-Liss, Inc.

1α,25(OH)2-Vitamin D3 Inhibits C2C12 Cell Differentiation by Activating c-Src and ERK1/2.

PubMed

Wang, Zhonghua; Jiang, Aijun; Mei, Jingwei; Zhang, Xinyan

2018-05-01

The steroid hormone 1α,25(OH)2-vitamin D3 (1,25-D3) induced some biological responses through activation of MAPK cascades in various cell types. It seems that 1,25-D3 plays different roles at different stages of proliferating, differentiating, and differentiated C2C12 cells. We wanted to detect the effect of 1,25-D3 on myogenic differentiation and the role of ERK1/2 in differentiating stage induced by 2% horse serum with 1,25-D3. In this study, cells were induced to differentiate with 2% horse serum until the 7th day (with addition of 1,25-D3 every two days). The protein level of MHC (myosin heavy chain) and phosphorylation level of Src and ERK1/2 were determined with western blot. U0126 (MEK inhibitor) and PP2 (Src specific inhibitor) were used to confirm the relationship between 1,25-D3, MHC, Src, and ERK1/2. 1,25-D3 inhibited differentiation of C2C12 cells and fusion of myotubes by phosphorylating and activating Src and ERK1/2. Phosphorylation of ERK1/2 was inhibited, not only by U0126 but also by PP2 (a Src specific inhibitor) which led to the promotion of differentiation of C2C12 cells; however, U0126 did not inhibit Src phosphorylation. These results suggested that 1,25-D3 possibly inhibited C2C12 differentiation through Src and ERK1/2, and Src played an upstream role in this signaling pathway.

Steroid receptor coactivators 1 and 2 mediate fetal-to-maternal signaling that initiates parturition

PubMed Central

Gao, Lu; Rabbitt, Elizabeth H.; Condon, Jennifer C.; Renthal, Nora E.; Johnston, John M.; Mitsche, Matthew A.; Chambon, Pierre; Xu, Jianming; O’Malley, Bert W.; Mendelson, Carole R.

2015-01-01

The precise mechanisms that lead to parturition are incompletely defined. Surfactant protein-A (SP-A), which is secreted by fetal lungs into amniotic fluid (AF) near term, likely provides a signal for parturition; however, SP-A–deficient mice have only a relatively modest delay (~12 hours) in parturition, suggesting additional factors. Here, we evaluated the contribution of steroid receptor coactivators 1 and 2 (SRC-1 and SRC-2), which upregulate SP-A transcription, to the parturition process. As mice lacking both SRC-1 and SRC-2 die at birth due to respiratory distress, we crossed double-heterozygous males and females. Parturition was severely delayed (~38 hours) in heterozygous dams harboring SRC-1/-2–deficient embryos. These mothers exhibited decreased myometrial NF-κB activation, PGF2α, and expression of contraction-associated genes; impaired luteolysis; and elevated circulating progesterone. These manifestations also occurred in WT females bearing SRC-1/-2 double-deficient embryos, indicating that a fetal-specific defect delayed labor. SP-A, as well as the enzyme lysophosphatidylcholine acyltransferase-1 (LPCAT1), required for synthesis of surfactant dipalmitoylphosphatidylcholine, and the proinflammatory glycerophospholipid platelet-activating factor (PAF) were markedly reduced in SRC-1/-2–deficient fetal lungs near term. Injection of PAF or SP-A into AF at 17.5 days post coitum enhanced uterine NF-κB activation and contractile gene expression, promoted luteolysis, and rescued delayed parturition in SRC-1/-2–deficient embryo-bearing dams. These findings reveal that fetal lungs produce signals to initiate labor when mature and that SRC-1/-2–dependent production of SP-A and PAF is crucial for this process. PMID:26098214

Randomized, placebo-controlled window trial of EGFR, Src, or combined blockade in head and neck cancer.

PubMed

Bauman, Julie E; Duvvuri, Umamaheswar; Gooding, William E; Rath, Tanya J; Gross, Neil D; Song, John; Jimeno, Antonio; Yarbrough, Wendell G; Johnson, Faye M; Wang, Lin; Chiosea, Simion; Sen, Malabika; Kass, Jason; Johnson, Jonas T; Ferris, Robert L; Kim, Seungwon; Hirsch, Fred R; Ellison, Kimberly; Flaherty, John T; Mills, Gordon B; Grandis, Jennifer R

2017-03-23

BACKGROUND. EGFR and Src family kinases are upregulated in head and neck squamous cell carcinoma (HNSCC). EGFR interacts with Src to activate STAT3 signaling, and dual EGFR-Src targeting is synergistic in HNSCC preclinical models. pSrc overexpression predicted resistance to the EGFR inhibitor, erlotinib, in a prior window trial. We conducted a 4-arm window trial to identify biomarkers associated with response to EGFR and/or Src inhibition. METHODS. Patients with operable stage II-IVa HNSCC were randomized to 7-21 days of neoadjuvant erlotinib, the Src inhibitor dasatinib, the combination of both, or placebo. Paired tumor specimens were collected before and after treatment. Pharmacodynamic expression of EGFR and Src pathway components was evaluated by IHC of tissue microarrays and reverse-phase protein array of tissue lysates. Candidate biomarkers were assessed for correlation with change in tumor size. RESULTS. From April 2009 to December 2012, 58 patients were randomized and 55 were treated. There was a significant decrease in tumor size in both erlotinib arms ( P = 0.0014); however, no effect was seen with dasatinib alone ( P = 0.24). High baseline pMAPK expression was associated with response to erlotinib ( P = 0.03). High baseline pSTAT3 was associated with resistance to dasatinib ( P = 0.099). CONCLUSIONS. Brief exposure to erlotinib significantly decreased tumor size in operable HNSCC, with no additive effect from dasatinib. Baseline pMAPK expression warrants further study as a response biomarker for anti-EGFR therapy. Basal expression of pSTAT3 may be independent of Src, explain therapeutic resistance, and preclude development of dasatinib in biomarker-unselected cohorts. TRIAL REGISTRATION. NCT00779389. FUNDING. National Cancer Institute, American Cancer Society, Pennsylvania Department of Health, V Foundation for Cancer Research, Bristol-Myers Squibb, and Astellas Pharma.

Randomized, placebo-controlled window trial of EGFR, Src, or combined blockade in head and neck cancer

PubMed Central

Bauman, Julie E.; Duvvuri, Umamaheswar; Gooding, William E.; Rath, Tanya J.; Gross, Neil D.; Song, John; Jimeno, Antonio; Yarbrough, Wendell G.; Johnson, Faye M.; Wang, Lin; Chiosea, Simion; Sen, Malabika; Kass, Jason; Johnson, Jonas T.; Ferris, Robert L.; Kim, Seungwon; Hirsch, Fred R.; Ellison, Kimberly; Flaherty, John T.; Mills, Gordon B.

2017-01-01

BACKGROUND. EGFR and Src family kinases are upregulated in head and neck squamous cell carcinoma (HNSCC). EGFR interacts with Src to activate STAT3 signaling, and dual EGFR-Src targeting is synergistic in HNSCC preclinical models. pSrc overexpression predicted resistance to the EGFR inhibitor, erlotinib, in a prior window trial. We conducted a 4-arm window trial to identify biomarkers associated with response to EGFR and/or Src inhibition. METHODS. Patients with operable stage II–IVa HNSCC were randomized to 7–21 days of neoadjuvant erlotinib, the Src inhibitor dasatinib, the combination of both, or placebo. Paired tumor specimens were collected before and after treatment. Pharmacodynamic expression of EGFR and Src pathway components was evaluated by IHC of tissue microarrays and reverse-phase protein array of tissue lysates. Candidate biomarkers were assessed for correlation with change in tumor size. RESULTS. From April 2009 to December 2012, 58 patients were randomized and 55 were treated. There was a significant decrease in tumor size in both erlotinib arms (P = 0.0014); however, no effect was seen with dasatinib alone (P = 0.24). High baseline pMAPK expression was associated with response to erlotinib (P = 0.03). High baseline pSTAT3 was associated with resistance to dasatinib (P = 0.099). CONCLUSIONS. Brief exposure to erlotinib significantly decreased tumor size in operable HNSCC, with no additive effect from dasatinib. Baseline pMAPK expression warrants further study as a response biomarker for anti-EGFR therapy. Basal expression of pSTAT3 may be independent of Src, explain therapeutic resistance, and preclude development of dasatinib in biomarker-unselected cohorts. TRIAL REGISTRATION. NCT00779389. FUNDING. National Cancer Institute, American Cancer Society, Pennsylvania Department of Health, V Foundation for Cancer Research, Bristol-Myers Squibb, and Astellas Pharma. PMID:28352657

Roles of steroid receptor coactivator (SRC)-1 and transcriptional intermediary factor (TIF) 2 in androgen receptor activity in mice

PubMed Central

Ye, Xiangcang; Han, Sang Jun; Tsai, Sophia Y.; DeMayo, Francesco J.; Xu, Jianming; Tsai, Ming-Jer; O'Malley, Bert W.

2005-01-01

Genetic disruption of the steroid receptor coactivator (SRC)-1 and transcriptional intermediary factor (TIF)2/SRC-2 in mouse resulted in distinctive mutant phenotypes. To quantify their roles in the function of androgen receptor (AR) transcriptional activity in vivo, we generated a unique transgenic AR-reporter mouse and analyzed the cell-specific contributions of SRC-1 and TIF2 to the activity of AR in mouse testis. Transgenic AR-luciferase and transgenic AR-lacZ mice harbor a recombinant mouse AR gene, ARGAL4DBD, which is functionally coupled with a upstream activation sequence-mediated reporter gene (AR activity indicator). After characterization of these mice in terms of AR function, we further derived bigenic mice by crossing AR activity indicator mice with the SRC-1-/- or TIF2+/- mutant mice. Analyses of the resultant bigenic mice by in vivo imaging and luciferase assays showed that testicular AR activity was decreased significantly in those with the TIF2+/- mutation but not in the SRC-1+/- background, suggesting that TIF2 serves as the preferential coactivator for AR in testis. Immunohistological analysis confirmed that AR and TIF2 coexist in mouse testicular Sertoli cell nuclei under normal conditions. Although SRC-1 concentrates in Sertoli cell nuclei in the absence of TIF2, nuclear SRC-1 is not able to rescue AR activity in the TIF2 mutant background. Interestingly, SRC-1 appears to negatively influence AR activity, thereby counterbalancing the TIF2-stimulated AR activity. Our results provide unique in vivo insights to the multidimensional cell-type-specific interactions between AR and coregulators. PMID:15983373

Roles of steroid receptor coactivator (SRC)-1 and transcriptional intermediary factor (TIF) 2 in androgen receptor activity in mice.

PubMed

Ye, Xiangcang; Han, Sang Jun; Tsai, Sophia Y; DeMayo, Francesco J; Xu, Jianming; Tsai, Ming-Jer; O'Malley, Bert W

2005-07-05

Genetic disruption of the steroid receptor coactivator (SRC)-1 and transcriptional intermediary factor (TIF)2/SRC-2 in mouse resulted in distinctive mutant phenotypes. To quantify their roles in the function of androgen receptor (AR) transcriptional activity in vivo, we generated a unique transgenic AR-reporter mouse and analyzed the cell-specific contributions of SRC-1 and TIF2 to the activity of AR in mouse testis. Transgenic AR-luciferase and transgenic AR-lacZ mice harbor a recombinant mouse AR gene, AR(GAL4DBD), which is functionally coupled with a upstream activation sequence-mediated reporter gene (AR activity indicator). After characterization of these mice in terms of AR function, we further derived bigenic mice by crossing AR activity indicator mice with the SRC-1-/- or TIF2+/- mutant mice. Analyses of the resultant bigenic mice by in vivo imaging and luciferase assays showed that testicular AR activity was decreased significantly in those with the TIF2+/- mutation but not in the SRC-1+/- background, suggesting that TIF2 serves as the preferential coactivator for AR in testis. Immunohistological analysis confirmed that AR and TIF2 coexist in mouse testicular Sertoli cell nuclei under normal conditions. Although SRC-1 concentrates in Sertoli cell nuclei in the absence of TIF2, nuclear SRC-1 is not able to rescue AR activity in the TIF2 mutant background. Interestingly, SRC-1 appears to negatively influence AR activity, thereby counterbalancing the TIF2-stimulated AR activity. Our results provide unique in vivo insights to the multidimensional cell-type-specific interactions between AR and coregulators.

Control of vascular smooth muscle function by Src-family kinases and reactive oxygen species in health and disease

PubMed Central

MacKay, Charles E; Knock, Greg A

2015-01-01

Abstract Reactive oxygen species (ROS) are now recognised as second messenger molecules that regulate cellular function by reversibly oxidising specific amino acid residues of key target proteins. Amongst these are the Src-family kinases (SrcFKs), a multi-functional group of non-receptor tyrosine kinases highly expressed in vascular smooth muscle (VSM). In this review we examine the evidence supporting a role for ROS-induced SrcFK activity in normal VSM contractile function and in vascular remodelling in cardiovascular disease. VSM contractile responses to G-protein-coupled receptor stimulation, as well as hypoxia in pulmonary artery, are shown to be dependent on both ROS and SrcFK activity. Specific phosphorylation targets are identified amongst those that alter intracellular Ca2+ concentration, including transient receptor potential channels, voltage-gated Ca2+ channels and various types of K+ channels, as well as amongst those that regulate actin cytoskeleton dynamics and myosin phosphatase activity, including focal adhesion kinase, protein tyrosine kinase-2, Janus kinase, other focal adhesion-associated proteins, and Rho guanine nucleotide exchange factors. We also examine a growing weight of evidence in favour of a key role for SrcFKs in multiple pro-proliferative and anti-apoptotic signalling pathways relating to oxidative stress and vascular remodelling, with a particular focus on pulmonary hypertension, including growth-factor receptor transactivation and downstream signalling, hypoxia-inducible factors, positive feedback between SrcFK and STAT3 signalling and positive feedback between SrcFK and NADPH oxidase dependent ROS production. We also discuss evidence for and against the potential therapeutic targeting of SrcFKs in the treatment of pulmonary hypertension. PMID:25384773

Steroid receptor coactivators 1 and 2 mediate fetal-to-maternal signaling that initiates parturition.

PubMed

Gao, Lu; Rabbitt, Elizabeth H; Condon, Jennifer C; Renthal, Nora E; Johnston, John M; Mitsche, Matthew A; Chambon, Pierre; Xu, Jianming; O'Malley, Bert W; Mendelson, Carole R

2015-07-01

The precise mechanisms that lead to parturition are incompletely defined. Surfactant protein-A (SP-A), which is secreted by fetal lungs into amniotic fluid (AF) near term, likely provides a signal for parturition; however, SP-A-deficient mice have only a relatively modest delay (~12 hours) in parturition, suggesting additional factors. Here, we evaluated the contribution of steroid receptor coactivators 1 and 2 (SRC-1 and SRC-2), which upregulate SP-A transcription, to the parturition process. As mice lacking both SRC-1 and SRC-2 die at birth due to respiratory distress, we crossed double-heterozygous males and females. Parturition was severely delayed (~38 hours) in heterozygous dams harboring SRC-1/-2-deficient embryos. These mothers exhibited decreased myometrial NF-κB activation, PGF2α, and expression of contraction-associated genes; impaired luteolysis; and elevated circulating progesterone. These manifestations also occurred in WT females bearing SRC-1/-2 double-deficient embryos, indicating that a fetal-specific defect delayed labor. SP-A, as well as the enzyme lysophosphatidylcholine acyltransferase-1 (LPCAT1), required for synthesis of surfactant dipalmitoylphosphatidylcholine, and the proinflammatory glycerophospholipid platelet-activating factor (PAF) were markedly reduced in SRC-1/-2-deficient fetal lungs near term. Injection of PAF or SP-A into AF at 17.5 days post coitum enhanced uterine NF-κB activation and contractile gene expression, promoted luteolysis, and rescued delayed parturition in SRC-1/-2-deficient embryo-bearing dams. These findings reveal that fetal lungs produce signals to initiate labor when mature and that SRC-1/-2-dependent production of SP-A and PAF is crucial for this process.

Reconstructing Solvent Density of Myoglobin Unit Cell from Proximal Radial Distribution Functions of Amino Acids

NASA Astrophysics Data System (ADS)

Galbraith, Madeline; Lynch, Gc; Pettitt, Bm

Understanding the solvent density around a protein crystal structure is an important step for refining accurate crystal structures for use in dynamics simulations or in free energy calculations. The free energy of solvation has typically been approximated using an implicit continuum solvent model or an all atom MD simulation, with a trade-off between accuracy and computation time. For proteins, using precomputed proximal radial distribution functions (pRDFs) of the solvent to reconstruct solvent density on a grid is much faster than all atom MD simulations and more accurate than using implicit solvent models. MD simulations were run for the 20 common amino acids and pRDFs were calculated for several atom type data sets with and without hydrogens, using atom types representative of amino acid side chain atoms. Preliminary results from reconstructions suggest using a data set with 15 heavy atoms and 3 hydrogen yields results with the lowest error without a tradeoff on time. The results of using precomputed pRDFs to reconstruct the solvent density of water for the myoglobin (pdb ID 2mgk) unit cell quantifies the accuracy of the method in comparison with the crystallographic data. Funding Acknowledgement: This research was funded by the CPRIT Summer Undergraduate Program in Computational Cancer Biology, training Grant award RP 140113 from the Cancer Prevention & Research Institute of Texas (CPRIT).

77 FR 58868 - Teleconference for the National Park Service Alaska Region's Subsistence Resource Commission Program

Federal Register 2010, 2011, 2012, 2013, 2014

2012-09-24

... Park Subsistence Resource Commission (SRC) and the Wrangell-St. Elias National Park SRC will meet to... and Location for Next Meeting 12. Adjourn Meeting Wrangell-St. Elias National Park SRC Meeting Date and Location: The [[Page 58869

Phosphatidylinositol Phosphate 5-Kinase Iγi2 in Association with Src Controls Anchorage-independent Growth of Tumor Cells*

PubMed Central

Thapa, Narendra; Choi, Suyong; Hedman, Andrew; Tan, Xiaojun; Anderson, Richard A.

2013-01-01

A fundamental property of tumor cells is to defy anoikis, cell death caused by a lack of cell-matrix interaction, and grow in an anchorage-independent manner. How tumor cells organize signaling molecules at the plasma membrane to sustain oncogenic signals in the absence of cell-matrix interactions remains poorly understood. Here, we describe a role for phosphatidylinositol 4-phosphate 5-kinase (PIPK) Iγi2 in controlling anchorage-independent growth of tumor cells in coordination with the proto-oncogene Src. PIPKIγi2 regulated Src activation downstream of growth factor receptors and integrins. PIPKIγi2 directly interacted with the C-terminal tail of Src and regulated its subcellular localization in concert with talin, a cytoskeletal protein targeted to focal adhesions. Co-expression of PIPKIγi2 and Src synergistically induced the anchorage-independent growth of nonmalignant cells. This study uncovers a novel mechanism where a phosphoinositide-synthesizing enzyme, PIPKIγi2, functions with the proto-oncogene Src, to regulate oncogenic signaling. PMID:24151076

Hsp90 dependence of a kinase is determined by its conformational landscape

PubMed Central

Luo, Qi; Boczek, Edgar E.; Wang, Qi; Buchner, Johannes; Kaila, Ville R. I.

2017-01-01

Heat shock protein 90 (Hsp90) is an abundant molecular chaperone, involved in the folding and activation of 60% of the human kinome. The oncogenic tyrosine kinase v-Src is one of the most stringent client proteins of Hsp90, whereas its almost identical homolog c-Src is only weakly affected by the chaperone. Here, we perform atomistic molecular simulations and in vitro kinase assays to explore the mechanistic differences in the activation of v-Src and c-Src. While activation in c-Src is strictly controlled by ATP-binding and phosphorylation, we find that activating conformational transitions are spontaneously sampled in Hsp90-dependent Src mutants. Phosphorylation results in an enrichment of the active conformation and in an increased affinity for Hsp90. Thus, the conformational landscape of the mutated kinase is reshaped by a broken “control switch”, resulting in perturbations of long-range electrostatics, higher activity and increased Hsp90-dependence. PMID:28290541

Differential Requirements for Src-Family Kinases in SYK or ZAP70-Mediated SLP-76 Phosphorylation in Lymphocytes

PubMed Central

Fasbender, Frank; Claus, Maren; Wingert, Sabine; Sandusky, Mina; Watzl, Carsten

2017-01-01

In a synthetic biology approach using Schneider (S2) cells, we show that SLP-76 is directly phosphorylated at tyrosines Y113 and Y128 by SYK in the presence of ITAM-containing adapters such as CD3ζ, DAP12, or FcεRγ. This phosphorylation was dependent on at least one functional ITAM and a functional SH2 domain within SYK. Inhibition of Src-kinases by inhibitors PP1 and PP2 did not reduce SLP-76 phosphorylation in S2 cells, suggesting an ITAM and SYK dependent, but Src-kinase independent signaling pathway. This direct ITAM/SYK/SLP-76 signaling pathway therefore differs from previously described ITAM signaling. However, the SYK-family kinase ZAP70 required the additional co-expression of the Src-family kinases Fyn or Lck to efficiently phosphorylate SLP-76 in S2 cells. This difference in Src-family kinase dependency of SYK versus ZAP70-mediated ITAM-based signaling was further demonstrated in human lymphocytes. ITAM signaling in ZAP70-expressing T cells was dependent on the activity of Src-family kinases. In contrast, Src-family kinases were partially dispensable for ITAM signaling in SYK-expressing B cells or in natural killer cells, which express SYK and ZAP70. This demonstrates that SYK can signal using a Src-kinase independent ITAM-based signaling pathway, which may be involved in calibrating the threshold for lymphocyte activation. PMID:28736554

Differential Requirements for Src-Family Kinases in SYK or ZAP70-Mediated SLP-76 Phosphorylation in Lymphocytes.

PubMed

Fasbender, Frank; Claus, Maren; Wingert, Sabine; Sandusky, Mina; Watzl, Carsten

2017-01-01

In a synthetic biology approach using Schneider (S2) cells, we show that SLP-76 is directly phosphorylated at tyrosines Y113 and Y128 by SYK in the presence of ITAM-containing adapters such as CD3ζ, DAP12, or FcεRγ. This phosphorylation was dependent on at least one functional ITAM and a functional SH2 domain within SYK. Inhibition of Src-kinases by inhibitors PP1 and PP2 did not reduce SLP-76 phosphorylation in S2 cells, suggesting an ITAM and SYK dependent, but Src-kinase independent signaling pathway. This direct ITAM/SYK/SLP-76 signaling pathway therefore differs from previously described ITAM signaling. However, the SYK-family kinase ZAP70 required the additional co-expression of the Src-family kinases Fyn or Lck to efficiently phosphorylate SLP-76 in S2 cells. This difference in Src-family kinase dependency of SYK versus ZAP70-mediated ITAM-based signaling was further demonstrated in human lymphocytes. ITAM signaling in ZAP70-expressing T cells was dependent on the activity of Src-family kinases. In contrast, Src-family kinases were partially dispensable for ITAM signaling in SYK-expressing B cells or in natural killer cells, which express SYK and ZAP70. This demonstrates that SYK can signal using a Src-kinase independent ITAM-based signaling pathway, which may be involved in calibrating the threshold for lymphocyte activation.

Src-dependent EGFR transactivation regulates lung inflammation via downstream signaling involving ERK1/2, PI3Kδ/Akt and NFκB induction in a murine asthma model.

PubMed

El-Hashim, Ahmed Z; Khajah, Maitham A; Renno, Waleed M; Babyson, Rhema S; Uddin, Mohib; Benter, Ibrahim F; Ezeamuzie, Charles; Akhtar, Saghir

2017-08-30

The molecular mechanisms underlying asthma pathogenesis are poorly characterized. In this study, we investigated (1) whether Src mediates epidermal growth factor receptor (EGFR) transactivation; (2) if ERK1/2, PI3Kδ/Akt and NF-κB are signaling effectors downstream of Src/EGFR activation; and (3) if upstream inhibition of Src/EGFR is more effective in downregulating the allergic inflammation than selective inhibition of downstream signaling pathways. Allergic inflammation resulted in increased phosphorylation of EGFR, Akt, ERK1/2 and IκB in the lung tissues from ovalbumin (OVA)-challenged BALB/c mice. Treatment with inhibitors of Src (SU6656) or EGFR (AG1478) reduced EGFR phosphorylation and downstream signaling which resulted in the inhibition of the OVA-induced inflammatory cell influx in bronchoalveolar lavage fluid (BALF), perivascular and peribronchial inflammation, fibrosis, goblet cell hyper/metaplasia and airway hyper-responsiveness. Treatment with pathway-selective inhibitors for ERK1/2 (PD89059) and PI3Kδ/Akt (IC-87114) respectively, or an inhibitor of NF-κB (BAY11-7085) also reduced the OVA-induced asthmatic phenotype but to a lesser extent compared to Src/EGFR inhibition. Thus, Src via EGFR transactivation and subsequent downstream activation of multiple pathways regulates the allergic airway inflammatory response. Furthermore, a broader upstream inhibition of Src/EGFR offers an attractive therapeutic alternative in the treatment of asthma relative to selectively targeting the individual downstream signaling effectors.

Examining Recovery Trajectories Following Sport-related Concussion Using a Multi-Modal Clinical Assessment Approach

PubMed Central

Henry, Luke C.; Elbin, RJ; Collins, Michael W.; Marchetti, Gregory; Kontos, Anthony P.

2016-01-01

Background Previous research estimates that the majority of athletes with sport-related concussion (SRC) will recover between 7–10 days following injury. This short, temporal window of recovery is predominately based on symptom resolution and cognitive improvement, and does not accurately reflect recent advances to the clinical assessment model. Objective To characterize SRC recovery at 1-week post-injury time intervals on symptom, neurocognitive, and vestibular-oculomotor outcomes, and examine gender differences on SRC recovery time. Methods A prospective, repeated measures design was used to examine the temporal resolution of neurocognitive, symptom, and vestibular-oculomotor impairment in 66 subjects (16.5 ± 1.9 years, range 14–23, 64% male) with SRC. Results Recovery time across all outcomes was between 21–28 days post SRC for most athletes. Symptoms demonstrated the greatest improvement in the first 2 weeks, while neurocognitive impairment lingered across various domains up to 28 days post SRC. Vestibular-oculomotor decrements also resolved between one to three weeks post injury. There were no gender differences in neurocognitive recovery. Males were more likely to be asymptomatic by the fourth week and reported less vestibular-oculomotor impairment than females at weeks 1 and 2. Conclusion When utilizing the recommended “comprehensive” approach for concussion assessment, recovery time for SRC is approximately three to four weeks, which is longer than the commonly reported 7–14 days. Sports medicine clinicians should use a variety of complementing assessment tools to capture the heterogeneity of SRC. PMID:26445375

The transcriptional coactivators p/CIP and SRC-1 control insulin resistance through IRS1 in obesity models.

PubMed

Wang, Zhiyong; Shah, O Jameel; Hunter, Tony

2012-01-01

Three p160 family members, p/CIP, SRC1, and TIF2, have been identified as transcriptional coactivators for nuclear hormone receptors and other transcription factors in vitro. In a previous study, we reported initial characterization of the obesity-resistant phenotypes of p/CIP and SRC-1 double knockout (DKO) mice, which exhibit increased energy expenditure, and suggested that nuclear hormone receptor target genes were involved in these phenotypes. In this study, we demonstrate that p/CIP and SRC1 control insulin signaling in a cell-autonomous manner both in vitro and in vivo. Genetic deletion of p/CIP and SRC-1 increases glucose uptake and enhances insulin sensitivity in both regular chow- and high fat diet-fed DKO mice despite increased food intake. Interestingly, we discover that loss of p/CIP and SRC-1 results in resistance to age-related obesity and glucose intolerance. We show that expression levels of a key insulin signaling component, insulin receptor substrate 1 (IRS1), are significantly increased in two cell lines representing fat and muscle lineages with p/CIP and SRC-1 deletions and in white adipose tissue and skeletal muscle of DKO mice; this may account for increased glucose metabolism and insulin sensitivity. This is the first evidence that the p160 coactivators control insulin signaling and glucose metabolism through IRS1. Therefore, our studies indicate that p/CIP and SRC-1 are potential therapeutic targets not only for obesity but also for diabetes.

The Transcriptional Coactivators p/CIP and SRC-1 Control Insulin Resistance through IRS1 in Obesity Models

PubMed Central

Wang, Zhiyong; Shah, O. Jameel; Hunter, Tony

2012-01-01

Three p160 family members, p/CIP, SRC1, and TIF2, have been identified as transcriptional coactivators for nuclear hormone receptors and other transcription factors in vitro. In a previous study, we reported initial characterization of the obesity-resistant phenotypes of p/CIP and SRC-1 double knockout (DKO) mice, which exhibit increased energy expenditure, and suggested that nuclear hormone receptor target genes were involved in these phenotypes. In this study, we demonstrate that p/CIP and SRC1 control insulin signaling in a cell-autonomous manner both in vitro and in vivo. Genetic deletion of p/CIP and SRC-1 increases glucose uptake and enhances insulin sensitivity in both regular chow- and high fat diet-fed DKO mice despite increased food intake. Interestingly, we discover that loss of p/CIP and SRC-1 results in resistance to age-related obesity and glucose intolerance. We show that expression levels of a key insulin signaling component, insulin receptor substrate 1 (IRS1), are significantly increased in two cell lines representing fat and muscle lineages with p/CIP and SRC-1 deletions and in white adipose tissue and skeletal muscle of DKO mice; this may account for increased glucose metabolism and insulin sensitivity. This is the first evidence that the p160 coactivators control insulin signaling and glucose metabolism through IRS1. Therefore, our studies indicate that p/CIP and SRC-1 are potential therapeutic targets not only for obesity but also for diabetes. PMID:22859932

Examining Recovery Trajectories After Sport-Related Concussion With a Multimodal Clinical Assessment Approach.

PubMed

Henry, Luke C; Elbin, R J; Collins, Michael W; Marchetti, Gregory; Kontos, Anthony P

2016-02-01

Previous research estimates that the majority of athletes with sport-related concussion (SRC) will recover between 7 and 10 days after injury. This short temporal window of recovery is based predominately on symptom resolution and cognitive improvement and does not accurately reflect recent advances in the clinical assessment model. To characterize SRC recovery at 1-week postinjury time intervals on symptom, neurocognitive, and vestibular-oculomotor outcomes and to examine sex differences in SRC recovery time. A prospective, repeated-measures design was used to examine the temporal resolution of neurocognitive, symptom, and vestibular-oculomotor impairment in 66 subjects (age, 16.5 ± 1.9 years; range, 14-23 years; 64% male) with SRC. Recovery time across all outcomes was between 21 and 28 days after SRC for most athletes. Symptoms demonstrated the greatest improvement in the first 2 weeks, although neurocognitive impairment lingered across various domains up to 28 days after SRC. Vestibular-oculomotor decrements also resolved between 1 and 3 weeks after injury. There were no sex differences in neurocognitive recovery. Male subjects were more likely to be asymptomatic by the fourth week and reported less vestibular-oculomotor impairment than female subjects at weeks 1 and 2. When the recommended "comprehensive" approach is used for concussion assessment, recovery time for SRC is approximately 3 to 4 weeks, which is longer than the commonly reported 7 to 14 days. Sports medicine clinicians should use a variety of complementing assessment tools to capture the heterogeneity of SRC.

Soil CO2 flux from three ecosystems in tropical peatland of Sarawak, Malaysia

NASA Astrophysics Data System (ADS)

Melling, Lulie; Hatano, Ryusuke; Goh, Kah Joo

2005-02-01

Soil CO2 flux was measured monthly over a year from tropical peatland of Sarawak, Malaysia using a closed-chamber technique. The soil CO2 flux ranged from 100 to 533 mg C m2 h1 for the forest ecosystem, 63 to 245 mg C m2 h1 for the sago and 46 to 335 mg C m2 h1 for the oil palm. Based on principal component analysis (PCA), the environmental variables over all sites could be classified into three components, namely, climate, soil moisture and soil bulk density, which accounted for 86% of the seasonal variability. A regression tree approach showed that CO2 flux in each ecosystem was related to different underlying environmental factors. They were relative humidity for forest, soil temperature at 5 cm for sago and water-filled pore space for oil palm. On an annual basis, the soil CO2 flux was highest in the forest ecosystem with an estimated production of 2.1 kg C m2 yr1 followed by oil palm at 1.5 kg C m2 yr1 and sago at 1.1 kg C m2 yr1. The different dominant controlling factors in CO2 flux among the studied ecosystems suggested that land use affected the exchange of CO2 between tropical peatland and the atmosphere.

Improving Protocols for Protein Mapping through Proper Comparison to Crystallography Data

PubMed Central

Lexa, Katrina W.; Carlson, Heather A.

2013-01-01

Computational approaches to fragment-based drug design (FBDD) can complement experiments and facilitate the identification of potential hot spots along the protein surface. However, the evaluation of computational methods for mapping binding sites frequently focuses upon the ability to reproduce crystallographic coordinates to within a low RMSD threshold. This dependency on the deposited coordinate data overlooks the original electron density from the experiment, thus techniques may be developed based upon subjective - or even erroneous - atomic coordinates. This can become a significant drawback in applications to systems where the location of hot spots is unknown. Based on comparison to crystallographic density, we previously showed that mixed-solvent molecular dynamics (MixMD) accurately identifies the active site for HEWL, with acetonitrile as an organic solvent. Here, we concentrated on the influence of protic solvent on simulation and refined the optimal MixMD approach for extrapolation of the method to systems without established sites. Our results establish an accurate approach for comparing simulations to experiment. We have outlined the most efficient strategy for MixMD, based on simulation length and number of runs. The development outlined here makes MixMD a robust method which should prove useful across a broad range of target structures. Lastly, our results with MixMD match experimental data so well that consistency between simulations and density may be a useful way to aid the identification of probes vs waters during the refinement of future MSCS crystallographic structures. PMID:23327200

KoBaMIN: a knowledge-based minimization web server for protein structure refinement.

PubMed

Rodrigues, João P G L M; Levitt, Michael; Chopra, Gaurav

2012-07-01

The KoBaMIN web server provides an online interface to a simple, consistent and computationally efficient protein structure refinement protocol based on minimization of a knowledge-based potential of mean force. The server can be used to refine either a single protein structure or an ensemble of proteins starting from their unrefined coordinates in PDB format. The refinement method is particularly fast and accurate due to the underlying knowledge-based potential derived from structures deposited in the PDB; as such, the energy function implicitly includes the effects of solvent and the crystal environment. Our server allows for an optional but recommended step that optimizes stereochemistry using the MESHI software. The KoBaMIN server also allows comparison of the refined structures with a provided reference structure to assess the changes brought about by the refinement protocol. The performance of KoBaMIN has been benchmarked widely on a large set of decoys, all models generated at the seventh worldwide experiments on critical assessment of techniques for protein structure prediction (CASP7) and it was also shown to produce top-ranking predictions in the refinement category at both CASP8 and CASP9, yielding consistently good results across a broad range of model quality values. The web server is fully functional and freely available at http://csb.stanford.edu/kobamin.

Protein structure modeling and refinement by global optimization in CASP12.

PubMed

Hong, Seung Hwan; Joung, InSuk; Flores-Canales, Jose C; Manavalan, Balachandran; Cheng, Qianyi; Heo, Seungryong; Kim, Jong Yun; Lee, Sun Young; Nam, Mikyung; Joo, Keehyoung; Lee, In-Ho; Lee, Sung Jong; Lee, Jooyoung

2018-03-01

For protein structure modeling in the CASP12 experiment, we have developed a new protocol based on our previous CASP11 approach. The global optimization method of conformational space annealing (CSA) was applied to 3 stages of modeling: multiple sequence-structure alignment, three-dimensional (3D) chain building, and side-chain re-modeling. For better template selection and model selection, we updated our model quality assessment (QA) method with the newly developed SVMQA (support vector machine for quality assessment). For 3D chain building, we updated our energy function by including restraints generated from predicted residue-residue contacts. New energy terms for the predicted secondary structure and predicted solvent accessible surface area were also introduced. For difficult targets, we proposed a new method, LEEab, where the template term played a less significant role than it did in LEE, complemented by increased contributions from other terms such as the predicted contact term. For TBM (template-based modeling) targets, LEE performed better than LEEab, but for FM targets, LEEab was better. For model refinement, we modified our CASP11 molecular dynamics (MD) based protocol by using explicit solvents and tuning down restraint weights. Refinement results from MD simulations that used a new augmented statistical energy term in the force field were quite promising. Finally, when using inaccurate information (such as the predicted contacts), it was important to use the Lorentzian function for which the maximal penalty arising from wrong information is always bounded. © 2017 Wiley Periodicals, Inc.

Sport-Related Concussions

ERIC Educational Resources Information Center

Brady, Don; Brady, Flo

2011-01-01

Sport-related concussions (SRC) are not limited to specific age ranges, professional athletes, or gender. The primary focus of much of SRC research pertains to the assessment, management, and return to play (RTP) of the concussed athlete. This article highlights some major issues of SRC along with some controversies that presently exist within the…

Sex differences in sport-related concussion long-term outcomes.

PubMed

Covassin, Tracey; Savage, Jennifer L; Bretzin, Abigail C; Fox, Meghan E

2017-09-18

Approximately 1.6 to 3.8 million recreational and sports-related concussions (SRC) occur each year in the Unites States. Research suggest that female athletes are at a greater risk for a SRC compared to male athletes competing in comparable sports (i.e., soccer, basketball). Moreover, female athletes have reported more total symptoms and greater neurocognitive impairments following a SRC. Female athletes have been found to report greater symptom provocation as measured by the Vestibular/Ocular Motor Screening (VOMS), and increased brain activation compared to males. There is a scarcity of research on long-term effects of SRC in male and female athletes. Therefore, the aim of this review article is to summarize the existing literature on sex differences in acute and sub-acute SRC outcomes. Copyright © 2017. Published by Elsevier B.V.

Src is required for migration, phagocytosis, and interferon beta production in Toll-like receptor-engaged macrophages.

PubMed

Maa, Ming-Chei; Leu, Tzeng-Horng

2016-06-01

As an evolutionarily conserved mechanism, innate immunity controls self-nonself discrimination to protect a host from invasive pathogens. Macrophages are major participants of the innate immune system. Through the activation of diverse Toll-like receptors (TLRs), macrophages are triggered to initiate a variety of functions including locomotion, phagocytosis, and secretion of cytokines that requires the participation of tyrosine kinases. Fgr, Hck, and Lyn are myeloid-specific Src family kinases. Despite their constitutively high expression in macrophages, their absence does not impair LPS responsiveness. In contrast, Src, a barely detectable tyrosine kinase in resting macrophages, becomes greatly inducible in response to TLR engagement, implicating its role in macrophage activation. Indeed, silencing Src suppresses the activated TLR-mediated migration, phagocytosis, and interferon-beta (IFN-β) secretion in macrophages. And these physiological defects can be restored by the introduction of siRNA-resistant Src. Notably, the elevated expression and activity of Src is inducible nitric oxide synthase (iNOS)-dependent. Due to (1) iNOS being a NF-κB target, which can be induced by various TLR ligands, (2) Src can mediate NF-κB activation, therefore, there ought to exist a loop of signal amplification that regulates macrophage physiology in response to the engagement of TLRs.

Interaction between Sam68 and Src family tyrosine kinases, Fyn and Lck, in T cell receptor signaling.

PubMed

Fusaki, N; Iwamatsu, A; Iwashima, M; Fujisawa, J i

1997-03-07

The Src family protein-tyrosine kinase, Fyn, is associated with the T cell receptor (TCR) and plays an important role in TCR-mediated signaling. We found that a human T cell leukemia virus type 1-infected T cell line, Hayai, overexpressed Fyn. To identify the molecules downstream of Fyn, we analyzed the tyrosine phosphorylation of cellular proteins in the cells. In Hayai, a 68-kDa protein was constitutively tyrosine-phosphorylated. The 68-kDa protein was coimmunoprecipitated with various signaling proteins such as phospholipase C gamma1, the phosphatidylinositol 3-kinase p85 subunit, Grb2, SHP-1, Cbl, and Jak3, implying that the protein might function as an adapter. Purification and microsequencing of this protein revealed that it was the RNA-binding protein, Sam68 (Src associated in mitosis, 68 kDa). Sam68 was associated with the Src homology 2 and 3 domains of Fyn and also those of another Src family kinase, Lck. CD3 cross-linking induced tyrosine phosphorylation of Sam68 in uninfected T cells. These data suggest that Sam68 participates in the signal transduction pathway downstream of TCR-coupled Src family kinases Fyn and Lck in lymphocytes, that is not only in the mitotic pathway downstream of c-Src in fibroblasts.

Nanometer Scale Titanium Surface Texturing Are Detected by Signaling Pathways Involving Transient FAK and Src Activations

PubMed Central

Zambuzzi, Willian F.; Bonfante, Estevam A.; Jimbo, Ryo; Hayashi, Mariko; Andersson, Martin; Alves, Gutemberg; Takamori, Esther R.; Beltrão, Paulo J.; Coelho, Paulo G.; Granjeiro, José M.

2014-01-01

Background It is known that physico/chemical alterations on biomaterial surfaces have the capability to modulate cellular behavior, affecting early tissue repair. Such surface modifications are aimed to improve early healing response and, clinically, offer the possibility to shorten the time from implant placement to functional loading. Since FAK and Src are intracellular proteins able to predict the quality of osteoblast adhesion, this study evaluated the osteoblast behavior in response to nanometer scale titanium surface texturing by monitoring FAK and Src phosphorylations. Methodology Four engineered titanium surfaces were used for the study: machined (M), dual acid-etched (DAA), resorbable media microblasted and acid-etched (MBAA), and acid-etch microblasted (AAMB). Surfaces were characterized by scanning electron microscopy, interferometry, atomic force microscopy, x-ray photoelectron spectroscopy and energy dispersive X-ray spectroscopy. Thereafter, those 4 samples were used to evaluate their cytotoxicity and interference on FAK and Src phosphorylations. Both Src and FAK were investigated by using specific antibody against specific phosphorylation sites. Principal Findings The results showed that both FAK and Src activations were differently modulated as a function of titanium surfaces physico/chemical configuration and protein adsorption. Conclusions It can be suggested that signaling pathways involving both FAK and Src could provide biomarkers to predict osteoblast adhesion onto different surfaces. PMID:24999733

Dynamic organization of myristoylated Src in the live cell plasma membrane

DOE Office of Scientific and Technical Information (OSTI.GOV)

Smith, Adam W.; Huang, Hector H.; Endres, Nicholas F.

The spatial organization of lipid-anchored proteins in the plasma membrane directly influences cell signaling, but measuring such organization in situ is experimentally challenging. The canonical oncogene, c-Src, is a lipid anchored protein that plays a key role in integrin-mediated signal transduction within focal adhesions and cell–cell junctions. Because of its activity in specific plasma membrane regions, structural motifs within the protein have been hypothesized to play an important role in its subcellular localization. This study used a combination of time-resolved fluorescence fluctuation spectroscopy and super-resolution microscopy to quantify the dynamic organization of c-Src in live cell membranes. Pulsed-interleaved excitation fluorescencemore » cross-correlation spectroscopy (PIE–FCCS) showed that a small fraction of c-Src transiently sorts into membrane clusters that are several times larger than the monomers. Photoactivated localization microscopy (PALM) confirmed that c-Src partitions into clusters with low probability and showed that the characteristic size of the clusters is 10–80 nm. Finally, time-resolved fluorescence anisotropy measurements were used to quantify the rotational mobility of c-Src to determine how it interacts with its local environment. Altogether, these results build a quantitative description of the mobility and clustering behavior of the c-Src nonreceptor tyrosine kinase in the live cell plasma membrane.« less

Dynamic organization of myristoylated Src in the live cell plasma membrane

DOE PAGES

Smith, Adam W.; Huang, Hector H.; Endres, Nicholas F.; ...

2016-01-15

The spatial organization of lipid-anchored proteins in the plasma membrane directly influences cell signaling, but measuring such organization in situ is experimentally challenging. The canonical oncogene, c-Src, is a lipid anchored protein that plays a key role in integrin-mediated signal transduction within focal adhesions and cell–cell junctions. Because of its activity in specific plasma membrane regions, structural motifs within the protein have been hypothesized to play an important role in its subcellular localization. This study used a combination of time-resolved fluorescence fluctuation spectroscopy and super-resolution microscopy to quantify the dynamic organization of c-Src in live cell membranes. Pulsed-interleaved excitation fluorescencemore » cross-correlation spectroscopy (PIE–FCCS) showed that a small fraction of c-Src transiently sorts into membrane clusters that are several times larger than the monomers. Photoactivated localization microscopy (PALM) confirmed that c-Src partitions into clusters with low probability and showed that the characteristic size of the clusters is 10–80 nm. Finally, time-resolved fluorescence anisotropy measurements were used to quantify the rotational mobility of c-Src to determine how it interacts with its local environment. Altogether, these results build a quantitative description of the mobility and clustering behavior of the c-Src nonreceptor tyrosine kinase in the live cell plasma membrane.« less

SRC-induced disintegration of adherens junctions of madin-darby canine kidney cells is dependent on endocytosis of cadherin and antagonized by Tiam-1.

PubMed

Palovuori, Riitta; Sormunen, Raija; Eskelinen, Sinikka

2003-12-01

The effects of Src tyrosine kinase activation in subconfluent temperature sensitive (ts)-Src-transformed Madin-Darby canine kidney (MDCK) cells were analyzed by shifting them from nonpermissive (40.5 degrees C) to permissive (35 degrees C) temperature. Already, in 15 minutes, adherens junction components were released from the lateral walls and accumulated to basal surfaces. Simultaneously, membranous actin staining vanished, actin bundles appeared at the basal surface, and the cells flattened. The only component phosphorylated and translocated after the shift to 35 degrees C was p120ctn. The epithelial-mesenchymal transition could be inhibited by a specific inhibitor of Src kinase, PP2, or by inhibiting endocytosis. Therefore, Src activation was responsible for the transition, but not because of phosphorylation of adherens junction components but by way of activation of endocytic machinery and RhoGTPase. Expression of an RacGEF, Tiam-1 (T-lymphoma invasion and metastasis gene 1), prevented flattening of Src-transformed MDCK cells at 35 degrees C and resulted in accumulation of cadherin to lateral membranes. In the case where the Src-MDCK cells were cultivated at 35 degrees C and shifted for short time periods to 40.5 degrees C, cadherin rapidly returned to lateral membranes, whereas actin and p120ctn followed hours afterward. This further supports the view that cadherin internalization is the primary target of Src kinase. We also looked at the cell morphology and distribution of cadherin and Tiam-1 in cells grown in three-dimensional gels composed of collagen and laminin or in Matrigel. At nonpermissive temperature, both Src-MDCK and Tiam-1-transfected Src-MDCK cells exhibited nonpolarized morphology in collagen I, a loose cluster in the mixture of collagen I and laminin, and a differentiated cyst in Matrigel. In growth factor-depleted Matrigel, the Src-MDCK cells grew in nondifferentiated clusters, whereas Tiam-1-transfected cells went to apoptosis. The differentiated phenotype of both cell lines could be rescued by Matrigel-conditioned medium, platelet-derived growth factor, or cholera toxin. Concomitantly, both cadherin and Tiam-1 were recruited to lateral membranes. Therefore, cadherin and Tiam-1 seem to be the key players in the differentiation process of MDCK cells.

Contribution of Hydrogen Bonds to Paper Strength Properties.

PubMed

Przybysz, Piotr; Dubowik, Marcin; Kucner, Marta Anna; Przybysz, Kazimierz; Przybysz Buzała, Kamila

2016-01-01

The objective of this work was to investigate the influence of hydrogen bonds between fibres on static and dynamic strength properties of paper. A commercial bleached pinewood kraft pulp was soaked in water, refined in a PFI, and used to form paper webs in different solvents, such as water, methanol, ethanol, n-propanol and n-butanol, to determine the effect of their dipole moment on static and dynamic strength properties of resulting paper sheets. Paper which was formed in water, being the solvent of the highest dipole moment among the tested ones, showed the highest breaking length and tear resistance. When paper webs were formed in n-butanol, which was the least polar among the solvents, these parameters were reduced by around 75%. These results provide evidence of the importance of water in paper web formation and strong impact of hydrogen bonds between fibres on strength properties of paper.

Contribution of Hydrogen Bonds to Paper Strength Properties

PubMed Central

Przybysz, Piotr; Dubowik, Marcin; Kucner, Marta Anna; Przybysz, Kazimierz; Przybysz Buzała, Kamila

2016-01-01

The objective of this work was to investigate the influence of hydrogen bonds between fibres on static and dynamic strength properties of paper. A commercial bleached pinewood kraft pulp was soaked in water, refined in a PFI, and used to form paper webs in different solvents, such as water, methanol, ethanol, n-propanol and n-butanol, to determine the effect of their dipole moment on static and dynamic strength properties of resulting paper sheets. Paper which was formed in water, being the solvent of the highest dipole moment among the tested ones, showed the highest breaking length and tear resistance. When paper webs were formed in n-butanol, which was the least polar among the solvents, these parameters were reduced by around 75%. These results provide evidence of the importance of water in paper web formation and strong impact of hydrogen bonds between fibres on strength properties of paper. PMID:27228172

Acute, subchronic, and developmental toxicological properties of lubricating oil base stocks.

PubMed

Dalbey, Walden E; McKee, Richard H; Goyak, Katy Olsavsky; Biles, Robert W; Murray, Jay; White, Russell

2014-01-01

Lubricating oil base stocks (LOBs) are substances used in the manufacture of finished lubricants and greases. They are produced from residue remaining after atmospheric distillation of crude oil that is subsequently fractionated by vacuum distillation and additional refining steps. Initial LOB streams that have been produced by vacuum distillation but not further refined may contain polycyclic aromatic compounds (PACs) and may present carcinogenic hazards. In modern refineries, LOBs are further refined by multistep processes including solvent extraction and/or hydrogen treatment to reduce the levels of PACs and other undesirable constituents. Thus, mildly (insufficiently) refined LOBs are potentially more hazardous than more severely (sufficiently) refined LOBs. This article discusses the evaluation of LOBs using statistical models based on content of PACs; these models indicate that insufficiently refined LOBs (potentially carcinogenic LOBs) can also produce systemic and developmental effects with repeated dermal exposure. Experimental data were also obtained in ten 13-week dermal studies in rats, eight 4-week dermal studies in rabbits, and seven dermal developmental toxicity studies with sufficiently refined LOBs (noncarcinogenic and commonly marketed) in which no observed adverse effect levels for systemic toxicity and developmental toxicity were 1000 to 2000 mg/kg/d with dermal exposures, typically the highest dose tested. Results in both oral and inhalation developmental toxicity studies were similar. This absence of toxicologically relevant findings was consistent with lower PAC content of sufficiently refined LOBs. Based on data on reproductive organs with repeated dosing and parameters in developmental toxicity studies, sufficiently refined LOBs are likely to have little, if any, effect on reproductive parameters.

NEPRILYSIN REGULATES PULMONARY ARTERY SMOOTH MUSCLE CELL PHENOTYPE THROUGH A PDGF RECEPTOR DEPENDENT MECHANISM

PubMed Central

Karoor, Vijaya; Oka, Masahiko; Walchak, Sandra J.; Hersh, Louis B.; Miller, York E.; Dempsey, Edward C.

2013-01-01

Reduced neprilysin (NEP), a cell surface metallopeptidase, which cleaves and inactivates pro-inflammatory and vasoactive peptides, predisposes the lung vasculature to exaggerated remodeling in response to hypoxia. We hypothesize that loss of NEP in pulmonary artery smooth muscle cells (PASMCs) results in increased migration and proliferation. PASMCs isolated from NEP−/− mice exhibited enhanced migration and proliferation in response to serum and PDGF, which was attenuated by NEP replacement. Inhibition of NEP by overexpression of a peptidase dead mutant or knockdown by siRNA in NEP+/+ cells increased migration and proliferation. Loss of NEP led to an increase in Src kinase activity and phosphorylation of PTEN resulting in activation of the PDGF receptor (PDGFR). Knockdown of Src kinase with siRNA or inhibition with PP2 a src kinase inhibitor decreased PDGFRY751 phosphorylation and attenuated migration and proliferation in NEP−/− SMCs. NEP substrates, endothelin-1(ET-1) or fibroblast growth factor-2 (FGF2), increased activation of Src and PDGFR in NEP+/+ cells, which was decreased by an ETAR antagonist, neutralizing antibody to FGF2 and Src inhibitor. Similar to the observations in PASMCs levels of p-PDGFR, p-Src and p-PTEN were elevated in NEP−/− lungs. ETAR antagonist also attenuated the enhanced responses in NEP−/−PASMCs and lungs. Taken together our results suggest a novel mechanism for regulation of PDGFR signaling by NEP substrates involving Src and PTEN. Strategies that increase lung NEP activity/expression or target key downstream effectors, like Src, PTEN or PDGFR, may be of therapeutic benefit in pulmonary vascular disease. PMID:23381789

Roles of the SH2 and SH3 domains in the regulation of neuronal Src kinase functions.

PubMed

Groveman, Bradley R; Xue, Sheng; Marin, Vedrana; Xu, Jindong; Ali, Mohammad K; Bienkiewicz, Ewa A; Yu, Xian-Min

2011-02-01

Previous studies demonstrated that intra-domain interactions between Src family kinases (SFKs), stabilized by binding of the phosphorylated C-terminus to the SH2 domain and/or binding of the SH2 kinase linker to the SH3 domain, lock the molecules in a closed conformation, disrupt the kinase active site, and inactivate SFKs. Here we report that the up-regulation of N-methyl-D-aspartate receptors (NMDARs) induced by expression of constitutively active neuronal Src (n-Src), in which the C-terminus tyrosine is mutated to phenylalanine (n-Src/Y535F), is significantly reduced by dysfunctions of the SH2 and/or SH3 domains of the protein. Furthermore, we found that dysfunctions of SH2 and/or SH3 domains reduce auto-phosphorylation of the kinase activation loop, depress kinase activity, and decrease NMDAR phosphorylation. The SH2 domain plays a greater regulatory role than the SH3 domain. Our data also show that n-Src binds directly to the C-terminus of the NMDAR NR2A subunit in vitro, with a K(D) of 108.2 ± 13.3 nM. This binding is not Src kinase activity-dependent, and dysfunctions of the SH2 and/or SH3 domains do not significantly affect the binding. These data indicate that the SH2 and SH3 domains may function to promote the catalytic activity of active n-Src, which is important in the regulation of NMDAR functions. © 2010 The Authors Journal compilation © 2010 FEBS.

Formation and reduction of 3-monochloropropane-1,2-diol esters in peanut oil during physical refining.

PubMed

Li, Chang; Li, Linyan; Jia, Hanbing; Wang, Yuting; Shen, Mingyue; Nie, Shaoping; Xie, Mingyong

2016-05-15

In the present study, lab-scale physical refining processes were investigated for their effects on the formation of 3-monochloropropane-1,2-diol (3-MCPD) esters. The potential precursors, partial acylglycerols and chlorines were determined before each refining step. 3-MCPD esters were not detected in degummed and bleached oil when the crude oils were extracted by solvent. While in the hot squeezed crude oils, 3-MCPD esters were detected with low amounts. 3-MCPD esters were generated with maximum values in 1-1.5h at a certain deodorizing temperature (220-260°C). Chlorine seemed to be more effective precursor than partial acylglycerol. By washing bleached oil before deodorization with ethanol solution, the precursors were removed partially and the content of 3-MCPD esters decreased to some extent accordingly. Diacetin was found to reduce 3-MCPD esters effectively. Copyright © 2015 Elsevier Ltd. All rights reserved.

NEW METABOLITES FROM THE MICROBIAL OXIDATION OF FLUORINATED AROMATIC COMPOUNDS. (R826113)

EPA Science Inventory

Abstract

m-Bromo-,,A role for Pyk2 and Src in linking G-protein-coupled receptors with MAP kinase activation.

PubMed

Dikic, I; Tokiwa, G; Lev, S; Courtneidge, S A; Schlessinger, J

1996-10-10

The mechanisms by which mitogenic G-protein-coupled receptors activate the MAP kinase signalling pathway are poorly understood. Candidate protein tyrosine kinases that link G-protein-coupled receptors with MAP kinase include Src family kinases, the epidermal growth factor receptor, Lyn and Syk. Here we show that lysophosphatidic acid (LPA) and bradykinin induce tyrosine phosphorylation of Pyk2 and complex formation between Pyk2 and activated Src. Moreover, tyrosine phosphorylation of Pyk2 leads to binding of the SH2 domain of Src to tyrosine 402 of Pyk2 and activation of Src. Transient overexpression of a dominant interfering mutant of Pyk2 or the protein tyrosine kinase Csk reduces LPA- or bradykinin-induced activation of MAP kinase. LPA- or bradykinin-induced MAP kinase activation was also inhibited by overexpression of dominant interfering mutants of Grb2 and Sos. We propose that Pyk2 acts with Src to link Gi- and Gq-coupled receptors with Grb2 and Sos to activate the MAP kinase signalling pathway in PC12 cells.

C-terminal Src kinase (Csk) regulates the tricellular junction protein Gliotactin independent of Src

PubMed Central

Samarasekera, G. D. N. Gayathri; Auld, Vanessa Jane

2018-01-01

Tricellular junctions (TCJs) are uniquely placed permeability barriers formed at the corners of polarized epithelia where tight junctions in vertebrates or septate junctions (SJ) in invertebrates from three cells converge. Gliotactin is a Drosophila TCJ protein, and loss of Gliotactin results in SJ and TCJ breakdown and permeability barrier loss. When overexpressed, Gliotactin spreads away from the TCJs, resulting in disrupted epithelial architecture, including overproliferation, cell delamination, and migration. Gliotactin levels are tightly controlled at the mRNA level and at the protein level through endocytosis and degradation triggered by tyrosine phosphorylation. We identified C-terminal Src kinase (Csk) as a tyrosine kinase responsible for regulating Gliotactin endocytosis. Increased Csk suppresses the Gliotactin overexpression phenotypes by increasing endocytosis. Loss of Csk causes Gliotactin to spread away from the TCJ. Although Csk is known as a negative regulator of Src kinases, the effects of Csk on Gliotactin are independent of Src and likely occur through an adherens junction associated complex. Overall, we identified a new Src-independent role for Csk in the control of Gliotactin, a key tricellular junction protein. PMID:29167383

Simian Immunodeficiency Virus and Human Immunodeficiency Virus Type 1 Nef Proteins Show Distinct Patterns and Mechanisms of Src Kinase Activation

PubMed Central

Greenway, Alison L.; Dutartre, Hélène; Allen, Kelly; McPhee, Dale A.; Olive, Daniel; Collette, Yves

1999-01-01

The nef gene from human and simian immunodeficiency viruses (HIV and SIV) regulates cell function and viral replication, possibly through binding of the nef product to cellular proteins, including Src family tyrosine kinases. We show here that the Nef protein encoded by SIVmac239 interacts with and also activates the human Src kinases Lck and Hck. This is in direct contrast to the inhibitory effect of HIV type 1 (HIV-1) Nef on Lck catalytic activity. Unexpectedly, however, the interaction of SIV Nef with human Lck or Hck is not mediated via its consensus proline motif, which is known to mediate HIV-1 Nef binding to Src homology 3 (SH3) domains, and various experimental analyses failed to show significant interaction of SIV Nef with the SH3 domain of either kinase. Instead, SIV Nef can bind Lck and Hck SH2 domains, and its N-terminal 50 amino acid residues are sufficient for Src kinase binding and activation. Our results provide evidence for multiple mechanisms by which Nef binds to and regulates Src kinases. PMID:10364375

Anxiety and Mood Clinical Profile following Sport-related Concussion: From Risk Factors to Treatment.

PubMed

Sandel, Natalie; Reynolds, Erin; Cohen, Paul E; Gillie, Brandon L; Kontos, Anthony P

2017-08-01

Conceptual models for assessing and treating sport-related concussion (SRC) have evolved from a homogenous approach to include different clinical profiles that reflect the heterogeneous nature of this injury and its effects. There are six identified clinical profiles, or subtypes from SRC, and one such clinical profile is the anxiety/mood profile. Athletes with this profile experience predominant emotional disturbance and anxiety following SRC. The purpose of this targeted review was to present an overview of the empirical evidence to support factors contributing to the anxiety/mood profile, along with methods of evaluation and treatment of this clinical profile following SRC. We discuss the potential underlying mechanisms and risk factors for this clinical profile, describe comprehensive assessments to evaluate concussed athletes with an anxiety/mood clinical profile, and explore behavioral and other interventions for treating these athletes. Although there is limited, but growing empirical evidence for the anxiety/mood clinical profile following SRC, understanding this clinical profile is germane for clinicians who are treating athletes with emotional sequelae after SRC.

Interaction between src family kinases and rho-kinase in agonist-induced Ca2+-sensitization of rat pulmonary artery.

PubMed

Knock, Greg A; Shaifta, Yasin; Snetkov, Vladimir A; Vowles, Benjamin; Drndarski, Svetlana; Ward, Jeremy P T; Aaronson, Philip I

2008-02-01

We investigated the role of src family kinases (srcFK) in agonist-mediated Ca2+-sensitization in pulmonary artery and whether this involves interaction with the rho/rho-kinase pathway. Intra-pulmonary arteries (IPAs) and cultured pulmonary artery smooth muscle cells (PASMC) were obtained from rat. Expression of srcFK was determined at the mRNA and protein levels. Ca2+-sensitization was induced by prostaglandin F(2 alpha) (PGF(2 alpha)) in alpha-toxin-permeabilized IPAs. Phosphorylation of the regulatory subunit of myosin phosphatase (MYPT-1) and of myosin light-chain-20 (MLC20) and translocation of rho-kinase in response to PGF(2 alpha) were also determined. Nine srcFK were expressed at the mRNA level, including src, fyn, and yes, and PGF(2 alpha) enhanced phosphorylation of three srcFK proteins at tyr-416. In alpha-toxin-permeabilized IPAs, PGF(2 alpha) enhanced the Ca2+-induced contraction (pCa 6.9) approximately three-fold. This enhancement was inhibited by the srcFK blockers SU6656 and PP2 and by the rho-kinase inhibitor Y27632. Y27632, but not SU6656 or PP2, also inhibited the underlying pCa 6.9 contraction. PGF(2 alpha) enhanced phosphorylation of MYPT-1 at thr-697 and thr-855 and of MLC20 at ser-19. This enhancement, but not the underlying basal phosphorylation, was inhibited by SU6656. Y27632 suppressed both basal and PGF(2 alpha)-mediated phosphorylation. The effects of SU6656 and Y27632, on both contraction and MYPT-1 and MLC20 phosphorylation, were not additive. PGF(2 alpha) triggered translocation of rho-kinase in PASMC, and this was inhibited by SU6656. srcFK are activated by PGF(2 alpha) in the rat pulmonary artery and may contribute to Ca2+-sensitization and contraction via rho-kinase translocation and phosphorylation of MYPT-1.

Cell Signaling Associated with Na+/K+-ATPase: Activation of Phosphatidylinositide 3-Kinase IA/Akt by Ouabain Is Independent of Src

PubMed Central

2013-01-01

Exposure of intact cells to selective inhibitors of Na+/K+-ATPase such as ouabain activates several growth-related cell signaling pathways. It has been suggested that the initial event of these pathways is the binding of ouabain to a preexisting complex of Src with Na+/K+-ATPase of the plasma membrane. The aim of this work was to evaluate the role of Src in the ouabain-induced activation of phosphatidylinositide 3-kinase 1A (PI3K1A) and its downstream consequences. When fibroblasts devoid of Src (SYF cells) and controls (Src++ cells) were exposed to ouabain, PI3K1A, Akt, and proliferative growth were similarly stimulated in both cell lines. Ouabain-induced activation of Akt was not prevented by the Src inhibitor PP2. In contrast, ERK1/2 were not activated by ouabain in SYF cells but were stimulated in Src++ cells; this was prevented by PP2. In isolated adult mouse cardiac myocytes, where ouabain induces hypertrophic growth, PP2 also did not prevent ouabain-induced activation of Akt and the resulting hypertrophy. Ouabain-induced increases in the levels of co-immunoprecipitation of the α-subunit of Na+/K+-ATPase with the p85 subunit of PI3K1A were noted in SYF cells, Src++ cells, and adult cardiac myocytes. In conjunction with previous findings, the results presented here indicate that (a) if there is a preformed complex of Src and Na+/K+-ATPase, it is irrelevant to ouabain-induced activation of the PI3K1A/Akt pathway through Na+/K+-ATPase and (b) a more likely, but not established, mechanism of linkage of Na+/K+-ATPase to PI3K1A is the ouabain-induced interaction of a proline-rich domain of the α-subunit of Na+/K+-ATPase with the SH3 domain of the p85 subunit of PI3K1A. PMID:24266852

Src Drives Growth of Antiestrogen Resistant Breast Cancer Cell Lines and Is a Marker for Reduced Benefit of Tamoxifen Treatment

PubMed Central

Larsen, Sarah L.; Laenkholm, Anne-Vibeke; Duun-Henriksen, Anne Katrine; Bak, Martin; Lykkesfeldt, Anne E.; Kirkegaard, Tove

2015-01-01

The underlying mechanisms leading to antiestrogen resistance in estrogen-receptor α (ER)-positive breast cancer is still poorly understood. The aim of this study was therefore to identify biomarkers and novel treatments for antiestrogen resistant breast cancer. We performed a kinase inhibitor screen on antiestrogen responsive T47D breast cancer cells and T47D-derived tamoxifen and fulvestrant resistant cell lines. We found that dasatinib, a broad-spectrum kinase inhibitor, inhibited growth of the antiestrogen resistant cells compared to parental T47D cells. Furthermore western blot analysis showed increased expression and phosphorylation of Src in the resistant cells and that dasatinib inhibited phosphorylation of Src and also signaling via Akt and Erk in all cell lines. Immunoprecipitation revealed Src: ER complexes only in the parental T47D cells. In fulvestrant resistant cells, Src formed complexes with the Human Epidermal growth factor Receptor (HER)1 and HER2. Neither HER receptors nor ER were co-precipitated with Src in the tamoxifen resistant cell lines. Compared to treatment with dasatinib alone, combined treatment with dasatinib and fulvestrant had a stronger inhibitory effect on tamoxifen resistant cell growth, whereas dasatinib in combination with tamoxifen had no additive inhibitory effect on fulvestrant resistant growth. When performing immunohistochemical staining on 268 primary tumors from breast cancer patients who had received tamoxifen as first line endocrine treatment, we found that membrane expression of Src in the tumor cells was significant associated with reduced disease-free and overall survival. In conclusion, Src was identified as target for treatment of antiestrogen resistant T47D breast cancer cells. For tamoxifen resistant T47D cells, combined treatment with dasatinib and fulvestrant was superior to treatment with dasatinib alone. Src located at the membrane has potential as a new biomarker for reduced benefit of tamoxifen. PMID:25706943

Interaction between src family kinases and rho-kinase in agonist-induced Ca2+-sensitization of rat pulmonary artery

PubMed Central

Knock, Greg A.; Shaifta, Yasin; Snetkov, Vladimir A.; Vowles, Benjamin; Drndarski, Svetlana; Ward, Jeremy P.T.; Aaronson, Philip I.

2008-01-01

Abstract Aims We investigated the role of src family kinases (srcFK) in agonist-mediated Ca2+-sensitization in pulmonary artery and whether this involves interaction with the rho/rho-kinase pathway. Methods and results Intra-pulmonary arteries (IPAs) and cultured pulmonary artery smooth muscle cells (PASMC) were obtained from rat. Expression of srcFK was determined at the mRNA and protein levels. Ca2+-sensitization was induced by prostaglandin F2α (PGF2α) in α-toxin-permeabilized IPAs. Phosphorylation of the regulatory subunit of myosin phosphatase (MYPT-1) and of myosin light-chain-20 (MLC20) and translocation of rho-kinase in response to PGF2α were also determined. Nine srcFK were expressed at the mRNA level, including src, fyn, and yes, and PGF2α enhanced phosphorylation of three srcFK proteins at tyr-416. In α-toxin-permeabilized IPAs, PGF2α enhanced the Ca2+-induced contraction (pCa 6.9) approximately three-fold. This enhancement was inhibited by the srcFK blockers SU6656 and PP2 and by the rho-kinase inhibitor Y27632. Y27632, but not SU6656 or PP2, also inhibited the underlying pCa 6.9 contraction. PGF2α enhanced phosphorylation of MYPT-1 at thr-697 and thr-855 and of MLC20 at ser-19. This enhancement, but not the underlying basal phosphorylation, was inhibited by SU6656. Y27632 suppressed both basal and PGF2α-mediated phosphorylation. The effects of SU6656 and Y27632, on both contraction and MYPT-1 and MLC20 phosphorylation, were not additive. PGF2α triggered translocation of rho-kinase in PASMC, and this was inhibited by SU6656. Conclusions srcFK are activated by PGF2α in the rat pulmonary artery and may contribute to Ca2+-sensitization and contraction via rho-kinase translocation and phosphorylation of MYPT-1. PMID:18032393

High-performance liquid chromatography with fluorescence detection for the rapid analysis of pheophytins and pyropheophytins in virgin olive oil.

PubMed

Li, Xueqi; Woodman, Michael; Wang, Selina C

2015-08-01

Pheophytins and pyropheophytin are degradation products of chlorophyll pigments, and their ratios can be used as a sensitive indicator of stress during the manufacturing and storage of olive oil. They increase over time depending on the storage condition and if the oil is exposed to heat treatments during the refining process. The traditional analysis method includes solvent- and time-consuming steps of solid-phase extraction followed by analysis by high-performance liquid chromatography with ultraviolet detection. We developed an improved dilute/fluorescence method where multi-step sample preparation was replaced by a simple isopropanol dilution before the high-performance liquid chromatography injection. A quaternary solvent gradient method was used to include a fourth strong solvent wash on a quaternary gradient pump, which avoided the need to premix any solvents and greatly reduced the oil residues on the column from previous analysis. This new method not only reduces analysis cost and time but shows reliability, repeatability, and improved sensitivity, especially important for low-level samples. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Chronic traumatic encephalopathy in sports: a historical and narrative review.

PubMed

Solomon, Gary

2018-01-01

My objectives are to review: 1) a brief history of sport-related concussion (SRC) and chronic traumatic encephalopathy (CTE), 2) the evolution of CTE in American professional football, 3) the data regarding SRC/CTE as they relate to depression and suicide, 4) the data on the neurocognitive effects of subconcussion/repetitive head trauma (with emphases on heading the ball in soccer and early exposure to football), 5) the evidence related to SRC and neurodegenerative diseases, 6) the published studies of CTE, 7) the NINDS neuropathological criteria for CTE, 8) public beliefs about SRC/CTE, and 9) the scientific questions regarding CTE.

Genetic and Environmental Models of Circadian Disruption Link SRC-2 Function to Hepatic Pathology

PubMed Central

Fleet, Tiffany; Stashi, Erin; Zhu, Bokai; Rajapakshe, Kimal; Marcelo, Kathrina L.; Kettner, Nicole M.; Gorman, Blythe K.; Coarfa, Cristian; Fu, Loning; O’Malley, Bert W.; York, Brian

2017-01-01

Circadian rhythmicity is a fundamental process that synchronizes behavioral cues with metabolic homeostasis. Disruption of daily cycles due to jet lag or shift work results in severe physiological consequences including advanced aging, metabolic syndrome, and even cancer. Our understanding of the molecular clock, which is regulated by intricate positive feedforward and negative feedback loops, has expanded to include an important metabolic transcriptional coregulator, Steroid Receptor Coactivator-2 (SRC-2), that regulates both the central clock of the suprachiasmatic nucleus (SCN) and peripheral clocks including the liver. We hypothesized that an environmental uncoupling of the light-dark phases, termed chronic circadian disruption (CCD), would lead to pathology similar to the genetic circadian disruption observed with loss of SRC-2. We found that CCD and ablation of SRC-2 in mice led to a common comorbidity of metabolic syndrome also found in humans with circadian disruption, non-alcoholic fatty liver disease (NAFLD). The combination of SRC-2−/− and CCD results in a more robust phenotype that correlates with human non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) gene signatures. Either CCD or SRC-2 ablation produces an advanced aging phenotype leading to increased mortality consistent with other circadian mutant mouse models. Collectively, our studies demonstrate that SRC-2 provides an essential link between the behavioral activities influenced by light cues and the metabolic homeostasis maintained by the liver. PMID:27432117

Combining Src inhibitors and aromatase inhibitors: a novel strategy for overcoming endocrine resistance and bone loss.

PubMed

Hiscox, Stephen; Barrett-Lee, Peter; Borley, Annabel C; Nicholson, Robert I

2010-08-01

Aromatase inhibitors have largely replaced tamoxifen as the first-line treatment for postmenopausal women with metastatic, hormone receptor-positive (HR+) breast cancer. However, many patients develop clinical resistance with prolonged treatment, and oestrogen deprivation following aromatase inhibition can result in loss of bone mineral density. Furthermore, most patients with metastatic breast cancer develop bone metastases, and the resulting adverse skeletal-related events are a significant cause of patient morbidity. Src, a non-receptor tyrosine kinase, is a component of signalling pathways that regulate breast cancer cell proliferation, invasion and metastasis as well as osteoclast-mediated bone turnover. Preclinical evidence also suggests a role for Src in acquired endocrine resistance. As such, Src inhibition represents a logical strategy for the treatment of metastatic breast cancer. In vitro, combination therapy with Src inhibitors and endocrine agents, including aromatase inhibitors, has been shown to inhibit the proliferation and metastasis of both endocrine-responsive and endocrine-resistant breast cancer cell lines more effectively than either of the therapy alone. Src inhibition has also been shown to suppress osteoclast formation and activity. Combination therapy with aromatase inhibitors and Src inhibitors therefore represents a novel approach through which the development of both acquired resistance and bone pathology could be delayed. Data from clinical trials utilising such combinations will reveal if this strategy has the potential to improve patient outcomes. Copyright 2010 Elsevier Ltd. All rights reserved.

Identification of N-Terminal Lobe Motifs that Determine the Kinase Activity of the Catalytic Domains and Regulatory Strategies of Src and Csk Protein Tyrosine Kinases†

PubMed Central

Huang, Kezhen; Wang, Yue-Hao; Brown, Alex; Sun, Gongqin

2009-01-01

Csk and Src protein tyrosine kinases are structurally homologous, but use opposite regulatory strategies. The isolated catalytic domain of Csk is intrinsically inactive and is activated by interactions with the regulatory SH3 and SH2 domains, while the isolated catalytic domain of Src is intrinsically active and is suppressed by interactions with the regulatory SH3 and SH2 domains. The structural basis for why one isolated catalytic domain is intrinsically active while the other is inactive is not clear. In this current study, we identify the structural elements in the N-terminal lobe of the catalytic domain that render the Src catalytic domain active. These structural elements include the α-helix C region, a β-turn between the β-4 and β-5 strands, and an Arg residue at the beginning of the catalytic domain. These three motifs interact with each other to activate the Src catalytic domain, but the equivalent motifs in Csk directly interact with the regulatory domains that are important for Csk activation. The Src motifs can be grafted to the Csk catalytic domain to obtain an active Csk catalytic domain. These results, together with available Src and Csk tertiary structures, reveal an important structural switch that determines the kinase activity of a catalytic domain and dictates the regulatory strategy of a kinase. PMID:19244618

A RNA Interference Screen Identifies the Protein Phosphatase 2A Subunit PR55γ as a Stress-Sensitive Inhibitor of c-SRC

PubMed Central

Eichhorn, Pieter J. A; Creyghton, Menno P; Wilhelmsen, Kevin; van Dam, Hans; Bernards, René

2007-01-01

Protein Phosphatase type 2A (PP2A) represents a family of holoenzyme complexes with diverse biological activities. Specific holoenzyme complexes are thought to be deregulated during oncogenic transformation and oncogene-induced signaling. Since most studies on the role of this phosphatase family have relied on the use of generic PP2A inhibitors, the contribution of individual PP2A holoenzyme complexes in PP2A-controlled signaling pathways is largely unclear. To gain insight into this, we have constructed a set of shRNA vectors targeting the individual PP2A regulatory subunits for suppression by RNA interference. Here, we identify PR55γ and PR55δ as inhibitors of c-Jun NH2-terminal kinase (JNK) activation by UV irradiation. We show that PR55γ binds c-SRC and modulates the phosphorylation of serine 12 of c-SRC, a residue we demonstrate to be required for JNK activation by c-SRC. We also find that the physical interaction between PR55γ and c-SRC is sensitive to UV irradiation. Our data reveal a novel mechanism of c-SRC regulation whereby in response to stress c-SRC activity is regulated, at least in part, through loss of the interaction with its inhibitor, PR55γ. PMID:18069897

Unified interpretation of exciplex formation and marcus electron transfer on the basis of two-dimensional free energy surfaces.

PubMed

Murata, Shigeo; Tachiya, M

2007-09-27

The mechanism of exciplex formation proposed in a previous paper has been refined to show how exciplex formation and Marcus electron transfer (ET) in fluorescence quenching are related to each other. This was done by making simple calculations of the free energies of the initial (DA*) and final (D+A-) states of ET. First it was shown that the decrease in D-A distance can induce intermolecular ET even in nonpolar solvents where solvent orientational polarization is absent, and that it leads to exciplex formation. This is consistent with experimental results that exciplex is most often observed in nonpolar solvents. The calculation was then extended to ET in polar solvents where the free energies are functions of both D-A distance and solvent orientational polarization. This enabled us to discuss both exciplex formation and Marcus ET in the same D-A pair and solvent on the basis of 2-dimensional free energy surfaces. The surfaces contain more information about the rates of these reactions, the mechanism of fluorescence quenching by ET, etc., than simple reaction schemes. By changing the parameters such as the free energy change of reaction, solvent dielectric constants, etc., one can construct the free energy surfaces for various systems. The effects of free energy change of reaction and of solvent polarity on the mechanism and relative importance of exciplex formation and Marcus ET in fluorescence quenching can be well explained. The free energy surface will also be useful for discussion of other phenomena related to ET reactions.

Principles and Practices of Enhanced Anaerobic Bioremediation of Chlorinated Solvents

DTIC Science & Technology

2004-09-01

high - fructose corn syrup (HFCS), whey, bark mulch and compost, chitin, and gaseous hydrogen. Table 1.2...Benzoate Injection wells or circulation systems Dissolved in water Continuous to monthly Molasses, High Fructose Corn Syrup Injection wells...to 0.35 High (> 100) Refined Sugars ( high fructose corn syrup ) 0.25 to 0.30 Moderate (> 20) Soluble substrates may be used for source

Cookie- versus cracker-baking--what's the difference? Flour functionality requirements explored by SRC and alveography.

PubMed

Kweon, Meera; Slade, Louise; Levine, Harry; Gannon, Diane

2014-01-01

The many differences between cookie- and cracker-baking are discussed and described in terms of the functionality, and functional requirements, of the major biscuit ingredients--flour and sugar. Both types of products are similar in their major ingredients, but different in their formulas and processes. One of the most important and consequential differences between traditional cracker and cookie formulas is sugar (i.e., sucrose) concentration: usually lower than 30% in a typical cracker formula and higher than 30% in a typical cookie formula. Gluten development is facilitated in lower-sugar cracker doughs during mixing and sheeting; this is a critical factor linked to baked-cracker quality. Therefore, soft wheat flours with greater gluten quality and strength are typically preferred for cracker production. In contrast, the concentrated aqueous sugar solutions existing in high-sugar cookie doughs generally act as an antiplasticizer, compared with water alone, so gluten development during dough mixing and starch gelatinization/pasting during baking are delayed or prevented in most cookie systems. Traditional cookies and crackers are low-moisture baked goods, which are desirably made from flours with low water absorption [low water-holding capacity (WHC)], and low levels of damaged starch and water-soluble pentosans (i.e., water-accessible arabinoxylans). Rheological (e.g., alveography) and baking tests are often used to evaluate flour quality for baked-goods applications, but the solvent retention capacity (SRC) method (AACC 56-11) is a better diagnostic tool for predicting the functional contribution of each individual flour functional component, as well as the overall functionality of flours for cookie- and/or cracker-baking.

Expression, Purification, Crystallisation and X-ray Crystallographic Analysis of a Truncated Form of Human Src Homology 2 Containing Inositol 5-Phosphatase 2.

PubMed

Le Coq, Johanne; Heredia Gallego, Luis; Lietha, Daniel

2016-06-01

The Src homology 2 containing inositol 5-phosphatase 2 (SHIP2) catalyses the dephosphorylation of the phospholipid phosphatidylinositol 3,4,5-triphosphate (PI(3,4,5)P3) to form PI(3,4)P2. PI(3,4,5)P3 is a key lipid second messenger, which can recruit signalling proteins to the plasma membrane and subsequently initiate numerous downstream signalling pathways responsible for the regulation of a plethora of cellular events such as proliferation, growth, apoptosis and cytoskeletal rearrangements. SHIP2 has been heavily implicated with several serious diseases such as cancer and type 2 diabetes but its regulation remains poorly understood. In order to gain insight into the mechanisms of SHIP2 regulation, a fragment of human SHIP2 containing the phosphatase domain and a region proposed to resemble a C2 domain was crystallized. Currently, no structural information is available on the putative C2-related domain or its relative position with respect to the phosphatase domain. Initial crystals were polycrystalline, but were optimized to obtain diffraction data to a resolution of 2.1 Å. Diffraction data analysis revealed a P212121 space group with unit cell parameters a = 136.04 Å, b = 175.84 Å, c = 176.89 Å. The Matthews coefficient is 2.54 Å(3) Da(-1) corresponding to 8 molecules in the asymmetric unit with a solvent content of 51.7 %.

Multiple Sparse Representations Classification

PubMed Central

Plenge, Esben; Klein, Stefan S.; Niessen, Wiro J.; Meijering, Erik

2015-01-01

Sparse representations classification (SRC) is a powerful technique for pixelwise classification of images and it is increasingly being used for a wide variety of image analysis tasks. The method uses sparse representation and learned redundant dictionaries to classify image pixels. In this empirical study we propose to further leverage the redundancy of the learned dictionaries to achieve a more accurate classifier. In conventional SRC, each image pixel is associated with a small patch surrounding it. Using these patches, a dictionary is trained for each class in a supervised fashion. Commonly, redundant/overcomplete dictionaries are trained and image patches are sparsely represented by a linear combination of only a few of the dictionary elements. Given a set of trained dictionaries, a new patch is sparse coded using each of them, and subsequently assigned to the class whose dictionary yields the minimum residual energy. We propose a generalization of this scheme. The method, which we call multiple sparse representations classification (mSRC), is based on the observation that an overcomplete, class specific dictionary is capable of generating multiple accurate and independent estimates of a patch belonging to the class. So instead of finding a single sparse representation of a patch for each dictionary, we find multiple, and the corresponding residual energies provides an enhanced statistic which is used to improve classification. We demonstrate the efficacy of mSRC for three example applications: pixelwise classification of texture images, lumen segmentation in carotid artery magnetic resonance imaging (MRI), and bifurcation point detection in carotid artery MRI. We compare our method with conventional SRC, K-nearest neighbor, and support vector machine classifiers. The results show that mSRC outperforms SRC and the other reference methods. In addition, we present an extensive evaluation of the effect of the main mSRC parameters: patch size, dictionary size, and sparsity level. PMID:26177106

Steroid receptor coactivator-1 mediates letrozole induced downregulation of postsynaptic protein PSD-95 in the hippocampus of adult female rats.

PubMed

Liu, Mengying; Huangfu, Xuhong; Zhao, Yangang; Zhang, Dongmei; Zhang, Jiqiang

2015-11-01

Hippocampus local estrogen which is converted from androgen that catalyzed by aromatase has been shown to play important roles in the regulation of learning and memory as well as cognition through action on synaptic plasticity, but the underlying mechanisms are poorly understood. Steroid receptor coactivator-1 (SRC-1) is one of the coactivators of steroid nuclear receptors; it is widely distributed in brain areas that related to learning and memory, reproductive regulation, sensory and motor information integration. Previous studies have revealed high levels of SRC-1 immunoreactivities in the hippocampus; it is closely related to the levels of synaptic proteins such as PSD-95 under normal development or gonadectomy, but its exact roles in the regulation of these proteins remains unclear. In this study, we used aromatase inhibitor letrozole in vivo and SRC-1 RNA interference in vitro to investigate whether SRC-1 mediated endogenous estrogen regulation of hippocampal PSD-95. The results revealed that letrozole injection synchronously decreased hippocampal SRC-1 and PSD-95 in a dose-dependant manner. Furthermore, when SRC-1 specific shRNA pool was applied to block the expression of SRC-1 in the primary hippocampal neuron culture, both immunocytochemistry and Western blot revealed that levels of PSD-95 were also decreased significantly. Taking together, these results provided the first evidence that SRC-1 mediated endogenous estrogen regulation of hippocampal synaptic plasticity by targeting the expression of synaptic protein PSD-95. Additionally, since letrozole is frequently used to treat estrogen-sensitive breast cancer, the above results also indicate its potential side effects in clinical administration. Copyright © 2015 Elsevier Ltd. All rights reserved.

All-trans-retinoic acid activates the pro-invasive Src-YAP-Interleukin 6 axis in triple-negative MDA-MB-231 breast cancer cells while cerivastatin reverses this action.

PubMed

Mezquita, Belén; Mezquita, Pau; Pau, Montserrat; Gasa, Laura; Navarro, Lourdes; Samitier, Mireia; Pons, Miquel; Mezquita, Cristóbal

2018-05-04

All-trans-retinoic acid (RA), the active metabolite of vitamin A, can reduce the malignant phenotype in some types of cancer and paradoxically also can promote cancer growth and invasion in others. For instance, it has been reported that RA induces tumor suppression in tumor xenografts of MDA-MB-468 breast cancer cells while increasing tumor growth and metastases in xenografts of MDA-MB-231 breast cancer cells. The signaling pathways involved in the pro-invasive action of retinoic acid remain mostly unknown. We show here that RA activates the pro-invasive axis Src-YAP-Interleukin 6 (Src-YAP-IL6) in triple negative MDA-MB-231 breast cancer cells, yielding to increased invasion of these cells. On the contrary, RA inhibits the Src-YAP-IL6 axis of triple-negative MDA-MB-468 cells, which results in decreased invasion phenotype. In both types of cells, inhibition of the Src-YAP-IL6 axis by the Src inhibitor PP2 drastically reduces migration and invasion. Src inhibition also downregulates the expression of a pro-invasive isoform of VEGFR1 in MDA-MB-231 breast cancer cells. Furthermore, interference of YAP nuclear translocation using the statin cerivastatin reverses the upregulation of Interleukin 6 (IL-6) and the pro-invasive effect of RA on MDA-MB-231 breast cancer cells and also decreases invasion and viability of MDA-MB-468 breast cancer cells. These results altogether suggest that RA induces pro-invasive or anti-invasive actions in two triple-negative breast cancer cell lines due to its ability to activate or inhibit the Src-YAP-IL6 axis in different cancer cells. The pro-invasive effect of RA can be reversed by the statin cerivastatin.

Hyaline Articular Matrix Formed by Dynamic Self-Regenerating Cartilage and Hydrogels.

PubMed

Meppelink, Amanda M; Zhao, Xing; Griffin, Darvin J; Erali, Richard; Gill, Thomas J; Bonassar, Lawrence J; Redmond, Robert W; Randolph, Mark A

2016-07-01

Injuries to the articular cartilage surface are challenging to repair because cartilage possesses a limited capacity for self-repair. The outcomes of current clinical procedures aimed to address these injuries are inconsistent and unsatisfactory. We have developed a novel method for generating hyaline articular cartilage to improve the outcome of joint surface repair. A suspension of 10(7) swine chondrocytes was cultured under reciprocating motion for 14 days. The resulting dynamic self-regenerating cartilage (dSRC) was placed in a cartilage ring and capped with fibrin and collagen gel. A control group consisted of chondrocytes encapsulated in fibrin gel. Constructs were implanted subcutaneously in nude mice and harvested after 6 weeks. Gross, histological, immunohistochemical, biochemical, and biomechanical analyses were performed. In swine patellar groove, dSRC was implanted into osteochondral defects capped with collagen gel and compared to defects filled with osteochondral plugs, collagen gel, or left empty after 6 weeks. In mice, the fibrin- and collagen-capped dSRC constructs showed enhanced contiguous cartilage matrix formation over the control of cells encapsulated in fibrin gel. Biochemically, the fibrin and collagen gel dSRC groups were statistically improved in glycosaminoglycan and hydroxyproline content compared to the control. There was no statistical difference in the biomechanical data between the dSRC groups and the control. The swine model also showed contiguous cartilage matrix in the dSRC group but not in the collagen gel and empty defects. These data demonstrate the survivability and successful matrix formation of dSRC under the mechanical forces experienced by normal hyaline cartilage in the knee joint. The results from this study demonstrate that dSRC capped with hydrogels successfully engineers contiguous articular cartilage matrix in both nonload-bearing and load-bearing environments.

Src mediates cigarette smoke-induced resistance to tyrosine kinase inhibitors in NSCLC cells.

PubMed

Filosto, Simone; Baston, David S; Chung, Samuel; Becker, Cathleen R; Goldkorn, Tzipora

2013-08-01

The EGF receptor (EGFR) is a proto-oncogene commonly dysregulated in several cancers including non-small cell lung carcinoma (NSCLC) and, thus, is targeted for treatment using tyrosine kinase inhibitors (TKI) such as erlotinib. However, despite the efficacy observed in patients with NSCLC harboring oncogenic variants of the EGFR, general ineffectiveness of TKIs in patients with NSCLC who are current and former smokers necessitates identification of novel mechanisms to overcome this phenomenon. Previously, we showed that NSCLC cells harboring either wild-type (WT) EGFR or oncogenic mutant (MT) L858R EGFR become resistant to the effects of TKIs when exposed to cigarette smoke, evidenced by their autophosphorylation and prolonged downstream signaling. Here, we present Src as a target mediating cigarette smoke-induced resistance to TKIs in both WT EGFR- and L858R MT EGFR-expressing NSCLC cells. First, we show that cigarette smoke exposure of A549 cells leads to time-dependent activation of Src, which then abnormally binds to the WT EGFR causing TKI resistance, contrasting previous observations of constitutive binding between inactive Src and TKI-sensitive L858R MT EGFR. Next, we show that Src inhibition restores TKI sensitivity in cigarette smoke-exposed NSCLC cells, preventing EGFR autophosphorylation in the presence of erlotinib. Furthermore, we show that overexpression of a dominant-negative Src (Y527F/K295R) restores TKI sensitivity to A549 exposed to cigarette smoke. Importantly, the TKI resistance that emerges even in cigarette smoke-exposed L858R EGFR-expressing NSCLC cells could be eliminated with Src inhibition. Together, these findings offer new rationale for using Src inhibitors for treating TKI-resistant NSCLC commonly observed in smokers.

Src mediates cigarette smoke-induced resistance to tyrosine kinase inhibitors in NSCLC cells

PubMed Central

Filosto, Simone; Baston, David S.; Chung, Samuel; Becker, Cathleen R.; Goldkorn, Tzipora

2015-01-01

The EGF Receptor (EGFR) is a proto-oncogene commonly dysregulated in several cancers including non-small cell lung cancer (NSCLC) and, thus, is targeted for treatment using tyrosine kinase inhibitors (TKIs) such as Erlotinib. However, despite the efficacy observed in NSCLC patients harboring oncogenic variants of the EGFR, general ineffectiveness of TKIs in NSCLC patients who are current and former smokers necessitates identification of novel mechanisms to overcome this phenomenon. Previously, we showed that NSCLC cells harboring either wild-type (WT) EGFR or oncogenic mutant (MT) L858R EGFR become resistant to the effects of TKIs when exposed to cigarette smoke (CS), evidenced by their auto-phosphorylation and prolonged downstream signaling. Here, we present Src as a target mediating CS-induced resistance to TKIs in both WT EGFR and L858R MT EGFR expressing NSCLC cells. First, we show that CS exposure of A549 cells leads to time-dependent activation of Src which then abnormally binds to the WT EGFR causing TKI resistance, contrasting previous observations of constitutive binding between inactive Src and TKI-sensitive L858R MT EGFR. Next, we demonstrate that Src inhibition restores TKI sensitivity in CS-exposed NSCLC cells, preventing EGFR auto-phosphorylation in the presence of Erlotinib. Furthermore, we show that over-expression of a dominant-negative Src (Y527F/K295R) restores TKI sensitivity to A549 exposed to CS. Importantly, the TKI resistance that emerges even in CS-exposed L858R EGFR expressing NSCLC cells could be eliminated with Src inhibition. Together, these findings offer new rationale for using Src inhibitors for treating TKI-resistant NSCLC commonly observed in smokers. PMID:23686837

A cell-penetrating peptide based on the interaction between c-Src and connexin43 reverses glioma stem cell phenotype

PubMed Central

Gangoso, E; Thirant, C; Chneiweiss, H; Medina, J M; Tabernero, A

2014-01-01

Connexin43 (Cx43), the main gap junction channel-forming protein in astrocytes, is downregulated in malignant gliomas. These tumors are composed of a heterogeneous population of cells that include many with stem-cell-like properties, called glioma stem cells (GSCs), which are highly tumorigenic and lack Cx43 expression. Interestingly, restoring Cx43 reverses GSC phenotype and consequently reduces their tumorigenicity. In this study, we investigated the mechanism by which Cx43 exerts its antitumorigenic effects on GSCs. We have focused on the tyrosine kinase c-Src, which interacts with the intracellular carboxy tail of Cx43. We found that Cx43 regulates c-Src activity and proliferation in human GSCs expanded in adherent culture. Thus, restoring Cx43 in GSCs inhibited c-Src activity, which in turn promoted the downregulation of the inhibitor of differentiation Id1. Id1 sustains stem cell phenotype as it controls the expression of Sox2, responsible for stem cell self-renewal, and promotes cadherin switching, which has been associated to epithelial–mesenchymal transition. Our results show that both the ectopic expression of Cx43 and the inhibition of c-Src reduced Id1, Sox2 expression and promoted the switch from N- to E-cadherin, suggesting that Cx43, by inhibiting c-Src, downregulates Id1 with the subsequent changes in stem cell phenotype. On the basis of this mechanism, we found that a cell-penetrating peptide, containing the region of Cx43 that interacts with c-Src, mimics the effect of Cx43 on GSC phenotype, confirming the relevance of the interaction between Cx43 and c-Src in the regulation of the malignant phenotype and pinpointing this interaction as a promising therapeutic target. PMID:24457967

Increased risk of kidney damage among Chinese adults with simple renal cyst.

PubMed

Kong, Xianglei; Ma, Xiaojing; Zhang, Chengyin; Su, Hong; Gong, Xiaojie; Xu, Dongmei

2018-05-04

The presence of simple renal cyst (SRC) has been related to hypertension, the early and long-term allograft function, and aortic disease, but the relationship with kidney damage was still controversial. Accordingly, we conducted a large sample cross-sectional study to explore the association of SRC with indicators of kidney damage among Chinese adults. A total of 42,369 adults (aged 45.8 ± 13.67 years, 70.6% males) who visited the Health Checkup Clinic were consecutively enrolled. SRC was assessed by ultrasonography according to Bosniak category. Multiple regression models were applied to explore the relationships between SRC and indicators of kidney damage [proteinuria (dipstick urine protein ≥ 1+) and decreased estimated glomerular filtration rate (DeGFR) < 60 ml/min/1.73 m 2 ]. Among all participants in the study, the prevalence of SRC was 10.5%. As a categorical outcome, participants with more 1 cyst and with 1 cyst had higher percentage of proteinuria [53 (5.3%) and 93 (2.7%) vs. 596 (1.6%), p < 0.001] and DeGFR [57 (5.7%) and 85 (2.5%) vs. 278 (0.7%), p < 0.001] compared with participants with no cyst. SRC significantly correlated with proteinuria [OR 1.59 (95% CI 1.30-1.95)] and DeGFR [OR 1.97 (95% CI 1.56-2.47)] after adjusting for potential confounders. Furthermore, the results also demonstrated that maximum diameter (per 1 cm increase), bilateral location, and multiple cysts significantly correlated with DeGFR in the multiple logistic regression analysis. The study revealed that SRC significantly correlated with kidney damage and special attention should be paid among Chinese adults with SRC.

Petroleum mineral oil refining and evaluation of cancer hazard.

PubMed

Mackerer, Carl R; Griffis, Larry C; Grabowski, John S; Reitman, Fred A

2003-11-01

Petroleum base oils (petroleum mineral oils) are manufactured from crude oils by vacuum distillation to produce several distillates and a residual oil that are then further refined. Aromatics including alkylated polycyclic aromatic compounds (PAC) are undesirable constituents of base oils because they are deleterious to product performance and are potentially carcinogenic. In modern base oil refining, aromatics are reduced by solvent extraction, catalytic hydrotreating, or hydrocracking. Chronic exposure to poorly refined base oils has the potential to cause skin cancer. A chronic mouse dermal bioassay has been the standard test for estimating carcinogenic potential of mineral oils. The level of alkylated 3-7-ring PAC in raw streams from the vacuum tower must be greatly reduced to render the base oil noncarcinogenic. The processes that can reduce PAC levels are known, but the operating conditions for the processing units (e.g., temperature, pressure, catalyst type, residence time in the unit, unit engineering design, etc.) needed to achieve adequate PAC reduction are refinery specific. Chronic dermal bioassays provide information about whether conditions applied can make a noncarcinogenic oil, but cannot be used to monitor current production for quality control or for conducting research or developing new processes since this test takes at least 78 weeks to conduct. Three short-term, non-animal assays all involving extraction of oil with dimethylsulfoxide (DMSO) have been validated for predicting potential carcinogenic activity of petroleum base oils: a modified Ames assay of a DMSO extract, a gravimetric assay (IP 346) for wt. percent of oil extracted into DMSO, and a GC-FID assay measuring 3-7-ring PAC content in a DMSO extract of oil, expressed as percent of the oil. Extraction with DMSO concentrates PAC in a manner that mimics the extraction method used in the solvent refining of noncarcinogenic oils. The three assays are described, data demonstrating the validation of the assays are shown, and test results of currently manufactured base oils are summarized to illustrate the general lack of cancer hazard for the base oils now being manufactured.

Aladdin: Transforming science at SRC

NASA Astrophysics Data System (ADS)

Bisognano, J.; Bissen, M.; Green, M.; Jacobs, K.; Moore, C.; Olson, E.; Severson, M.; Wehlitz, R.

2011-09-01

The Synchrotron Radiation Center (SRC) is dedicated to enabling of innovative research using IR, ultraviolet, and soft X-ray synchrotron radiation. It delivers beam time with high reliability (99%) and continues to improve the Aladdin storage ring complex. A lower emittance tuning has been commissioned to support a microfocus capability. SRC successfully installed an APPLE II undulator providing elliptically polarized light with lattice compensation for flexible scanning. Installation of a new IR beamline at SRC is providing synchrotron chemical imaging with unprecedented structural and chemical information, simultaneously. In addition, SRC has established a strong education and outreach program to bring the knowledge and power of light source science to a wider national community. It is moving forward into the future by developing a new micro focus beamline producing a diffraction-limited focus of about 500 nm at 22 eV, proposing an additional diffraction-limited chemical imaging beamline, and advancing the Wisconsin Free Electron Laser (WiFEL) concept.

CHLAMYDIA TRACHOMATIS TARP IS PHOSPHORYLATED BY SRC FAMILY TYROSINE KINASES

PubMed Central

Jewett, Travis J.; Dooley, Cheryl A.; Mead, David J.; Hackstadt, Ted

2008-01-01

The translocated actin recruiting phosphoprotein (Tarp) is injected into the cytosol shortly after Chlamydia trachomatis attachment to a target cell and subsequently phosphorylated by an unidentified tyrosine kinase. A role for Tarp phosphorylation in bacterial entry is unknown. In this study, recombinant C. trachomatis Tarp was employed to identify the host cell kinase(s) required for phosphorylation. Each tyrosine rich repeat of L2 Tarp harbors a sequence similar to a Src and Abl kinase consensus target. Furthermore, purified p60-src, Yes, Fyn, and Abl kinases were able to phosphorylate Tarp. Mutagenesis of potential tyrosines within a single tyrosine rich repeat peptide indicated that both Src and Abl kinases phosphorylate the same residues suggesting that C. trachomatis Tarp may serve as a substrate for multiple host cell kinases. Surprisingly, chemical inhibition of Src and Abl kinases prevented Tarp phosphorylation in culture and had no measurable effect on bacterial entry into host cells. PMID:18442471

Chromosomal localization of the mouse Src-like adapter protein (Slap) gene and its putative human homolog SLA

DOE Office of Scientific and Technical Information (OSTI.GOV)

Angrist, M.; Chakravarti, A.; Wells, D.E.

1995-12-10

Molecules containing Src-homology 2 (SH2) and Src-homology 3 (SH3) domains are critical components of signal transduction pathways that serve to relay signals originating from the cell surface to the interior of the cell. Src-like adapter protein (SLAP) is a recently described adapter protein that binds activated the Eck receptor protein-tyrosine kinase. Although SLAP bears a striking homology to the SH3 and SH2 domains of the Src family of nonreceptor tyrosine kinases, it does not contain a tyrosine kinase catalytic domain. In this report, the Slap gene was mapped by linkage analysis to mouse chromosome 15, while its putative human homologmore » (SLA) was identified and mapped to human 8q22.3-qter using a panel of somatic cell hybrids. 10 refs., 2 figs.« less

SRC Residual fuel oils

DOEpatents

Tewari, Krishna C.; Foster, Edward P.

1985-01-01

Coal solids (SRC) and distillate oils are combined to afford single-phase blends of residual oils which have utility as fuel oils substitutes. The components are combined on the basis of their respective polarities, that is, on the basis of their heteroatom content, to assure complete solubilization of SRC. The resulting composition is a fuel oil blend which retains its stability and homogeneity over the long term.

75 FR 65377 - Notice of Public Meeting for the National Park Service (NPS) Alaska Region's Subsistence Resource...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-10-22

... Resource Commission (SRC) program. SUMMARY: The Gates of the Arctic National Park SRC will meet to develop.... Gates of the Arctic National Park SRC Meeting Date and Location: The Gates of the Arctic National Park... meeting may end early if all business is completed. For Further Information On the Gates of the Arctic...

76 FR 1458 - Public Meeting for the National Park Service Alaska Region's Subsistence Resource Commission (SRC...

Federal Register 2010, 2011, 2012, 2013, 2014

2011-01-10

... Plan Update. c. Subsistence Uses of Horns, Antlers, Bones and Plants EA Update. 13. New Business. 14... guarantee that we will be able to do so. Wrangell-St. Elias National Park SRC Meeting Date and Location: The... if all business is completed. For Further Information on the Gates of the Arctic National Park SRC...

ERK3 signals through SRC-3 coactivator to promote human lung cancer cell invasion

PubMed Central

Long, Weiwen; Foulds, Charles E.; Qin, Jun; Liu, Jian; Ding, Chen; Lonard, David M.; Solis, Luisa M.; Wistuba, Ignacio I.; Qin, Jun; Tsai, Sophia Y.; Tsai, Ming-Jer; O’Malley, Bert W.

2012-01-01

In contrast to the well-studied classic MAPKs, such as ERK1/2, little is known concerning the regulation and substrates of the atypical MAPK ERK3 signaling cascade and its function in cancer progression. Here, we report that ERK3 interacted with and phosphorylated steroid receptor coactivator 3 (SRC-3), an oncogenic protein overexpressed in multiple human cancers at serine 857 (S857). This ERK3-mediated phosphorylation at S857 was essential for interaction of SRC-3 with the ETS transcription factor PEA3, which promotes upregulation of MMP gene expression and proinvasive activity in lung cancer cells. Importantly, knockdown of ERK3 or SRC-3 inhibited the ability of lung cancer cells to invade and form tumors in the lung in a xenograft mouse model. In addition, ERK3 was found to be highly upregulated in human lung carcinomas. Our study identifies a previously unknown role for ERK3 in promoting lung cancer cell invasiveness by phosphorylating SRC-3 and regulating SRC-3 proinvasive activity by site-specific phosphorylation. As such, ERK3 protein kinase may be an attractive target for therapeutic treatment of invasive lung cancer. PMID:22505454

Src is activated by the nuclear receptor peroxisome proliferator-activated receptor β/δ in ultraviolet radiation-induced skin cancer

PubMed Central

Montagner, Alexandra; Delgado, Maria B; Tallichet-Blanc, Corinne; Chan, Jeremy S K; Sng, Ming K; Mottaz, Hélène; Degueurce, Gwendoline; Lippi, Yannick; Moret, Catherine; Baruchet, Michael; Antsiferova, Maria; Werner, Sabine; Hohl, Daniel; Al Saati, Talal; Farmer, Pierre J; Tan, Nguan S; Michalik, Liliane; Wahli, Walter

2014-01-01

Although non-melanoma skin cancer (NMSC) is the most common human cancer and its incidence continues to rise worldwide, the mechanisms underlying its development remain incompletely understood. Here, we unveil a cascade of events involving peroxisome proliferator-activated receptor (PPAR) β/δ and the oncogene Src, which promotes the development of ultraviolet (UV)-induced skin cancer in mice. UV-induced PPARβ/δ activity, which directly stimulated Src expression, increased Src kinase activity and enhanced the EGFR/Erk1/2 signalling pathway, resulting in increased epithelial-to-mesenchymal transition (EMT) marker expression. Consistent with these observations, PPARβ/δ-null mice developed fewer and smaller skin tumours, and a PPARβ/δ antagonist prevented UV-dependent Src stimulation. Furthermore, the expression of PPARβ/δ positively correlated with the expression of SRC and EMT markers in human skin squamous cell carcinoma (SCC), and critically, linear models applied to several human epithelial cancers revealed an interaction between PPARβ/δ and SRC and TGFβ1 transcriptional levels. Taken together, these observations motivate the future evaluation of PPARβ/δ modulators to attenuate the development of several epithelial cancers. PMID:24203162

Saracatinib Impairs Head and Neck Squamous Cell Carcinoma Invasion by Disrupting Invadopodia Function

PubMed Central

Ammer, Amanda Gatesman; Kelley, Laura C.; Hayes, Karen E.; Evans, Jason V.; Lopez-Skinner, Lesly Ann; Martin, Karen H.; Frederick, Barbara; Rothschild, Brian L.; Raben, David; Elvin, Paul; Green, Tim P.; Weed, Scott A.

2010-01-01

Elevated Src kinase activity is linked to the progression of solid tumors, including head and neck squamous cell carcinoma (HNSCC). Src regulates HNSCC proliferation and tumor invasion, with the Src-targeted small molecule inhibitor saracatinib displaying potent anti-invasive effects in preclinical studies. However, the pro-invasive cellular mechanism(s) perturbed by saracatinib are unclear. The anti-proliferative and anti-invasive effects of saracatinib on HNSCC cell lines were therefore investigated in pre-clinical cell and mouse model systems. Saracatinib treatment inhibited growth, cell cycle progression and transwell Matrigel invasion in HNSCC cell lines. Dose-dependent decreases in Src activation and phosphorylation of the invasion-associated substrates focal adhesion kinase, p130 CAS and cortactin were also observed. While saracatinib did not significantly impact HNSCC tumor growth in a mouse orthotopic model of tongue squamous cell carcinoma, impaired perineural invasion and cervical lymph node metastasis was observed. Accordingly, saracatinib treatment displayed a dose-dependent inhibitory effect on invadopodia formation, extracellular matrix degradation and matrix metalloprotease 9 activation. These results suggest that inhibition of Src kinase by saracatinib impairs the pro-invasive activity of HNSCC by inhibiting Src substrate phosphorylation important for invadopodia formation and associated matrix metalloprotease activity. PMID:20505783

C-Cbl reverses HER2-mediated tamoxifen resistance in human breast cancer cells.

PubMed

Li, Wei; Xu, Ling; Che, Xiaofang; Li, Haizhou; Zhang, Ye; Song, Na; Wen, Ti; Hou, Kezuo; Yang, Yi; Zhou, Lu; Xin, Xing; Xu, Lu; Zeng, Xue; Shi, Sha; Liu, Yunpeng; Qu, Xiujuan; Teng, Yuee

2018-05-02

Tamoxifen is a frontline therapy for estrogen receptor (ER)-positive breast cancer in premenopausal women. However, many patients develop resistance to tamoxifen, and the mechanism underlying tamoxifen resistance is not well understood. Here we examined whether ER-c-Src-HER2 complex formation is involved in tamoxifen resistance. MTT and colony formation assays were used to measure cell viability and proliferation. Western blot was used to detect protein expression and protein complex formations were detected by immunoprecipitation and immunofluorescence. SiRNA was used to examine the function of HER2 in of BT474 cells. An in vivo xenograft animal model was established to examine the role of c-Cbl in tumor growth. MTT and colony formation assay showed that BT474 cells are resistant to tamoxifen and T47D cells are sensitive to tamoxifen. Immunoprecipitation experiments revealed ER-c-Src-HER2 complex formation in BT474 cells but not in T47D cells. However, ER-c-Src-HER2 complex formation was detected after overexpressing HER2 in T47D cells and these cells were more resistant to tamoxifen. HER2 knockdown by siRNA in BT474 cells reduced ER-c-Src-HER2 complex formation and reversed tamoxifen resistance. ER-c-Src-HER2 complex formation was also disrupted and tamoxifen resistance was reversed in BT474 cells by the c-Src inhibitor PP2 and HER2 antibody trastuzumab. Nystatin, a lipid raft inhibitor, reduced ER-c-Src-HER2 complex formation and partially reversed tamoxifen resistance. ER-c-Src-HER2 complex formation was disrupted by overexpression of c-Cbl but not by the c-Cbl ubiquitin ligase mutant. In addition, c-Cbl could reverse tamoxifen resistance in BT474 cells, but the ubiquitin ligase mutant had no effect. The effect of c-Cbl was validated in BT474 tumor-bearing nude mice in vivo. Immunofluorescence also revealed ER-c-Src-HER2 complex formation was reduced in tumor tissues of nude mice with c-Cbl overexpression. Our results suggested that c-Cbl can reverse tamoxifen resistance in HER2-overexpressing breast cancer cells by inhibiting the formation of the ER-c-Src-HER2 complex.

Comparison of oil refining and biodiesel production process between screw press and n-hexane techniques from beauty leaf feedstock

NASA Astrophysics Data System (ADS)

Bhuiya, M. M. K.; Rasul, M. G.; Khan, M. M. K.; Ashwath, N.

2016-07-01

The Beauty Leaf Tree (Callophylum inophyllum) is regarded as an alternative source of energy to produce 2nd generation biodiesel due to its potentiality as well as high oil yield content in the seed kernels. The treating process is indispensable during the biodiesel production process because it can augment the yield as well as quality of the product. Oil extracted from both mechanical screw press and solvent extraction using n-hexane was refined. Five replications each of 25 gm of crude oil for screw press and five replications each of 25 gm of crude oil for n-hexane were selected for refining as well as biodiesel conversion processes. The oil refining processes consists of degumming, neutralization as well as dewaxing. The degumming, neutralization and dewaxing processes were performed to remove all the gums (phosphorous-based compounds), free fatty acids, and waxes from the fresh crude oil before the biodiesel conversion process carried out, respectively. The results indicated that up to 73% and 81% of mass conversion efficiency of the refined oil in the screw press and n-hexane refining processes were obtained, respectively. It was also found that up to 88% and 90% of biodiesel were yielded in terms of mass conversion efficiency in the transesterification process for the screw press and n-hexane techniques, respectively. While the entire processes (refining and transesterification) were considered, the conversion of beauty leaf tree (BLT) refined oil into biodiesel was yielded up to 65% and 73% of mass conversion efficiency for the screw press and n-hexane techniques, respectively. Physico-chemical properties of crude and refined oil, and biodiesel were characterized according to the ASTM standards. Overall, BLT has the potential to contribute as an alternative energy source because of high mass conversion efficiency.

Determination of Microalgal Lipid Content and Fatty Acid for Biofuel Production

PubMed Central

Chen, Zhipeng; Wang, Lingfeng

2018-01-01

Biofuels produced from microalgal biomass have received growing worldwide recognition as promising alternatives to conventional petroleum-derived fuels. Among the processes involved, the downstream refinement process for the extraction of lipids from biomass greatly influences the sustainability and efficiency of the entire biofuel system. This review summarizes and compares the current techniques for the extraction and measurement of microalgal lipids, including the gravimetric methods using organic solvents, CO2-based solvents, ionic liquids and switchable solvents, Nile red lipid visualization method, sulfo-phospho-vanillin method, and the thin-layer chromatography method. Each method has its own competitive advantages and disadvantages. For example, the organic solvents-based gravimetric method is mostly used and frequently employed as a reference standard to validate other methods, but it requires large amounts of samples and is time-consuming and expensive to recover solvents also with low selectivity towards desired products. The pretreatment approaches which aimed to disrupt cells and support subsequent lipid extraction through bead beating, microwave, ultrasonication, chemical methods, and enzymatic disruption are also introduced. Moreover, the principles and procedures for the production and quantification of fatty acids are finally described in detail, involving the preparation of fatty acid methyl esters and their quantification and composition analysis by gas chromatography.

Soy Sauce Residue Oil Extracted by a Novel Continuous Phase Transition Extraction under Low Temperature and Its Refining Process.

PubMed

Zhao, Lichao; Zhang, Yong; He, Liping; Dai, Weijie; Lai, Yingyi; Yao, Xueyi; Cao, Yong

2014-04-09

On the basis of previous single-factor experiments, extraction parameters of soy sauce residue (SSR) oil extracted using a self-developed continuous phase transition extraction method at low temperature was optimized using the response surface methodology. The established optimal conditions for maximum oil yield were n-butane solvent, 0.5 MPa extraction pressure, 45 °C temperature, 62 min extraction time, and 45 mesh raw material granularity. Under these conditions, the actual yield was 28.43% ± 0.17%, which is relatively close to the predicted yield. Meanwhile, isoflavone was extracted from defatted SSR using the same method, but the parameters and solvent used were altered. The new solvent was 95% (v/v) ethanol, and extraction was performed under 1.0 MPa at 60 °C for 90 min. The extracted isoflavones, with 0.18% ± 0.012% yield, mainly comprised daidzein and genistein, two kinds of aglycones. The novel continuous phase transition extraction under low temperature could provide favorable conditions for the extraction of nonpolar or strongly polar substances. The oil physicochemical properties and fatty acids compositions were analyzed. Results showed that the main drawback of the crude oil was the excess of acid value (AV, 63.9 ± 0.1 mg KOH/g) and peroxide value (POV, 9.05 ± 0.3 mmol/kg), compared with that of normal soybean oil. However, through molecular distillation, AV and POV dropped to 1.78 ± 0.12 mg KOH/g and 5.9 ± 0.08 mmol/kg, respectively. This refined oil may be used as feedstuff oil.

Structural Refinement of Membrane Proteins by Restrained Molecular Dynamics and Solvent Accessibility Data

PubMed Central

Sompornpisut, Pornthep; Roux, Benoît; Perozo, Eduardo

2008-01-01

We present an approach for incorporating solvent accessibility data from electron paramagnetic resonance experiments in the structural refinement of membrane proteins through restrained molecular dynamics simulations. The restraints have been parameterized from oxygen (ΠO2) and nickel-ethylenediaminediacetic acid (ΠNiEdda) collision frequencies, as indicators of lipid or aqueous exposed spin-label sites. These are enforced through interactions between a pseudoatom representation of the covalently attached Nitroxide spin-label and virtual “solvent” particles corresponding to O2 and NiEdda in the surrounding environment. Interactions were computed using an empirical potential function, where the parameters have been optimized to account for the different accessibilities of the spin-label pseudoatoms to the surrounding environment. This approach, “pseudoatom-driven solvent accessibility refinement”, was validated by refolding distorted conformations of the Streptomyces lividans potassium channel (KcsA), corresponding to a range of 2–30 Å root mean-square deviations away from the native structure. Molecular dynamics simulations based on up to 58 electron paramagnetic resonance restraints derived from spin-label mutants were able to converge toward the native structure within 1–3 Å root mean-square deviations with minimal computational cost. The use of energy-based ranking and structure similarity clustering as selection criteria helped in the convergence and identification of correctly folded structures from a large number of simulations. This approach can be applied to a variety of integral membrane protein systems, regardless of oligomeric state, and should be particularly useful in calculating conformational changes from a known reference crystal structure. PMID:18676641

Longitudinal Brain Magnetic Resonance Imaging CO2 Stress Testing in Individual Adolescent Sports-Related Concussion Patients: A Pilot Study.

PubMed

Mutch, W Alan C; Ellis, Michael J; Ryner, Lawrence N; Morissette, Marc P; Pries, Philip J; Dufault, Brenden; Essig, Marco; Mikulis, David J; Duffin, James; Fisher, Joseph A

2016-01-01

Advanced neuroimaging studies in concussion have been limited to detecting group differences between concussion patients and healthy controls. In this small pilot study, we used brain magnetic resonance imaging (MRI) CO2 stress testing to longitudinally assess cerebrovascular responsiveness (CVR) in individual sports-related concussion (SRC) patients. Six SRC patients (three males and three females; mean age = 15.7, range = 15-17 years) underwent longitudinal brain MRI CO2 stress testing using blood oxygen level-dependent (BOLD) MRI and model-based prospective end-tidal CO2 targeting under isoxic conditions. First-level and second-level comparisons were undertaken using statistical parametric mapping (SPM) to score the scans and compare them to an atlas of 24 healthy control subjects. All tests were well tolerated and without any serious adverse events. Anatomical MRI was normal in all study participants. The CO2 stimulus was consistent between the SRC patients and control subjects and within SRC patients across the longitudinal study. Individual SRC patients demonstrated both quantitative and qualitative patient-specific alterations in CVR (p < 0.005) that correlated strongly with clinical findings, and that persisted beyond clinical recovery. Standardized brain MRI CO2 stress testing is capable of providing a longitudinal assessment of CVR in individual SRC patients. Consequently, larger prospective studies are needed to examine the utility of brain MRI CO2 stress testing as a clinical tool to help guide the evaluation, classification, and longitudinal management of SRC patients.

Accuracy of Monte Carlo photon transport simulation in characterizing brachytherapy dosimeter energy-response artefacts

NASA Astrophysics Data System (ADS)

Das, R. K.; Li, Z.; Perera, H.; Williamson, J. F.

1996-06-01

Practical dosimeters in brachytherapy, such as thermoluminescent dosimeters (TLD) and diodes, are usually calibrated against low-energy megavoltage beams. To measure absolute dose rate near a brachytherapy source, it is necessary to establish the energy response of the detector relative to that of the calibration energy. The purpose of this paper is to assess the accuracy of Monte Carlo photon transport (MCPT) simulation in modelling the absolute detector response as a function of detector geometry and photon energy. We have exposed two different sizes of TLD-100 (LiF chips) and p-type silicon diode detectors to calibrated , HDR source and superficial x-ray beams. For the Scanditronix electron-field diode, the relative detector response, defined as the measured detector readings per measured unit of air kerma, varied from (40 kVp beam) to ( beam). Similarly for the large and small chips the same quantity varied from and , respectively. Monte Carlo simulation was used to calculate the absorbed dose to the active volume of the detector per unit air kerma. If the Monte Carlo simulation is accurate, then the absolute detector response, which is defined as the measured detector reading per unit dose absorbed by the active detector volume, and is calculated by Monte Carlo simulation, should be a constant. For the diode, the absolute response is . For TLDs of size the absolute response is and for TLDs of it is . From the above results we can conclude that the absolute response function of detectors (TLDs and diodes) is directly proportional to absorbed dose by the active volume of the detector and is independent of beam quality.

A dynamic structural model of expanded RNA CAG repeats: A refined X-ray structure and computational investigations using molecular dynamics and umbrella sampling simulations

PubMed Central

Yildirim, Ilyas; Park, Hajeung; Disney, Matthew D.; Schatz, George C.

2013-01-01

One class of functionally important RNA is repeating transcripts that cause disease through various mechanisms. For example, expanded r(CAG) repeats can cause Huntington’s and other disease through translation of toxic proteins. Herein, crystal structure of r[5ʹUUGGGC(CAG)3GUCC]2, a model of CAG expanded transcripts, refined to 1.65 Å resolution is disclosed that show both anti-anti and syn-anti orientations for 1×1 nucleotide AA internal loops. Molecular dynamics (MD) simulations using Amber force field in explicit solvent were run for over 500 ns on model systems r(5ʹGCGCAGCGC)2 (MS1) and r(5ʹCCGCAGCGG)2 (MS2). In these MD simulations, both anti-anti and syn-anti AA base pairs appear to be stable. While anti-anti AA base pairs were dynamic and sampled multiple anti-anti conformations, no syn-anti↔anti-anti transformations were observed. Umbrella sampling simulations were run on MS2, and a 2D free energy surface was created to extract transformation pathways. In addition, over 800 ns explicit solvent MD simulation was run on r[5ʹGGGC(CAG)3GUCC]2, which closely represents the refined crystal structure. One of the terminal AA base pairs (syn-anti conformation), transformed to anti-anti conformation. The pathway followed in this transformation was the one predicted by umbrella sampling simulations. Further analysis showed a binding pocket near AA base pairs in syn-anti conformations. Computational results combined with the refined crystal structure show that global minimum conformation of 1×1 nucleotide AA internal loops in r(CAG) repeats is anti-anti but can adopt syn-anti depending on the environment. These results are important to understand RNA dynamic-function relationships and develop small molecules that target RNA dynamic ensembles. PMID:23441937

Endothelial Barrier Protection by Local Anesthetics: Ropivacaine and Lidocaine Block Tumor Necrosis Factor-α–induced Endothelial Cell Src Activation

PubMed Central

Piegeler, Tobias; Votta-Velis, E. Gina; Bakhshi, Farnaz R.; Mao, Mao; Carnegie, Graeme; Bonini, Marcelo G.; Schwartz, David E.; Borgeat, Alain; Beck-Schimmer, Beatrice; Minshall, Richard D.

2014-01-01

Background Pulmonary endothelial barrier dysfunction mediated in part by Src-kinase activation plays a crucial role in acute inflammatory disease. Proinflammatory cytokines, such as tumor necrosis factor-α (TNFα), activate Src via phosphatidylinositide 3-kinase/Akt-dependent nitric oxide generation, a process initiated by recruitment of phosphatidylinositide 3-kinase regulatory subunit p85 to TNF-receptor-1. Because amide-linked local anesthetics have well-established anti-inflammatory effects, the authors hypothesized that ropivacaine and lidocaine attenuate inflammatory Src signaling by disrupting the phosphatidylinositide 3-kinase–Akt–nitric oxide pathway, thus blocking Src-dependent neutrophil adhesion and endothelial hyperpermeability. Methods Human lung microvascular endothelial cells, incubated with TNFα in the absence or presence of clinically relevant concentrations of ropivacaine and lidocaine, were analyzed by Western blot, probing for phosphorylated/activated Src, endothelial nitric oxide synthase, Akt, intercellular adhesion molecule-1, and caveolin-1. The effect of ropivacaine on TNFα-induced nitric oxide generation, co-immunoprecipitation of TNF-receptor-1 with p85, neutrophil adhesion, and endothelial barrier disruption were assessed. Results Ropivacaine and lidocaine attenuated TNFα-induced Src activation (half-maximal inhibitory concentration [IC50] = 8.611 × 10−10 M for ropivacaine; IC50 = 5.864 × 10−10 M for lidocaine) and endothelial nitric oxide synthase phosphorylation (IC50 = 7.572 × 10−10 M for ropivacaine; IC50 = 6.377 × 10−10 M for lidocaine). Akt activation (n = 7; P = 0.006) and stimulus-dependent binding of TNF-receptor-1 and p85 (n = 6; P = 0.043) were blocked by 1 nM of ropivacaine. TNFα-induced neutrophil adhesion and disruption of endothelial monolayers via Src-dependent intercellular adhesion molecule-1- and caveolin-1-phosphorylation, respectively, were also attenuated. Conclusions Ropivacaine and lidocaine effectively blocked inflammatory TNFα signaling in endothelial cells by attenuating p85 recruitment to TNF-receptor-1. The resultant decrease in Akt, endothelial nitric oxide synthase, and Src phosphorylation reduced neutrophil adhesion and endothelial hyperpermeability. This novel anti-inflammatory “side-effect” of ropivacaine and lidocaine may provide therapeutic benefit in acute inflammatory disease. PMID:24525631

Endothelial barrier protection by local anesthetics: ropivacaine and lidocaine block tumor necrosis factor-α-induced endothelial cell Src activation.

PubMed

Piegeler, Tobias; Votta-Velis, E Gina; Bakhshi, Farnaz R; Mao, Mao; Carnegie, Graeme; Bonini, Marcelo G; Schwartz, David E; Borgeat, Alain; Beck-Schimmer, Beatrice; Minshall, Richard D

2014-06-01

Pulmonary endothelial barrier dysfunction mediated in part by Src-kinase activation plays a crucial role in acute inflammatory disease. Proinflammatory cytokines, such as tumor necrosis factor-α (TNFα), activate Src via phosphatidylinositide 3-kinase/Akt-dependent nitric oxide generation, a process initiated by recruitment of phosphatidylinositide 3-kinase regulatory subunit p85 to TNF-receptor-1. Because amide-linked local anesthetics have well-established anti-inflammatory effects, the authors hypothesized that ropivacaine and lidocaine attenuate inflammatory Src signaling by disrupting the phosphatidylinositide 3-kinase-Akt-nitric oxide pathway, thus blocking Src-dependent neutrophil adhesion and endothelial hyperpermeability. Human lung microvascular endothelial cells, incubated with TNFα in the absence or presence of clinically relevant concentrations of ropivacaine and lidocaine, were analyzed by Western blot, probing for phosphorylated/activated Src, endothelial nitric oxide synthase, Akt, intercellular adhesion molecule-1, and caveolin-1. The effect of ropivacaine on TNFα-induced nitric oxide generation, co-immunoprecipitation of TNF-receptor-1 with p85, neutrophil adhesion, and endothelial barrier disruption were assessed. Ropivacaine and lidocaine attenuated TNFα-induced Src activation (half-maximal inhibitory concentration [IC50] = 8.611 × 10 M for ropivacaine; IC50 = 5.864 × 10 M for lidocaine) and endothelial nitric oxide synthase phosphorylation (IC50 = 7.572 × 10 M for ropivacaine; IC50 = 6.377 × 10 M for lidocaine). Akt activation (n = 7; P = 0.006) and stimulus-dependent binding of TNF-receptor-1 and p85 (n = 6; P = 0.043) were blocked by 1 nM of ropivacaine. TNFα-induced neutrophil adhesion and disruption of endothelial monolayers via Src-dependent intercellular adhesion molecule-1- and caveolin-1-phosphorylation, respectively, were also attenuated. Ropivacaine and lidocaine effectively blocked inflammatory TNFα signaling in endothelial cells by attenuating p85 recruitment to TNF-receptor-1. The resultant decrease in Akt, endothelial nitric oxide synthase, and Src phosphorylation reduced neutrophil adhesion and endothelial hyperpermeability. This novel anti-inflammatory "side-effect" of ropivacaine and lidocaine may provide therapeutic benefit in acute inflammatory disease.

The prostaglandin receptor EP2 activates multiple signaling pathways and β-arrestin1 complex formation during mouse skin papilloma development

PubMed Central

Chun, Kyung-Soo; Lao, Huei-Chen; Trempus, Carol S.; Okada, Manabu; Langenbach, Robert

2009-01-01

Prostaglandin E2 (PGE2) is elevated in many tumor types, but PGE2's contributions to tumor growth are largely unknown. To investigate PGE2's roles, the contributions of one of its receptors, EP2, were studied using the mouse skin initiation/promotion model. Initial studies indicated that protein kinase A (PKA), epidermal growth factor receptor (EGFR) and several effectors—cyclic adenosine 3′,5′-monophosphate response element-binding protein (CREB), H-Ras, Src, protein kinase B (AKT) and extracellular signal-regulated kinase (ERK)1/2—were activated in 12-O-tetradecanoylphorbol-13-acetate (TPA)-promoted papillomas and that PKA and EGFR inhibition (H89 and AG1478, respectively) decreased papilloma formation. EP2's contributions to the activation of these pathways and papilloma development were determined by inhibiting endogenous TPA-induced PGE2 production with indomethacin (Indo) and concomitantly treating with the EP2 agonist, CAY10399 (CAY). CAY treatment restored papilloma formation in TPA/Indo-treated mice and increased cyclic adenosine 3′,5′-monophosphate and PKA activation as measured by p-CREB formation. CAY treatment also increased EGFR and Src activation and their inhibition by AG1478 and PP2 indicated that Src was upstream of EGFR. CAY also increased H-Ras, ERK1/2 and AKT activation, and AG1478 decreased their activation indicating EGFR being upstream. Supporting EP2's contribution, EP2−/− mice exhibited 65% fewer papillomas and reduced Src, EGFR, H-Ras, AKT and ERK1/2 activation. G protein-coupled receptor (GPCR) activation of EGFR has been reported to involve Src's activation via a GPCR–β-arrestin–Src complex. Indeed, immunoprecipitation of β-arrestin1 or p-Src indicated the presence of an EP2–β-arrestin1–p-Src complex in papillomas. The data indicated that EP2 contributed to tumor formation via activation of PKA and EGFR and that EP2 formed a complex with β-arrestin1 and Src that contributed to signaling and/or EP2 desensitization. PMID:19587094

Magnetic fields from domestic appliances in the UK

NASA Astrophysics Data System (ADS)

Preece, A. W.; Kaune, W.; Grainger, P.; Preece, S.; Golding, J.

1997-01-01

In a survey of 50 UK homes the 50 Hz fundamental and harmonic magnetic fields generated by 806 domestic appliances found in the homes, and used regularly by mothers, were measured. Measurements were made in the direction of most likely access, and from the surface of the appliances. Mothers completed a questionnaire on the use of appliances and were monitored for 24 h so that acquired exposure could be compared with the measured ambient fields in the home. Appliances were measured at standard distances and an algorithm was used to calculate fields at 100 and 50 cm to remove room background contributions. A few appliances generated fields in excess of at 1 m: microwave cookers ; washing machines ; dishwashers ; some electric showers and can openers . Of continuously operating devices, only central heating pumps (), central heating boilers () and fish-tank air pumps () produced significant fields at 0.5 m. There were no obvious ways to group different types of appliances as high- or low-strength sources. Mothers spent on average about 4.5 h per day in the kitchen, where the strongest sources of magnetic field were located.

A general ionic liquid pH-zone-refining countercurrent chromatography method for separation of alkaloids from Nelumbo nucifera Gaertn.

PubMed

Fang, Yingtong; Li, Quan; Shao, Qian; Wang, Binghai; Wei, Yun

2017-07-21

The alkaloids from lotus (Nelumbo nucifera Gaertn) are effective in lowering hyperlipemia and level of cholesterol. However, there is not a general method for their separation. In this work, a general ionic liquid pH-zone-refining countercurrent chromatography method for isolation and purification of six alkaloids from the whole lotus plant was successfully established by using ionic liquids as the modifier of the two-phase solvent system. The conditions of ionic liquid pH-zone-refining countercurrent chromatography, involving solvent systems, concentration of retainer and eluter, types of ionic liquids, the content of ionic liquids as well as ionic liquids posttreatment, were optimized to improve extraction efficiency. Finally, the separation of these six alkaloids was performed with a two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water-[C 4 mim][PF 6 ] at a volume ratio of 5:2:2:8:0.1, where 10mM TEA was added to the organic stationary phase as a retainer and 3mM HCl was added to the aqueous mobile phase as an eluter. As a result, six alkaloids including N-nornuciferine, liensinine, nuciferine, isoliensinine, roemerine and neferine were successfully separated with the purities of 97.0%, 90.2%, 94.7%, 92.8%, 90.4% and 95.9%, respectively. The established general method has been respectively applied to the crude samples of lotus leaves and lotus plumules. A total of 37.3mg of liensinine, 57.7mg of isoliensinine and 179.9mg of neferine were successfully purified in one run from 1.00g crude extract of lotus plumule with the purities of 93.2%, 96.5% and 98.8%, respectively. Amount of 45.6mg N-nornuciferine, 21.6mg nuciferine and 11.7mg roemerine was obtained in one step separation from 1.05g crude extract of lotus leaves with the purity of 96.9%, 95.6% and 91.33%, respectively. Copyright © 2017 Elsevier B.V. All rights reserved.

JunD/AP-1 Antagonizes the Induction of DAPK1 To Promote the Survival of v-Src-Transformed Cells.

PubMed

Maślikowski, Bart M; Wang, Lizhen; Wu, Ying; Fielding, Ben; Bédard, Pierre-André

2017-01-01

The increase in AP-1 activity is a hallmark of cell transformation by tyrosine kinases. Previously, we reported that blocking AP-1 using the c-Jun dominant negative mutant TAM67 induced senescence, adipogenesis, or apoptosis in v-Src-transformed chicken embryo fibroblasts (CEFs) whereas inhibition of JunD by short hairpin RNA (shRNA) specifically induced apoptosis. To investigate the role of AP-1 in Src-mediated transformation, we undertook a gene profiling study to characterize the transcriptomes of v-Src-transformed CEFs expressing either TAM67 or the JunD shRNA. Our study revealed a cluster of 18 probe sets upregulated exclusively in response to AP-1/JunD impairment and v-Src transformation. Four of these probe sets correspond to genes involved in the interferon pathway. One gene in particular, death-associated protein kinase 1 (DAPK1), is a C/EBPβ-regulated mediator of apoptosis in gamma interferon (IFN-γ)-induced cell death. Here, we show that inhibition of DAPK1 abrogates cell death in v-Src-transformed cells expressing the JunD shRNA. Chromatin immunoprecipitation data indicated that C/EBPβ was recruited to the DAPK1 promoter while the expression of a dominant negative mutant of C/EBPβ abrogated the induction of DAPK1 in response to the inhibition of AP-1. In contrast, as determined by chromatin immunoprecipitation (ChIP) assays, JunD was not detected on the DAPK1 promoter under any conditions, suggesting that JunD promotes survival by indirectly antagonizing the expression of DAPK1 in v-Src transformed cells. Transformation by the v-Src oncoprotein causes extensive changes in gene expression in primary cells such as chicken embryo fibroblasts. These changes, determining the properties of transformed cells, are controlled in part at the transcriptional level. Much attention has been devoted to transcription factors such as AP-1 and NF-κB and the control of genes associated with a more aggressive phenotype. In this report, we describe a novel mechanism of action determined by the JunD component of AP-1, a factor enhancing cell survival in v-Src-transformed cells. We show that the loss of JunD results in the aberrant activation of a genetic program leading to cell death. This program requires the activation of the tumor suppressor death-associated protein kinase 1 (DAPK1). Since DAPK1 is phosphorylated and inhibited by v-Src, these results highlight the importance of this kinase and the multiple mechanisms controlled by v-Src to antagonize the tumor suppressor function of DAPK1. Copyright © 2016 American Society for Microbiology.

Stimulus-response correspondence effect as a function of temporal overlap between relevant and irrelevant information processing.

PubMed

Wang, Dong-Yuan Debbie; Richard, F Dan; Ray, Brittany

2016-01-01

The stimulus-response correspondence (SRC) effect refers to advantages in performance when stimulus and response correspond in dimensions or features, even if the common features are irrelevant to the task. Previous research indicated that the SRC effect depends on the temporal course of stimulus information processing. The current study investigated how the temporal overlap between relevant and irrelevant stimulus processing influences the SRC effect. In this experiment, the irrelevant stimulus (a previously associated tone) preceded the relevant stimulus (a coloured rectangle). The irrelevant and relevant stimuli onset asynchrony was varied to manipulate the temporal overlap between the irrelevant and relevant stimuli processing. Results indicated that the SRC effect size varied as a quadratic function of the temporal overlap between the relevant stimulus and irrelevant stimulus. This finding extends previous experimental observations that the SRC effect size varies in an increasing or decreasing function with reaction time. The current study demonstrated a quadratic function between effect size and the temporal overlap.

An evolutionary switch in ND2 enables Src kinase regulation of NMDA receptors

NASA Astrophysics Data System (ADS)

Scanlon, David P.; Bah, Alaji; Krzeminski, Mickaël; Zhang, Wenbo; Leduc-Pessah, Heather L.; Dong, Yi Na; Forman-Kay, Julie D.; Salter, Michael W.

2017-05-01

The non-receptor tyrosine kinase Src is a key signalling hub for upregulating the function of N-methyl D-aspartate receptors (NMDARs). Src is anchored within the NMDAR complex via NADH dehydrogenase subunit 2 (ND2), a mitochondrially encoded adaptor protein. The interacting regions between Src and ND2 have been broadly identified, but the interaction between ND2 and the NMDAR has remained elusive. Here we generate a homology model of ND2 and dock it onto the NMDAR via the transmembrane domain of GluN1. This interaction is enabled by the evolutionary loss of three helices in bilaterian ND2 proteins compared to their ancestral homologues. We experimentally validate our model and demonstrate that blocking this interaction with an ND2 fragment identified in our experimental studies prevents Src-mediated upregulation of NMDAR currents in neurons. Our findings establish the mode of interaction between an NMDAR accessory protein with one of the core subunits of the receptor.

Presence of an SH2 domain in the actin-binding protein tensin.

PubMed

Davis, S; Lu, M L; Lo, S H; Lin, S; Butler, J A; Druker, B J; Roberts, T M; An, Q; Chen, L B

1991-05-03

The molecular cloning of the complementary DNA coding for a 90-kilodalton fragment of tensin, an actin-binding component of focal contacts and other submembraneous cytoskeletal structures, is reported. The derived amino acid sequence revealed the presence of a Src homology 2 (SH2) domain. This domain is shared by a number of signal transduction proteins including nonreceptor tyrosine kinases such as Abl, Fps, Src, and Src family members, the transforming protein Crk, phospholipase C-gamma 1, PI-3 (phosphatidylinositol) kinase, and guanosine triphosphatase-activating protein (GAP). Like the SH2 domain found in Src, Crk, and Abl, the SH2 domain of tensin bound specifically to a number of phosphotyrosine-containing proteins from v-src-transformed cells. Tensin was also found to be phosphorylated on tyrosine residues. These findings suggest that by possessing both actin-binding and phosphotyrosine-binding activities and being itself a target for tyrosine kinases, tensin may link signal transduction pathways with the cytoskeleton.

The tumor suppressor DAPK is reciprocally regulated by tyrosine kinase Src and phosphatase LAR.

PubMed

Wang, Won-Jing; Kuo, Jean-Cheng; Ku, Wei; Lee, Yu-Ru; Lin, Feng-Chi; Chang, Yih-Leong; Lin, Yu-Min; Chen, Chun-Hau; Huang, Yuan-Ping; Chiang, Meng-Jung; Yeh, Sheng-Wen; Wu, Pei-Rung; Shen, Che-Hung; Wu, Chen-Tu; Chen, Ruey-Hwa

2007-09-07

Death-associated protein kinase (DAPK) is a calmodulin-regulated serine/threonine kinase and elicits tumor suppression function through inhibiting cell adhesion/migration and promoting apoptosis. Despite these biological functions, the signaling mechanisms through which DAPK is regulated remain largely elusive. Here, we show that the leukocyte common antigen-related (LAR) tyrosine phosphatase dephosphorylates DAPK at pY491/492 to stimulate the catalytic, proapoptotic, and antiadhesion/antimigration activities of DAPK. Conversely, Src phosphorylates DAPK at Y491/492, which induces DAPK intra-/intermolecular interaction and inactivation. Upon EGF stimulation, a rapid Src activation leads to subsequent LAR downregulation, and these two events act in synergism to inactivate DAPK, thereby facilitating tumor cell migration and invasion toward EGF. Finally, DAPK Y491/492 hyperphosphorylation is found in human cancers in which Src activity is aberrantly elevated. These results identify LAR and Src as a DAPK regulator through their reciprocal modification of DAPK Y491/492 residues and establish a functional link of this DAPK-regulatory circuit to tumor progression.

Losartan Improves Palmitate-Induced Insulin Resistance in 3T3-L1 Adipocytes Through Upregulation of Src Phosphorylation.

PubMed

Tian, X; Ye, M; Cao, Y; Wang, C

2017-02-01

Angiotensin II type 1 receptor blocker losartan has shown strongly anti-insulin resistance properties in vivo and in vitro ; however, the underlying mechanisms are poorly understood. In this study, we demonstrate that losartan administration increased phosphorylation of Akt and its downstream Akt substrate of 160 kDa (AS160), enhanced plasma membrane translocation of glucose transporter type 4 (GLUT4), and increased glucose uptake, along with increased Src phosphorylation as well as reduced expression of docking protein 1(DOK1) in palmitate-treated 3T3-L1 adipocytes. The beneficial impacts of losartan on insulin signaling were diminished in Src-deficient 3T3-L1 adipocytes. In addition, suppressed expression of DOK1 by losartan was abolished by Src knockdown. Our results suggest that anti-insulin resistance ability of losartan is mediated by Src/DOK1/Akt pathway. © Georg Thieme Verlag KG Stuttgart · New York.

Thermodynamic Approach to Boron Nitride Nanotube Solubility and Dispersion

NASA Technical Reports Server (NTRS)

Tiano, A. L.; Gibbons, L.; Tsui, M.; Applin, S. I.; Silva, R.; Park, C.; Fay, C. C.

2016-01-01

Inadequate dispersion of nanomaterials is a critical issue that significantly limits the potential properties of nanocomposites and when overcome, will enable further enhancement of material properties. The most common methods used to improve dispersion include surface functionalization, surfactants, polymer wrapping, and sonication. Although these approaches have proven effective, they often achieve dispersion by altering the surface or structure of the nanomaterial and ultimately, their intrinsic properties. Co-solvents are commonly utilized in the polymer, paint, and art conservation industries to selectively dissolve materials. These co-solvents are utilized based on thermodynamic interaction parameters and are chosen so that the original materials are not affected. The same concept was applied to enhance the dispersion of boron nitride nanotubes (BNNTs) to facilitate the fabrication of BNNT nanocomposites. Of the solvents tested, dimethylacetamide (DMAc) exhibited the most stable, uniform dispersion of BNNTs, followed by N,N-dimethylformamide (DMF), acetone, and N-methyl-2-pyrrolidone (NMP). Utilizing the known Hansen solubility parameters of these solvents in comparison to the BNNT dispersion state, a region of good solubility was proposed. This solubility region was used to identify co-solvent systems that led to improved BNNT dispersion in poor solvents such as toluene, hexane, and ethanol. Incorporating the data from the co-solvent studies further refined the proposed solubility region. From this region, the Hansen solubility parameters for BNNTs are thought to lie at the midpoint of the solubility sphere: 16.8, 10.7, and 9.0 MPa(exp 1/2) for delta d, delta p, and delta h, respectively, with a calculated Hildebrand parameter of 21.8 MPa)exp 1/2).

Thermodynamic approach to boron nitride nanotube solubility and dispersion.

PubMed

Tiano, A L; Gibbons, L; Tsui, M; Applin, S I; Silva, R; Park, C; Fay, C C

2016-02-21

Inadequate dispersion of nanomaterials is a critical issue that significantly limits the potential properties of nanocomposites and when overcome, will enable further enhancement of material properties. The most common methods used to improve dispersion include surface functionalization, surfactants, polymer wrapping, and sonication. Although these approaches have proven effective, they often achieve dispersion by altering the surface or structure of the nanomaterial and ultimately, their intrinsic properties. Co-solvents are commonly utilized in the polymer, paint, and art conservation industries to selectively dissolve materials. These co-solvents are utilized based on thermodynamic interaction parameters and are chosen so that the original materials are not affected. The same concept was applied to enhance the dispersion of boron nitride nanotubes (BNNTs) to facilitate the fabrication of BNNT nanocomposites. Of the solvents tested, dimethylacetamide (DMAc) exhibited the most stable, uniform dispersion of BNNTs, followed by N,N-dimethylformamide (DMF), acetone, and N-methyl-2-pyrrolidone (NMP). Utilizing the known Hansen solubility parameters of these solvents in comparison to the BNNT dispersion state, a region of good solubility was proposed. This solubility region was used to identify co-solvent systems that led to improved BNNT dispersion in poor solvents such as toluene, hexane, and ethanol. Incorporating the data from the co-solvent studies further refined the proposed solubility region. From this region, the Hansen solubility parameters for BNNTs are thought to lie at the midpoint of the solubility sphere: 16.8, 10.7, and 9.0 MPa(1/2) for δd, δp, and δh, respectively, with a calculated Hildebrand parameter of 21.8 MPa(1/2).

Activation of Stat3 Transcription Factor by Herpesvirus Saimiri STP-A Oncoprotein

PubMed Central

Chung, Young-Hwa; Cho, Nam-hyuk; Garcia, Maria Ines; Lee, Sun-Hwa; Feng, Pinghui; Jung, Jae U.

2004-01-01

The saimiri transforming protein (STP) oncogene of Herpesvirus saimiri subgroup A strain 11 (STP-A11) is not required for viral replication but is required for lymphoid cell immortalization in culture and lymphoma induction in primates. We previously showed that STP-A11 interacts with cellular Src kinase through its SH2 binding motif and that this interaction elicits Src signal transduction. Here we demonstrate that STP-A11 interacts with signal transducer and activator of transcription 3 (Stat3) independently of Src association and that the amino-terminal short proline-rich motif of STP-A11 and the central linker region of Stat3 are necessary for their interaction. STP-A11 formed a triple complex with Src kinase and Stat3 where Src kinase phosphorylated Stat3, resulting in the nuclear localization and transcriptional activation of Stat3. Consequently, the constitutively active Stat3 induced by STP-A11 elicited cellular signal transduction, which ultimately induced cell survival and proliferation upon serum deprivation. Furthermore, this activity was strongly correlated with the induction of Fos, cyclin D1, and Bcl-XL expression. These results demonstrate that STP-A11 independently targets two important cellular signaling molecules, Src and Stat3, and that these proteins cooperate efficiently to induce STP-A11-mediated transformation. PMID:15163742

Effects of sustained-release composite on the oxygen levels and sediment phosphorus fractions of an urban river in Shanghai.

PubMed

Li, Yang; Zhou, Yanbo; Zhou, Zhenhua; Wang, Ningsheng; Zhou, Qiang; Wang, Fuchen

2014-01-01

The eutrophication of many rivers and lakes is attributed to the anoxia and the increasing internal loading of nutrients from sediment. A novel sustained-release composite (SRC) synthesis of stearic acid and calcium peroxide (CaO2) was applied to supply a water body with oxygen endured in this study. The influences of SRC on the dissolved oxygen (DO) level, pH and total phosphorus (TP) of an urban river in Shanghai were studied. The results show that SRC has a longer oxygen-releasing cycle and a more tender effect on pH with the comparison of CaO2 powder. Reduction of 79.6% in the concentration of TP was observed in the water column. After 35 days of SRC addition, there was a significant positive correlation between TP and DO. As a consequence, the phosphorus fractions in sediment, including loosely sorbed P (NH4Cl-P), redox-sensitive P (Fe-P), calcium bound P (Ca-P), aluminium bound P (Al-P) and residual P (organic and refractory P) were affected by the addition of SRC. The NH4Cl-P and Fe-P fractions in the sediment that could release P easily were well constrained under the positive effect of SRC.

Selective Targeting of SH2 Domain–Phosphotyrosine Interactions of Src Family Tyrosine Kinases with Monobodies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kükenshöner, Tim; Schmit, Nadine Eliane; Bouda, Emilie

The binding of Src-homology 2 (SH2) domains to phosphotyrosine (pY) sites is critical for the autoinhibition and substrate recognition of the eight Src family kinases (SFKs). The high sequence conservation of the 120 human SH2 domains poses a significant challenge to selectively perturb the interactions of even the SFK SH2 family against the rest of the SH2 domains. We have developed synthetic binding proteins, termed monobodies, for six of the SFK SH2 domains with nanomolar affinity. Most of these monobodies competed with pY ligand binding and showed strong selectivity for either the SrcA (Yes, Src, Fyn, Fgr) or SrcB subgroupmore » (Lck, Lyn, Blk, Hck). Interactome analysis of intracellularly expressed monobodies revealed that they bind SFKs but no other SH2-containing proteins. Three crystal structures of monobody–SH2 complexes unveiled different and only partly overlapping binding modes, which rationalized the observed selectivity and enabled structure-based mutagenesis to modulate inhibition mode and selectivity. In line with the critical roles of SFK SH2 domains in kinase autoinhibition and T-cell receptor signaling, monobodies binding the Src and Hck SH2 domains selectively activated respective recombinant kinases, whereas an Lck SH2-binding monobody inhibited proximal signaling events downstream of the T-cell receptor complex. Our results show that SFK SH2 domains can be targeted with unprecedented potency and selectivity using monobodies. They are excellent tools for dissecting SFK functions in normal development and signaling and to interfere with aberrant SFK signaling networks in cancer cells.« less

SRC: marker or actor in prostate cancer aggressiveness.

PubMed

Vlaeminck-Guillem, Virginie; Gillet, Germain; Rimokh, Ruth

2014-01-01

A key question for urologic practitioners is whether an apparently organ-confined prostate cancer (PCa) is actually aggressive or not. The dilemma is to specifically identify among all prostate tumors the very aggressive high-grade cancers that will become life-threatening by developing extra-prostatic invasion and metastatic potential and the indolent cancers that will never modify a patient's life expectancy. A choice must be made between several therapeutic options to achieve the optimal personalized management of the disease that causes as little harm as possible to patients. Reliable clinical, biological, or pathological markers that would enable distinctions to be made between aggressive and indolent PCas in routine practice at the time of initial diagnosis are still lacking. The molecular mechanisms that explain why a PCa is aggressive or not are also poorly understood. Among the potential markers and/or actors in PCa aggressiveness, Src and other members of the Src kinase family, are valuable candidates. Activation of Src-dependent intracellular pathways is frequently observed in PCa. Indeed, Src is at the cross-roads of several pathways [including androgen receptor (AR), TGFbeta, Bcl-2, Akt/PTEN or MAPK, and ERK …], and is now known to influence some of the cellular and tissular events that accompany tumor progression: cell proliferation, cell motility, invasion, epithelial-to-mesenchymal transition, resistance to apoptosis, angiogenesis, neuroendocrine differentiation, and metastatic spread. Recent work even suggests that Src could also play a part in PCa initiation in coordination with the AR. The aim of this review is to gather data that explore the links between the Src kinase family and PCa progression and aggressiveness.

Removal From Play After Concussion and Recovery Time

PubMed Central

Sufrinko, Alicia; Schatz, Philip; French, Jon; Henry, Luke; Burkhart, Scott; Collins, Michael W.; Kontos, Anthony P.

2016-01-01

OBJECTIVE: Despite increases in education and awareness, many athletes continue to play with signs and symptoms of a sport-related concussion (SRC). The impact that continuing to play has on recovery is unknown. This study compared recovery time and related outcomes between athletes who were immediately removed from play and athletes who continued to play with an SRC. METHODS: A prospective, repeated measures design was used to compare neurocognitive performance, symptoms, and recovery time between 35 athletes (mean ± SD age, 15.61 ± 1.65 years) immediately removed after an SRC (REMOVED group) compared with 34 athletes (mean ± SD age, 15.35 ± 1.73 years) who continued to play (PLAYED group) with SRC. Neurocognitive and symptom data were obtained at baseline and at 1 to 7 days and 8 to 30 days after an SRC. RESULTS: The PLAYED group took longer to recover than the REMOVED group (44.4 ± 36.0 vs 22.0 ± 18.7 days; P = .003) and were 8.80 times more likely to demonstrate protracted recovery (≥21 days) (P < .001). Removal from play status was associated with the greatest risk of protracted recovery (adjusted odds ratio, 14.27; P = .001) compared with other predictors (eg, sex). The PLAYED group exhibited significantly worse neurocognitive and greater symptoms than the REMOVED group. CONCLUSIONS: SRC recovery time may be reduced if athletes are removed from participation. Immediate removal from play is the first step in mitigating prolonged SRC recovery, and these data support current consensus statements and management guidelines. PMID:27573089

The role of steroid receptor coactivator-3 (SRC-3) in human malignant disease.

PubMed

Gojis, O; Rudraraju, B; Alifrangis, C; Krell, J; Libalova, P; Palmieri, C

2010-03-01

The p160 steroid receptor coactivator (SRC) family is critical to the transcriptional activation function of nuclear hormone receptors. A key member of this family is SRC-3, initially found to be amplified and expressed in breast cancer it has subsequent been shown to be expressed in malignant disease arising from a wide range of other organs. An understanding of the potential role of SRC-3 in the pathogenesis and its possible prognostic role in a broad range of tumours will improve our general understanding of carcinogenesis as well as potentially leading to a new prognostic marker as well as new therapeutic targets. Relevant papers were identified by searching the PubMed and MEDLINE databases for article published until 28th February 2009. Only articles published in English were considered. The search terms included "SRC-3", "AIB1" in association with the following terms: "human", "cancer" and "malignant disease". The search focused on malignant disease arising outside of the mammary gland. Full articles were obtained and references were checked for additional material when appropriate. SRC-3 is amplified and expressed in a wide spectrum of human malignant diseases and appears to be a potential prognostic marker in a number of different tumours. SRC-3 appears to be implicated in the possible risk of developing prostate and ovarian cancer. Its presence appears to be a marker of aggressive disease. Further research is required to determine its predictive and prognostic utility given the relative paucity of studies for each specific malignant disease. Copyright (c) 2009. Published by Elsevier Ltd.

Thrombin increases hyposmotic taurine efflux and accelerates ICI-swell and RVD in 3T3 fibroblasts by a src-dependent EGFR transactivation.

PubMed

Vázquez-Juárez, E; Ramos-Mandujano, G; Lezama, R A; Cruz-Rangel, S; Islas, L D; Pasantes-Morales, H

2008-02-01

The present study in Swiss3T3 fibroblasts examines the effect of thrombin on hyposmolarity-induced osmolyte fluxes and RVD, and the contribution of the src/EGFR pathway. Thrombin (5 U/ml) added to a 30% hyposmotic medium markedly increased hyposmotic 3H-taurine efflux (285%), accelerated the volume-sensitive Cl- current (ICI-swell) and increased RVD rate. These effects were reduced (50-65%) by preventing the thrombin-induced intracellular Ca2+ [Ca2+]i rise with EGTA-AM, or with the phospholipase C (PLC) blocker U73122. Ca2+calmodulin (CaM) and calmodulin kinase II (CaMKII) also participate in this Ca2+-dependent pathway. Thrombin plus hyposmolarity increased src and EGFR phosphorylation, whose blockade by PP2 and AG1478, decreased by 30-50%, respectively, the thrombin effects on hyposmotic taurine efflux, ICI-swell and RVD. Ca2+- and src/EGFR-mediated pathways operate independently as shown by (1) the persistence of src and EGFR activation when [Ca2+]i rise is prevented and (2) the additive effect on taurine efflux, ICI-swell or RVD by simultaneous inhibition of the two pathways, which essentially suppressed these events. PLC-Ca2+- and src/EGFR-signaling pathways operate in the hyposmotic condition and because thrombin per se failed to increase taurine efflux and ICI-swell under isosmotic condition it seems that it is merely amplifying these previously activated mechanisms. The study shows that thrombin potentiates hyposmolarity-induced osmolyte fluxes and RVD by increasing src/EGFR-dependent signaling, in addition to the Ca2+-dependent pathway.

The Transmodulation of HER2 and EGFR by Substance P in Breast Cancer Cells Requires c-Src and Metalloproteinase Activation.

PubMed

Garcia-Recio, Susana; Pastor-Arroyo, Eva M; Marín-Aguilera, Mercedes; Almendro, Vanessa; Gascón, Pedro

2015-01-01

Substance P (SP) is a pleiotropic cytokine/neuropeptide that enhances breast cancer (BC) aggressiveness by transactivating tyrosine kinase receptors like EGFR and HER2. We previously showed that SP and its cognate receptor NK-1 (SP/NK1-R) signaling modulates the basal phosphorylation of HER2 and EGFR in BC, increasing aggressiveness and drug resistance. In order to elucidate the mechanisms responsible for NK-1R-mediated HER2 and EGFR transactivation, we investigated the involvement of c-Src (a ligand-independent mediator) and of metalloproteinases (ligand-dependent mediators) in HER2/EGFR activation. Overexpression of NK-1R in MDA-MB-231 and its chemical inhibition in SK-BR-3, BT-474 and MDA-MB-468 BC cells significantly modulated c-Src activation, suggesting that this protein is a mediator of NK-1R signaling. In addition, the c-Src inhibitor 4-(4'-phenoxyanilino)-6,7-dimethoxyquinazoline prevented SP-induced activation of HER2. On the other hand, SP-dependent phosphorylation of HER2 and EGFR decreased substantially in the presence of the MMP inhibitor 1-10, phenanthroline monohydrate, and the dual inhibition of both c-Src and MMP almost abolished the activation of HER2 and EGFR. Moreover, the use of these inhibitors demonstrated that this Src and MMP-dependent signaling is important to the cell viability and migration capacity of HER2+ and EGFR+ cell lines. Our results indicate that the transactivation of HER2 and EGFR by the pro-inflammatory cytokine/neuropeptide SP in BC cells is a c-Src and MMP-dependent process.

Antitumor Effect of KX-01 through Inhibiting Src Family Kinases and Mitosis.

PubMed

Kim, Seongyeong; Min, Ahrum; Lee, Kyung-Hun; Yang, Yaewon; Kim, Tae-Yong; Lim, Jee Min; Park, So Jung; Nam, Hyun-Jin; Kim, Jung Eun; Song, Sang-Hyun; Han, Sae-Won; Oh, Do-Youn; Kim, Jee Hyun; Kim, Tae-You; Hangauer, David; Lau, Johnson Yiu-Nam; Im, Kyongok; Lee, Dong Soon; Bang, Yung-Jue; Im, Seock-Ah

2017-07-01

KX-01 is a novel dual inhibitor of Src and tubulin. Unlike previous Src inhibitors that failed to show clinical benefit during treatment of breast cancer, KX-01 can potentially overcome the therapeutic limitations of current Src inhibitors through inhibition of both Src and tubulin. The present study further evaluates the activity and mechanism of KX-01 in vitro and in vivo . The antitumor effect of KX-01 in triple negative breast cancer (TNBC) cell lines was determined by MTT assay. Wound healing and immunofluorescence assays were performed to evaluate the action mechanisms of KX-01. Changes in the cell cycle and molecular changes induced by KX-01 were also evaluated. A MDA-MB-231 mouse xenograft model was used to demonstrate the in vivo effects. KX-01 effectively inhibited the growth of breast cancer cell lines. The expression of phospho-Src and proliferative-signaling molecules were down-regulated in KX-01-sensitive TNBC cell lines. In addition, migration inhibition was observed by wound healing assay. KX-01-induced G2/M cell cycle arrest and increased the aneuploid cell population in KX-01-sensitive cell lines. Multi-nucleated cells were significantly increased after KX-01 treatment. Furthermore, KX-01 effectively delayed tumor growth in a MDA-MB-231 mouse xenograft model. KX-01 effectively inhibited cell growth and migration of TNBC cells. Moreover, this study demonstrated that KX-01 showed antitumor effects through the inhibition of Src signaling and the induction of mitotic catastrophe. The antitumor effects of KX-01 were also demonstrated in vivo using a mouse xenograft model.

Src family kinase expression and subcellular localization in macrophages: implications for their role in CSF-1-induced macrophage migration.

PubMed

Dwyer, Amy R; Mouchemore, Kellie A; Steer, James H; Sunderland, Andrew J; Sampaio, Natalia G; Greenland, Eloise L; Joyce, David A; Pixley, Fiona J

2016-07-01

A major role of colony-stimulating factor-1 is to stimulate the differentiation of mononuclear phagocytic lineage cells into adherent, motile, mature macrophages. The colony-stimulating factor-1 receptor transduces colony-stimulating factor-1 signaling, and we have shown previously that phosphatidylinositol 3-kinase p110δ is a critical mediator of colony-stimulating factor-1-stimulated motility through the colony-stimulating factor-1 receptor pY721 motif. Src family kinases are also implicated in the regulation of macrophage motility and in colony-stimulating factor-1 receptor signaling, although functional redundancy of the multiple SFKs expressed in macrophages makes it challenging to delineate their specific functions. We report a comprehensive analysis of individual Src family kinase expression in macrophage cell lines and primary macrophages and demonstrate colony-stimulating factor-1-induced changes in Src family kinase subcellular localization, which provides clues to their distinct and redundant functions in macrophages. Moreover, expression of individual Src family kinases is both species specific and dependent on colony-stimulating factor-1-induced macrophage differentiation. Hck associated with the activated colony-stimulating factor-1 receptor, whereas Lyn associated with the receptor in a constitutive manner. Consistent with this, inhibitor studies revealed that Src family kinases were important for both colony-stimulating factor-1 receptor activation and colony-stimulating factor-1-induced macrophage spreading, motility, and invasion. Distinct colony-stimulating factor-1-induced changes in the subcellular localization of individual SFKs suggest specific roles for these Src family kinases in the macrophage response to colony-stimulating factor-1. © Society for Leukocyte Biology.

Valsartan regulates the interaction of angiotensin II type 1 receptor and endothelial nitric oxide synthase via Src/PI3K/Akt signalling.

PubMed

Su, Kuo-Hui; Tsai, Jin-Yi; Kou, Yu Ru; Chiang, An-Na; Hsiao, Sheng-Huang; Wu, Yuh-Lin; Hou, Hsin-Han; Pan, Ching-Chian; Shyue, Song-Kun; Lee, Tzong-Shyuan

2009-06-01

Valsartan, a selective angiotensin II type 1 receptor (AT1R) blocker, has beneficial effects in the cardiovascular system in part by its increase of nitric oxide (NO) bioavailability, yet the mechanisms are unclear. We investigated the molecular mechanisms underlying this effect in endothelial cells (ECs). NO production was examined by Griess reagent assay, DAF-2 DA fluorescence staining and cGMP ELISA kits. Protein interaction was determined by western blotting and immunoprecipitation. Treating bovine or human aortic ECs with valsartan increased NO production, as evidenced by elevated level of stable NO metabolites and intracellular cGMP. Valsartan increased the phosphorylation but not the protein level of endothelial NO synthase (eNOS). Inhibition of phosphoinositide-3 kinase (PI3K)/Akt and Src pathways by specific inhibitors suppressed valsartan-induced NO release. In addition, valsartan increased the tyrosine residue phosphorylation of AT1R, which was attenuated by inhibition of Src but not PI3K activities. Valsartan also suppressed the interaction of eNOS and AT1R, which was blocked by Src or PI3K inhibition. Valsartan-induced NO production in ECs is mediated through Src/PI3K/Akt-dependent phosphorylation of eNOS. Valsartan-induced AT1R phosphorylation depends on Src but not PI3K, whereas valsartan-induced suppression of AT1R-eNOS interaction depends on Src/PI3K/Akt signalling. These results indicate a novel vasoprotective mechanism of valsartan in upregulating NO production in ECs.

Steroid Receptor Coactivator-1 Knockdown Decreases Synaptic Plasticity and Impairs Spatial Memory in the Hippocampus of Mice.

PubMed

Bian, Chen; Huang, Yan; Zhu, Haitao; Zhao, Yangang; Zhao, Jikai; Zhang, Jiqiang

2018-05-01

Steroids have been demonstrated to play profound roles in the regulation of hippocampal function by acting on their receptors, which need coactivators for their transcriptional activities. Previous studies have shown that steroid receptor coactivator-1 (SRC-1) is the predominant coactivator in the hippocampus, but its exact role and the underlying mechanisms remain unclear. In this study, we constructed SRC-1 RNA interference (RNAi) lentiviruses, injected them into the hippocampus of male mice, and then examined the changes in the expression of selected synaptic proteins, CA1 synapse density, postsynaptic density (PSD) thickness, and in vivo long-term potentiation (LTP). Spatial learning and memory behavior changes were investigated using the Morris water maze. We then transfected the lentiviruses into cultured hippocampal cells and examined the changes in synaptic protein and phospho-cyclic AMP response element-binding protein (pCREB) expression. The in vivo results showed that SRC-1 knockdown significantly decreased the expression of synaptic proteins and CA1 synapse density as well as PSD thickness; SRC-1 knockdown also significantly impaired in vivo LTP and disrupted spatial learning and memory. The in vitro results showed that while the expression of synaptic proteins was significantly decreased by SRC-1 knockdown, pCREB expression was also significantly decreased. The above results suggest a pivotal role of SRC-1 in the regulation of hippocampal synaptic plasticity and spatial learning and memory, strongly indicating SRC-1 may serve as a novel therapeutic target for hippocampus-dependent memory disorders. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

Src-dependent phosphorylation of caveolin-1 Tyr-14 promotes swelling and release of caveolae

PubMed Central

Zimnicka, Adriana M.; Husain, Yawer S.; Shajahan, Ayesha N.; Sverdlov, Maria; Chaga, Oleg; Chen, Zhenlong; Toth, Peter T.; Klomp, Jennifer; Karginov, Andrei V.; Tiruppathi, Chinnaswamy; Malik, Asrar B.; Minshall, Richard D.

2016-01-01

Caveolin 1 (Cav1) is a required structural component of caveolae, and its phosphorylation by Src is associated with an increase in caveolae-mediated endocytosis. Here we demonstrate, using quantitative live-cell 4D, TIRF, and FRET imaging, that endocytosis and trafficking of caveolae are associated with a Cav1 Tyr-14 phosphorylation-dependent conformational change, which spatially separates, or loosens, Cav1 molecules within the oligomeric caveolar coat. When tracked by TIRF and spinning-disk microscopy, cells expressing phosphomimicking Cav1 (Y14D) mutant formed vesicles that were greater in number and volume than with Y14F-Cav1-GFP. Furthermore, we observed in HEK cells cotransfected with wild-type, Y14D, or Y14F Cav1-CFP and -YFP constructs that FRET efficiency was greater with Y14F pairs than with Y14D, indicating that pY14-Cav1 regulates the spatial organization of Cav1 molecules within the oligomer. In addition, albumin-induced Src activation or direct activation of Src using a rapamycin-inducible Src construct (RapR-Src) led to an increase in monomeric Cav1 in Western blots, as well as a simultaneous increase in vesicle number and decrease in FRET intensity, indicative of a Src-mediated conformational change in CFP/YFP-tagged WT-Cav1 pairs. We conclude that phosphorylation of Cav1 leads to separation or “spreading” of neighboring negatively charged N-terminal phosphotyrosine residues, promoting swelling of caveolae, followed by their release from the plasma membrane. PMID:27170175

Multidisciplinary Management of Pediatric Sports-Related Concussion.

PubMed

Ellis, Michael J; Ritchie, Lesley J; McDonald, Patrick J; Cordingley, Dean; Reimer, Karen; Nijjar, Satnam; Koltek, Mark; Hosain, Shahid; Johnston, Janine; Mansouri, Behzad; Sawyer, Scott; Silver, Norm; Girardin, Richard; Larkins, Shannon; Vis, Sara; Selci, Erin; Davidson, Michael; Gregoire, Scott; Sam, Angela; Black, Brian; Bunge, Martin; Essig, Marco; MacDonald, Peter; Leiter, Jeff; Russell, Kelly

2017-01-01

To summarize the clinical characteristics and outcomes of pediatric sports-related concussion (SRC) patients who were evaluated and managed at a multidisciplinary pediatric concussion program and examine the healthcare resources and personnel required to meet the needs of this patient population. We conducted a retrospective review of all pediatric SRC patients referred to the Pan Am Concussion Program from September 1st, 2013 to May 25th, 2015. Initial assessments and diagnoses were carried out by a single neurosurgeon. Return-to-Play decision-making was carried out by the multidisciplinary team. 604 patients, including 423 pediatric SRC patients were evaluated at the Pan Am Concussion Program during the study period. The mean age of study patients was 14.30 years (SD: 2.32, range 7-19 years); 252 (59.57%) were males. Hockey (182; 43.03%) and soccer (60; 14.18%) were the most commonly played sports at the time of injury. Overall, 294 (69.50%) of SRC patients met the clinical criteria for concussion recovery, while 75 (17.73%) were lost to follow-up, and 53 (12.53%) remained in active treatment at the end of the study period. The median duration of symptoms among the 261 acute SRC patients with complete follow-up was 23 days (IQR: 15, 36). Overall, 25.30% of pediatric SRC patients underwent at least one diagnostic imaging test and 32.62% received referral to another member of our multidisciplinary clinical team. Comprehensive care of pediatric SRC patients requires access to appropriate diagnostic resources and the multidisciplinary collaboration of experts with national and provincially-recognized training in TBI.

Analysis of macromolecules, ligands and macromolecule-ligand complexes

DOEpatents

Von Dreele, Robert B [Los Alamos, NM

2008-12-23

A method for determining atomic level structures of macromolecule-ligand complexes through high-resolution powder diffraction analysis and a method for providing suitable microcrystalline powder for diffraction analysis are provided. In one embodiment, powder diffraction data is collected from samples of polycrystalline macromolecule and macromolecule-ligand complex and the refined structure of the macromolecule is used as an approximate model for a combined Rietveld and stereochemical restraint refinement of the macromolecule-ligand complex. A difference Fourier map is calculated and the ligand position and points of interaction between the atoms of the macromolecule and the atoms of the ligand can be deduced and visualized. A suitable polycrystalline sample of macromolecule-ligand complex can be produced by physically agitating a mixture of lyophilized macromolecule, ligand and a solvent.

Potential for control of detrusor smooth muscle spontaneous rhythmic contraction by cyclooxygenase products released by interstitial cells of Cajal

PubMed Central

Collins, Clinton; Klausner, Adam P; Herrick, Benjamin; Koo, Harry P; Miner, Amy S; Henderson, Scott C; Ratz, Paul H

2009-01-01

Interstitial cells of Cajal (ICCs) have been identified as pacemaker cells in the upper urinary tract and urethra, but the role of ICCs in the bladder remains to be determined. We tested the hypotheses that ICCs express cyclooxygenase (COX), and that COX products (prostaglandins), are the cause of spontaneous rhythmic contraction (SRC) of isolated strips of rabbit bladder free of urothelium. SRC was abolished by 10 μM indomethacin and ibuprofen (non-selective COX inhibitors). SRC was concentration-dependently inhibited by selective COX-1 (SC-560 and FR-122047) and COX-2 inhibitors (NS-398 and LM-1685), and by SC-51089, a selective antagonist for the PGE-2 receptor (EP) and ICI-192,605 and SQ-29,548, selective antagonists for thromboxane receptors (TP). The partial agonist/antagonist of the PGF-2α receptor (FP), AL-8810, inhibited SRC by ∼50%. Maximum inhibition was ∼90% by SC-51089, ∼80–85% by the COX inhibitors and ∼70% by TP receptor antagonists. In the presence of ibuprofen to abolish SRC, PGE-2, sulprostone, misoprostol, PGF-2α and U-46619 (thromboxane mimetic) caused rhythmic contractions that mimicked SRC. Fluorescence immunohistochemistry coupled with confocal laser scanning microscopy revealed that c-Kit and vimentin co-localized to interstitial cells surrounding detrusor smooth muscle bundles, indicating the presence of extensive ICCs in rabbit bladder. Co-localization of COX-1 and vimentin, and COX-2 and vimentin by ICCs supports the hypothesis that ICCs were the predominant cell type in rabbit bladder expressing both COX isoforms. These data together suggest that ICCs appear to be an important source of prostaglandins that likely play a role in regulation of SRC. Additional studies on prostaglandin-dependent SRC may generate opportunities for the application of novel treatments for disorders leading to overactive bladder. PMID:19243470

Lipopolysaccharide induces VCAM-1 expression and neutrophil adhesion to human tracheal smooth muscle cells: Involvement of Src/EGFR/PI3-K/Akt pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, W.-N.; Luo, S.-F.; Wu, C.-B.

2008-04-15

In our previous study, LPS has been shown to induce vascular cell adhesion molecule-1(VCAM-1) expression through MAPKs and NF-{kappa}B in human tracheal smooth muscle cells (HTSMCs). In addition to these pathways, the non-receptor tyrosine kinases (Src), EGF receptor (EGFR), and phosphatidylinositol 3-kinase (PI3K) have been shown to be implicated in the expression of several inflammatory target proteins. Here, we reported that LPS-induced up-regulation of VCAM-1 enhanced the adhesion of neutrophils onto HTSMC monolayer, which was inhibited by LY294002 and wortmannin. LPS stimulated phosphorylation of protein tyrosine kinases including Src, PYK2, and EGFR, which were further confirmed using specific anti-phospho-Src, PYK2,more » or EGFR Ab, respectively, revealed by Western blotting. LPS-stimulated Src, PYK2, EGFR, and Akt phosphorylation and VCAM-1 expression were attenuated by the inhibitors of Src (PP1), EGFR (AG1478), PI3-K (LY294002 and wortmannin), and Akt (SH-5), respectively, or transfection with siRNAs of Src or Akt and shRNA of p110. LPS-induced VCAM-1 expression was also blocked by pretreatment with curcumin (a p300 inhibitor) or transfection with p300 siRNA. LPS-stimulated Akt activation translocated into nucleus and associated with p300 and VCAM-1 promoter region was further confirmed by immunofluorescence, immunoprecipitation, and chromatin immunoprecipitation assays. This association of Akt and p300 to VCAM-1 promoter was inhibited by pretreatment with PP1, AG1478, wortmannin, and SH-5. LPS-induced p300 activation enhanced VCAM-1 promoter activity and VCAM-1 mRNA expression. These results suggested that in HTSMCs, Akt phosphorylation mediated through transactivation of Src/PYK2/EGFR promoted the transcriptional p300 activity and eventually led to VCAM-1 expression induced by LPS.« less

Na/K-ATPase/src complex mediates regulation of CD40 in renal parenchyma.

PubMed

Xie, Jeffrey X; Zhang, Shungang; Cui, Xiaoyu; Zhang, Jue; Yu, Hui; Khalaf, Fatimah K; Malhotra, Deepak; Kennedy, David J; Shapiro, Joseph I; Tian, Jiang; Haller, Steven T

2017-12-22

Recent studies have highlighted a critical role for CD40 in the pathogenesis of renal injury and fibrosis. However, little is currently understood about the regulation of CD40 in this setting. We use novel Na/K-ATPase cell lines and inhibitors in order to demonstrate the regulatory function of Na/K-ATPase with regards to CD40 expression and function. We utilize 5/6 partial nephrectomy as well as direct infusion of a Na/K-ATPase ligand to demonstrate this mechanism exists in vivo. We demonstrate that knockdown of the α1 isoform of Na/K-ATPase causes a reduction in CD40 while rescue of the α1 but not the α2 isoform restores CD40 expression in renal epithelial cells. Second, because the major functional difference between α1 and α2 is the ability of α1 to form a functional signaling complex with Src, we examined whether the Na/K-ATPase/Src complex is important for CD40 expression. We show that a gain-of-Src binding α2 mutant restores CD40 expression while loss-of-Src binding α1 reduces CD40 expression. Furthermore, loss of a functional Na/K-ATPase/Src complex also disrupts CD40 signaling. Importantly, we show that use of a specific Na/K-ATPase/Src complex antagonist, pNaKtide, can attenuate cardiotonic steroid (CTS)-induced induction of CD40 expression in vitro. Because the Na/K-ATPase/Src complex is also a key player in the pathogenesis of renal injury and fibrosis, our new findings suggest that Na/K-ATPase and CD40 may comprise a pro-fibrotic feed-forward loop in the kidney and that pharmacological inhibition of this loop may be useful in the treatment of renal fibrosis. © The Author 2017. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

HSP27 phosphorylation modulates TRAIL-induced activation of Src-Akt/ERK signaling through interaction with β-arrestin2.

PubMed

Qi, Shimei; Xin, Yinqiang; Qi, Zhilin; Xu, Yimiao; Diao, Ying; Lan, Lei; Luo, Lan; Yin, Zhimin

2014-03-01

Heat shock protein 27 (HSP27) regulates critical cellular functions such as development, differentiation, cell growth and apoptosis. A variety of stimuli induce the phosphorylation of HSP27, which affects its cellular functions. However, most previous studies focused on the role of HSP27 protein itself in apoptosis, the particular role of its phosphorylation state in signaling transduction remains largely unclear. In the present study, we reported that HSP27 phosphorylation modulated TRAIL-triggered pro-survival signaling transduction. In HeLa cells, suppression of HSP27 phosphorylation by specific inhibitor KRIBB3 or MAPKAPK2 (MK2) knockdown and by overexpression of non-phosphorylatable HSP27(3A) mutant demonstrated that hindered HSP27 phosphorylation enhanced the TRAIL-induced apoptosis. In addition, reduced HSP27 phosphorylation by KRIBB3 treatment or MK2 knockdown attenuated the TRAIL-induced activation of Akt and ERK survival signaling through suppressing the phosphorylation of Src. By overexpression of HSP27(15A) or HSP27(78/82A) phosphorylation mutant, we further showed that phosphorylation of HSP27 at serine 78/82 residues was essential to TRAIL-triggered Src-Akt/ERK signaling transduction. Co-immunoprecipitation and confocal microscopy showed that HSP27 interacted with Src and scaffolding protein β-arrestin2 in response of TRAIL stimulation and suppression of HSP27 phosphorylation apparently disrupted the TRAIL-induced interaction of HSP27 and Src or interaction of HSP27 and β-arrestin2. We further demonstrated that β-arrestin2 mediated HSP27 action on TRAIL-induced Src activation, which was achieved by recruiting signaling complex of HSP27/β-arrestin2/Src in response to TRAIL. Taken together, our study revealed that HSP27 phosphorylation modulates TRAIL-triggered activation of Src-Akt/ERK pro-survival signaling via interacting with β-arrestin2 in HeLa cells. Copyright © 2013 Elsevier Inc. All rights reserved.

Isthmin is a novel vascular permeability inducer that functions through cell-surface GRP78-mediated Src activation.

PubMed

Venugopal, Shruthi; Chen, Mo; Liao, Wupeng; Er, Shi Yin; Wong, Wai-Shiu Fred; Ge, Ruowen

2015-07-01

Isthmin (ISM) is a recently identified 60 kDa secreted angiogenesis inhibitor. Two cell-surface receptors for ISM have been defined, the high-affinity glucose-regulated protein 78 kDa (GRP78) and the low-affinity αvβ5 integrin. As αvβ5 integrin plays an important role in pulmonary vascular permeability (VP) and ISM is highly expressed in mouse lung, we sought to clarify the role of ISM in VP. Recombinant ISM (rISM) dose-dependently enhances endothelial monolayer permeability in vitro and local dermal VP when administered intradermally in mice. Systemic rISM administration through intravenous injection leads to profound lung vascular hyperpermeability but not in other organs. Mechanistic investigations using molecular, biochemical approaches and specific chemical inhibitors revealed that ISM-GRP78 interaction triggers a direct interaction between GRP78 and Src, leading to Src activation and subsequent phosphorylation of adherens junction proteins and loss of junctional proteins from inter-endothelial junctions, resulting in enhanced VP. Dynamic studies of Src activation, VP and apoptosis revealed that ISM induces VP directly via Src activation while apoptosis contributes indirectly only after prolonged treatment. Furthermore, ISM is significantly up-regulated in lipopolysaccharide (LPS)-treated mouse lung. Blocking cell-surface GRP78 by systemic infusion of anti-GRP78 antibody significantly attenuates pulmonary vascular hyperpermeability in LPS-induced acute lung injury (ALI) in mice. ISM is a novel VP inducer that functions through cell-surface GRP78-mediated Src activation as well as induction of apoptosis. It induces a direct GRP78-Src interaction, leading to cytoplasmic Src activation. ISM contributes to pulmonary vascular hyperpermeability of LPS-induced ALI in mice. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2015. For permissions please email: journals.permissions@oup.com.

Hard X-ray spectral investigations of gamma-ray bursts 120521C and 130606A at high-redshift z ˜ 6

NASA Astrophysics Data System (ADS)

Yasuda, T.; Urata, Y.; Enomoto, J.; Tashiro, M. S.

2017-04-01

This study presents a temporal and spectral analysis of the prompt emission of two high-redshift gamma-ray bursts (GRBs), 120521C at z ˜ 6 and 130606A at z ˜ 5.91, using data obtained from the Swift-XRT/BAT and the Suzaku-WAM simultaneously. Based on follow-up XRT observations, the longest durations of the prompt emissions were approximately 80 s (120521C) and 360 s (130606A) in the rest-frames of the two GRBs. These objects are thus categorized as long-duration GRBs; however, the durations are short compared with the predicted duration of GRBs originating from first-generation stars. Because of the wide bandpass of the instruments, covering the ranges 15 keV-5 MeV (BAT-WAM) and 0.3 keV-5.0 MeV (XRT-BAT-WAM), we could successfully determine the νFν peak energies E_peak^src in the rest-frame and isotropic-equivalent radiated energies Eiso; E^src_peak = 682^{+845}_{-207} keV and E_iso = (8. 25^{+2.24}_{-1.96}) × 10^{52} erg for 120521C, and E^src_peak = 1209^{+553}_{-304} keV and E_iso = (2.82^{+0.17}_{-0.71}) × 10^{53} erg for 130606A. These obtained characteristic parameters are in accordance with the well-known relationship between E_peak^src and Eiso (Amati relationship). In addition, we examined the relationships between E_peak^src and the 1-s peak luminosity, Lp, and between E_peak^src and the geometrical corrected radiated energy, Eγ, and confirmed the E_peak^src-Lp (Yonetoku) and E_peak^src-Eγ (Ghirlanda) relationships. The results imply that these high-redshift GRBs at z ˜ 6, which are expected to have radiated during the reionization epoch, have properties similar to those of low-redshift GRBs regarding X-ray prompt emission.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cho, Nancy L., E-mail: nlcho@partners.org; Lin, Chi-Iou; Du, Jinyan

Highlights: Black-Right-Pointing-Pointer Kinome profiling is a novel technique for identifying activated kinases in human cancers. Black-Right-Pointing-Pointer Src activity is increased in invasive thyroid cancers. Black-Right-Pointing-Pointer Inhibition of Src activity decreased proliferation and invasion in vitro. Black-Right-Pointing-Pointer Further investigation of Src targeted therapies in thyroid cancer is warranted. -- Abstract: Background: Novel therapies are needed for the treatment of invasive thyroid cancers. Aberrant activation of tyrosine kinases plays an important role in thyroid oncogenesis. Because current targeted therapies are biased toward a small subset of tyrosine kinases, we conducted a study to reveal novel therapeutic targets for thyroid cancer using amore » bead-based, high-throughput system. Methods: Thyroid tumors and matched normal tissues were harvested from twenty-six patients in the operating room. Protein lysates were analyzed using the Luminex immunosandwich, a bead-based kinase phosphorylation assay. Data was analyzed using GenePattern 3.0 software and clustered according to histology, demographic factors, and tumor status regarding capsular invasion, size, lymphovascular invasion, and extrathyroidal extension. Survival and invasion assays were performed to determine the effect of Src inhibition in papillary thyroid cancer (PTC) cells. Results: Tyrosine kinome profiling demonstrated upregulation of nine tyrosine kinases in tumors relative to matched normal thyroid tissue: EGFR, PTK6, BTK, HCK, ABL1, TNK1, GRB2, ERK, and SRC. Supervised clustering of well-differentiated tumors by histology, gender, age, or size did not reveal significant differences in tyrosine kinase activity. However, supervised clustering by the presence of invasive disease showed increased Src activity in invasive tumors relative to non-invasive tumors (60% v. 0%, p < 0.05). In vitro, we found that Src inhibition in PTC cells decreased cell invasion and proliferation. Conclusion: Global kinome analysis enables the discovery of novel targets for thyroid cancer therapy. Further investigation of Src targeted therapy for advanced thyroid cancer is warranted.« less

Communication: Cosolvency and cononsolvency explained in terms of a Flory-Huggins type theory

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dudowicz, Jacek, E-mail: dudowicz@jfi.uchicago.edu; Freed, Karl F.; Douglas, Jack F.

2015-10-07

Standard Flory-Huggins (FH) theory is utilized to describe the enigmatic cosolvency and cononsolvency phenomena for systems of polymers dissolved in mixed solvents. In particular, phase boundaries (specifically upper critical solution temperature spinodals) are calculated for solutions of homopolymers B in pure solvents and in binary mixtures of small molecule liquids A and C. The miscibility (or immiscibility) patterns for the ternary systems are classified in terms of the FH binary interaction parameters (χ{sub αβ}) and the ratio r = ϕ{sub A}/ϕ{sub C} of the concentrations ϕ{sub A} and ϕ{sub C} of the two solvents. The trends in miscibility are comparedmore » to those observed for blends of random copolymers (A{sub x}C{sub 1−x}) with homopolymers (B) and to those deduced for A/B/C solutions of polymers B in liquid mixtures of small molecules A and C that associate into polymeric clusters (A{sub p}C{sub q}){sub i}, (i = 1, 2, …, ∞). Although the classic FH theory is able to explain cosolvency and cononsolvency phenomena, the theory does not include a consideration of the mutual association of the solvent molecules and the competitive association between the solvent molecules and the polymer. These interactions can be incorporated in refinements of the FH theory, and the present paper provides a foundation for such extensions for modeling the rich thermodynamics of polymers in mixed solvents.« less

Derivation of Reliable Geometries in QM Calculations of DNA Structures: Explicit Solvent QM/MM and Restrained Implicit Solvent QM Optimizations of G-Quadruplexes.

PubMed

Gkionis, Konstantinos; Kruse, Holger; Šponer, Jiří

2016-04-12

Modern dispersion-corrected DFT methods have made it possible to perform reliable QM studies on complete nucleic acid (NA) building blocks having hundreds of atoms. Such calculations, although still limited to investigations of potential energy surfaces, enhance the portfolio of computational methods applicable to NAs and offer considerably more accurate intrinsic descriptions of NAs than standard MM. However, in practice such calculations are hampered by the use of implicit solvent environments and truncation of the systems. Conventional QM optimizations are spoiled by spurious intramolecular interactions and severe structural deformations. Here we compare two approaches designed to suppress such artifacts: partially restrained continuum solvent QM and explicit solvent QM/MM optimizations. We report geometry relaxations of a set of diverse double-quartet guanine quadruplex (GQ) DNA stems. Both methods provide neat structures without major artifacts. However, each one also has distinct weaknesses. In restrained optimizations, all errors in the target geometries (i.e., low-resolution X-ray and NMR structures) are transferred to the optimized geometries. In QM/MM, the initial solvent configuration causes some heterogeneity in the geometries. Nevertheless, both approaches represent a decisive step forward compared to conventional optimizations. We refine earlier computations that revealed sizable differences in the relative energies of GQ stems computed with AMBER MM and QM. We also explore the dependence of the QM/MM results on the applied computational protocol.

Application of Physiologically Based Pharmacokinetic Modeling in Understanding Bosutinib Drug-Drug Interactions: Importance of Intestinal P-Glycoprotein.

PubMed

Yamazaki, Shinji; Loi, Cho-Ming; Kimoto, Emi; Costales, Chester; Varma, Manthena V

2018-05-08

Bosutinib is an orally available Src/Abl tyrosine kinase inhibitor indicated for the treatment of patients with Ph+ chronic myelogenous leukemia at a clinically recommended dose of 500 mg once daily. Clinical results indicated that increases in bosutinib oral exposures were supra-proportional at the lower doses (50 to 200 mg) and approximately dose-proportional at the higher doses (200 to 600 mg). Bosutinib is a substrate of CYP3A4 and P-glycoprotein and exhibits pH-dependent solubility with moderate intestinal permeability. These findings led us to investigate the factors influencing the underlying pharmacokinetic mechanisms of bosutinib with physiologically-based pharmacokinetic (PBPK) models. Our primary objectives were to: 1) refine the previously developed bosutinib PBPK model based on the latest oral bioavailability data and 2) verify the refined PBPK model with P-glycoprotein kinetics based on the bosutinib drug-drug interaction (DDI) results with ketoconazole and rifampin. Additionally, the verified PBPK model was applied to predict bosutinib DDIs with dual CYP3A/P-glycoprotein inhibitors. The results indicated that 1) the refined PBPK model adequately described the observed plasma concentration-time profiles of bosutinib and 2) the verified PBPK model reasonably predicted the effects of ketoconazole and rifampin on bosutinib exposures by accounting for intestinal P-gp inhibition/induction. These results suggested that bosutinib DDI mechanism could involve not only CYP3A4-mediated metabolism but also P-glycoprotein-mediated efflux on absorption. In summary, P-glycoprotein kinetics could constitute a critical element in the PBPK models to understand the pharmacokinetic mechanism of dual CYP3A/P-glycoprotein substrates such as bosutinib exhibiting nonlinear pharmacokinetics due largely to a saturation of intestinal P-glycoprotein-mediated efflux. The American Society for Pharmacology and Experimental Therapeutics.

Formation of Polyphenol-Denatured Protein Flocs in Alcohol Beverages Sweetened with Refined Cane Sugars.

PubMed

Eggleston, Gillian; Triplett, Alexa

2017-11-08

The sporadic appearance of floc from refined, white cane sugars in alcohol beverages remains a technical problem for both beverage manufacturers and sugar refiners. Cane invert sugars mixed with 60% pure alcohol and water increased light scattering by up to ∼1000-fold. Insoluble and soluble starch, fat, inorganic ash, oligosaccharides, Brix, and pH were not involved in the prevailing floc-formation mechanism. Strong polynomial correlations existed between the haze floc and indicator values (IVs) (color at 420 nm pH 9.0/color at pH 4.0-an indirect measure of polyphenolic and flavonoid colorants) (R 2 = 0.815) and protein (R 2 = 0.819) content of the invert sugars. Ethanol-induced denaturation of the protein exposed hydrophobic polyphenol-binding sites that were further exposed when heated to 80 °C. A tentative mechanism for floc formation was advanced by molecular probing with a haze (floc) active protein and polyphenol as well as polar, nonpolar, and ionic solvents.

Solution NMR structure of a designed metalloprotein and complementary molecular dynamics refinement.

PubMed

Calhoun, Jennifer R; Liu, Weixia; Spiegel, Katrin; Dal Peraro, Matteo; Klein, Michael L; Valentine, Kathleen G; Wand, A Joshua; DeGrado, William F

2008-02-01

We report the solution NMR structure of a designed dimetal-binding protein, di-Zn(II) DFsc, along with a secondary refinement step employing molecular dynamics techniques. Calculation of the initial NMR structural ensemble by standard methods led to distortions in the metal-ligand geometries at the active site. Unrestrained molecular dynamics using a nonbonded force field for the metal shell, followed by quantum mechanical/molecular mechanical dynamics of DFsc, were used to relax local frustrations at the dimetal site that were apparent in the initial NMR structure and provide a more realistic description of the structure. The MD model is consistent with NMR restraints, and in good agreement with the structural and functional properties expected for DF proteins. This work demonstrates that NMR structures of metalloproteins can be further refined using classical and first-principles molecular dynamics methods in the presence of explicit solvent to provide otherwise unavailable insight into the geometry of the metal center.

Stardust Hypervelocity Entry Observing Campaign Support

NASA Technical Reports Server (NTRS)

Kontinos, Dean A.; Jordan, David E.; Jenniskens, Peter

2009-01-01

In the early morning of January 15, 2006, the Stardust Sample Return Capsule (SRC) successfully delivered its precious cargo of cometary particles to the awaiting recovery team at the Utah Test and Training Range (UTTR). As the SRC entered at 12.8 km/s, the fastest manmade object to traverse the atmosphere, a team of researchers imaged the event aboard the NASA DC-8 airborne observatory. At SRC entry, the airplane was at an altitude of 11.9 km positioned within 6.4 km of the prescribed, preferred target view location. The incoming SRC was first acquired approximately 18 seconds (s) after atmospheric interface and tracked for approximately 60 s, an observation period that is roughly centered in time around predicted peak heating.

International Space Station (ISS) External Television (TV) Camera Shutdown Investigation

NASA Technical Reports Server (NTRS)

Kichak, Robert; Young, Eric; Pandipati, Chetty; Cooke, Robert

2009-01-01

In the early morning of January 15, 2006, the Stardust Sample Return Capsule (SRC) successfully delivered its precious cargo of cometary particles to the awaiting recovery team at the Utah Test and Training Range (UTTR). As the SRC entered at 12.8 km/s, the fastest manmade object to traverse the atmosphere, a team of researchers imaged the event aboard the NASA DC-8 airborne observatory. At SRC entry, the airplane was at an altitude of 11.9 km positioned within 6.4 km of the prescribed, preferred target view location. The incoming SRC was first acquired approximately 18 seconds (s) after atmospheric interface and tracked for approximately 60 s, an observation period that is roughly centered in time around predicted peak heating.

National Energy Policy: Report of the National Energy Policy Development Group

DTIC Science & Technology

2001-05-01

and the environ- ment. Oil spill technology has improved dur- ing the last decade and will continue to do so. Risk assessments ...chemical intermediate in paper pulping and ore refining, and as a solvent. Transportation Sector: Private and public vehicles that move people and ...Energy Security and International Relationships Appendix One Summary of Report Recommendations Appendix Two Glossary 1 2 3 4 5 6 7 8 NATIONAL

New electrolytes for aluminum production: Ionic liquids

NASA Astrophysics Data System (ADS)

Zhang, Mingming; Kamavarum, Venkat; Reddy, Ramana G.

2003-11-01

In this article, the reduction, refining/recycling, and electroplating of aluminum from room-temperature molten salts are reviewed. In addition, the characteristics of several non-conventional organic solvents, electrolytes, and molten salts are evaluated, and the applicability of these melts for production of aluminum is discussed with special attention to ionic liquids. Also reviewed are electrochemical processes and conditions for electrodeposition of aluminum using ionic liquids at near room temperatures.

Crystal structure of a new alpha-cyclodextrin hydrate form. Molecular geometry and packing features: disordered solvent contribution.

PubMed

Puliti, R; Mattia, C A; Paduano, L

1998-08-01

The crystallographic study of a new hydrated form of alpha-cyclodextrin (cyclohexaamylose) is reported. C36H60O30 . 11H2O; space group P2(1)2(1)2(1) with cell constants a = 13.839(3), b = 15.398(3), c = 24.209(7) A; final discrepancy index R = 0.057 for the 5182 observed reflections and 632 refined parameters. Besides four ordered water molecules placed outside alpha-cyclodextrins, the structure shows regions of severely disordered solvent mainly confined in the oligosaccharide cavities. The contribution of the observed disorder has been computed via Fourier inversions of the residual electron density and incorporated into the structure factors in further refinements of the ordered part. The alpha-cyclodextrin molecule assumes a relaxed round shape stabilised by a ring sequence of all the six possible O2 ... O3 intramolecular hydrogen bonds. The four ordered water molecules take part in an extensive network of hydrogen bonds (infinite chains and loops) without modifying the scheme of intramolecular H-bonds or the (-)gauche conformation of O-6-H hydroxyl groups. The structure shows a new molecular arrangement, for an "empty" hydrated alpha-cyclodextrin, like that "brick-type" observed for alpha-CD in the iodoanilide trihydrate complex crystallising in an isomorphous cell.

Integrated report on the toxicological mitigation of coal liquids by hydrotreatment and other processes. [Petroleum and coal-derived products

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guerin, M.R.; Griest, W.H.; Ho, C.H.

1986-06-01

Research here on the toxicological properties of coal-derived liquids focuses on characterizing the refining process and refined products. Principle attention is given to the potential tumorigenicity of coal-derived fuels and to the identification of means to further reduce tumorigenicity should this be found necessary. Hydrotreatment is studied most extensively because it will be almost certainly required to produce commercial products and because it is likely to also greatly reduce tumorigenic activity relative to that of crude coal-liquid feedstocks. This report presents the results of a lifetime C3H mouse skin tumorigenicity assay of an H-Coal series of oils and considers themore » relationships between tumorigenicity, chemistry, and processing. Lifetime assay results are reported for an H-Coal syncrude mode light oil/heavy oil blend, a low severity hydrotreatment product, a high severity hydrotreatment product, a naphtha reformate, a heating oil, a petroleum-derived reformate, and a petroleum derived heating oil. Data are compared with those for an earlier study of an SRC-II blend and products of its hydrotreatment. Adequate data are presented to allow an independent qualitative assessment of the conclusions while statistical evaluation of the data is being completed. The report also documents the physical and chemical properties of the oils tested. 33 refs., 14 figs., 53 tabs.« less

Changes in signal transducer and activator of transcription 3 (STAT3) dynamics induced by complexation with pharmacological inhibitors of Src homology 2 (SH2) domain dimerization.

PubMed

Resetca, Diana; Haftchenary, Sina; Gunning, Patrick T; Wilson, Derek J

2014-11-21

The activity of the transcription factor signal transducer and activator of transcription 3 (STAT3) is dysregulated in a number of hematological and solid malignancies. Development of pharmacological STAT3 Src homology 2 (SH2) domain interaction inhibitors holds great promise for cancer therapy, and a novel class of salicylic acid-based STAT3 dimerization inhibitors that includes orally bioavailable drug candidates has been recently developed. The compounds SF-1-066 and BP-1-102 are predicted to bind to the STAT3 SH2 domain. However, given the highly unstructured and dynamic nature of the SH2 domain, experimental confirmation of this prediction was elusive. We have interrogated the protein-ligand interaction of STAT3 with these small molecule inhibitors by means of time-resolved electrospray ionization hydrogen-deuterium exchange mass spectrometry. Analysis of site-specific evolution of deuterium uptake induced by the complexation of STAT3 with SF-1-066 or BP-1-102 under physiological conditions enabled the mapping of the in silico predicted inhibitor binding site to the STAT3 SH2 domain. The binding of both inhibitors to the SH2 domain resulted in significant local decreases in dynamics, consistent with solvent exclusion at the inhibitor binding site and increased rigidity of the inhibitor-complexed SH2 domain. Interestingly, inhibitor binding induced hot spots of allosteric perturbations outside of the SH2 domain, manifesting mainly as increased deuterium uptake, in regions of STAT3 important for DNA binding and nuclear localization. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Targeting Src in Mucinous Ovarian Carcinoma

PubMed Central

Matsuo, Koji; Nishimura, Masato; Bottsford-Miller, Justin N.; Huang1, Jie; Komurov, Kakajan; Armaiz-Pena, Guillermo N.; Shahzad, Mian M. K.; Stone, Rebecca L.; Roh, Ju Won; Sanguino, Angela M.; Lu, Chunhua; Im, Dwight D.; Rosenshien, Neil B.; Sakakibara, Atsuko; Nagano, Tadayoshi; Yamasaki, Masato; Enomoto, Takayuki; Kimura, Tadashi; Ram, Prahlad T.; Schmeler, Kathleen M.; Gallick, Gary E.; Wong, Kwong K.; Frumovitz, Michael; Sood, Anil K.

2014-01-01

PURPOSE Mucinous ovarian carcinomas have a distinct clinical pattern compared to other subtypes of ovarian carcinoma. Here, we evaluated (i) stage-specific clinical significance of mucinous ovarian carcinomas in a large cohort and (ii) the functional role of src kinase in pre-clinical models of mucinous ovarian carcinoma. EXPERIMENTAL DESIGN 1302 ovarian cancer patients including 122 (9.4%) cases of mucinous carcinoma were evaluated for survival analyses. Biological effects of src kinase inhibition were tested in a novel orthotopic mucinous ovarian cancer model (RMUG-S-ip2) using dasatinib-based therapy. RESULTS Patients with advanced-stage mucinous ovarian cancer had significantly worse survival compared to those with serous histology: median overall survival, 1.67 versus 3.41 years, p=0.002; and median survival time after recurrence of 0.53 versus 1.66 years, p<0.0001. Among multiple ovarian cancer cell lines, RMUG-S-ip2 mucinous ovarian cancer cells showed the highest src kinase activity. Moreover, oxaliplatin treatment induced phosphorylation of src kinase. This induced activity by oxaliplatin therapy was inhibited by concurrent administration of dasatinib. Targeting src with dasatinib in vivo showed significant anti-tumor effects in the RMUG-S-ip2 model, but not in the serous ovarian carcinoma (SKOV3-TR) model. Combination therapy of oxaliplatin with dasatinib further demonstrated significant effects on reducing cell viability, increasing apoptosis, and in vivo anti-tumor effects in the RMUG-S-ip2 model. CONCLUSIONS Our results suggest that poor survival of women with mucinous ovarian carcinoma is associated with resistance to cytotoxic therapy. Targeting src kinase with combination of dasatinib and oxaliplatin may be an attractive approach in this disease. PMID:21737505

Inhibiting Src family tyrosine kinase activity blocks glutamate signalling to ERK1/2 and Akt/PKB but not JNK in cultured striatal neurones.

PubMed

Crossthwaite, Andrew J; Valli, Haseeb; Williams, Robert J

2004-03-01

Glutamate receptor activation of mitogen-activated protein (MAP) kinase signalling cascades has been implicated in diverse neuronal functions such as synaptic plasticity, development and excitotoxicity. We have previously shown that Ca2+-influx through NMDA receptors in cultured striatal neurones mediates the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt/protein kinase B (PKB) through a phosphatidylinositol 3-kinase (PI 3-kinase)-dependent pathway. Exposing neurones to the Src family tyrosine kinase inhibitor PP2, but not the inactive analogue PP3, inhibited NMDA receptor-induced phosphorylation of ERK1/2 and Akt/PKB in a concentration-dependent manner, and reduced cAMP response element-binding protein (CREB) phosphorylation. To establish a link between Src family tyrosine kinase-mediated phosphorylation and PI 3-kinase signalling, affinity precipitation experiments were performed with the SH2 domains of the PI 3-kinase regulatory subunit p85. This revealed a Src-dependent phosphorylation of a focal adhesion kinase (FAK)-p85 complex on glutamate stimulation. Demonstrating that PI3-kinase is not ubiquitously involved in NMDA receptor signal transduction, the PI 3-kinase inhibitors wortmannin and LY294002 did not prevent NMDA receptor Ca2+-dependent phosphorylation of c-Jun N-terminal kinase 1/2 (JNK1/2). Further, inhibiting Src family kinases increased NMDA receptor-dependent JNK1/2 phosphorylation, suggesting that Src family kinase-dependent cascades may physiologically limit signalling to JNK. These results demonstrate that Src family tyrosine kinases and PI3-kinase are pivotal regulators of NMDA receptor signalling to ERK/Akt and JNK in striatal neurones.

Letrozole regulates actin cytoskeleton polymerization dynamics in a SRC-1 dependent manner in the hippocampus of mice.

PubMed

Zhao, Yangang; Yu, Yanlan; Zhang, Yuanyuan; He, Li; Qiu, Linli; Zhao, Jikai; Liu, Mengying; Zhang, Jiqiang

2017-03-01

In the hippocampus, local estrogens (E 2 ) derived from testosterone that is catalyzed by aromatase play important roles in the regulation of hippocampal neural plasticity, but the underlying mechanisms remain unclear. The actin cytoskeleton contributes greatly to hippocampal synaptic plasticity; however, whether it is regulated by local E 2 and the related mechanisms remain to be elucidated. In this study, we first examined the postnatal developmental profiles of hippocampal aromatase and specific proteins responsible for actin cytoskeleton dynamics. Then we used aromatase inhibitor letrozole (LET) to block local E 2 synthesis and examined the changes of these proteins and steroid receptor coactivator-1 (SRC-1), the predominant coactivator for steroid nuclear receptors. Finally, SRC-1 specific RNA interference was used to examine the effects of SRC-1 on the expression of these actin remodeling proteins. The results showed a V-type profile for aromatase and increased profiles for actin cytoskeleton proteins in both male and female hippocampus without obvious sex differences. LET treatment dramatically decreased the F-actin/G-actin ratio, the expression of Rictor, phospho-AKT (ser473), Profilin-1, phospho-Cofilin (Ser3), and SRC-1 in a dose-dependent manner. In vitro studies demonstrated that LET induced downregulation of these proteins could be reversed by E 2 , and E 2 induced increase of these proteins were significantly suppressed by SRC-1 shRNA interference. These results for the first time clearly demonstrated that local E 2 inhibition could induce aberrant actin polymerization; they also showed an important role of SRC-1 in the mediation of local E 2 action on hippocampal synaptic plasticity by regulation of actin cytoskeleton dynamics. Copyright © 2016 Elsevier Ltd. All rights reserved.

Molecular signaling in live cells studied by FRET

NASA Astrophysics Data System (ADS)

Chien, Shu; Wang, Yingxiao

2011-11-01

Genetically encoded biosensors based on fluorescence resonance energy transfer (FRET) enables visualization of signaling events in live cells with high spatiotemporal resolution. We have used FRET to assess temporal and spatial characteristics for signaling molecules, including tyrosine kinases Src and FAK, small GTPase Rac, calcium, and a membrane-bound matrix metalloproteinase MT1-MMP. Activations of Src and Rac by platelet derived growth factor (PDGF) led to distinct subcellular patterns during cell migration on micropatterned surface, and these two enzymes interact with each other to form a feedback loop with differential regulations at different subcellular locations. We have developed FRET biosensors to monitor FAK activities at rafts vs. non-raft regions of plasma membrane in live cells. In response to cell adhesion on matrix proteins or stimulation by PDGF, the raft-targeting FAK biosensor showed a stronger FRET response than that at non-rafts. The FAK activation at rafts induced by PDGF is mediated by Src. In contrast, the FAK activation at rafts induced by adhesion is independent of Src activity, but rather is essential for Src activation. Thus, Src is upstream to FAK in response to chemical stimulation (PDGF), but FAK is upstream to Src in response to mechanical stimulation (adhesion). A novel biosensor has been developed to dynamically visualize the activity of membrane type-1-matrix metalloproteinase (MT1-MMP), which proteolytically remodels the extracellular matrix. Epidermal growth factor (EGF) directed active MT1-MMP to the leading edge of migrating live cancer cells with local accumulation of EGF receptor via a process dependent on an intact cytoskeletal network. In summary, FRET-based biosensors enable the elucidation of molecular processes and hierarchies underlying spatiotemporal regulation of biological and pathological processes, thus advancing our knowledge on how cells perceive mechanical/chemical cues in space and time to coordinate molecular/cellular functions.

Molecular signaling in live cells studied by FRET

NASA Astrophysics Data System (ADS)

Chien, Shu; Wang, Yingxiao

2012-03-01

Genetically encoded biosensors based on fluorescence resonance energy transfer (FRET) enables visualization of signaling events in live cells with high spatiotemporal resolution. We have used FRET to assess temporal and spatial characteristics for signaling molecules, including tyrosine kinases Src and FAK, small GTPase Rac, calcium, and a membrane-bound matrix metalloproteinase MT1-MMP. Activations of Src and Rac by platelet derived growth factor (PDGF) led to distinct subcellular patterns during cell migration on micropatterned surface, and these two enzymes interact with each other to form a feedback loop with differential regulations at different subcellular locations. We have developed FRET biosensors to monitor FAK activities at rafts vs. non-raft regions of plasma membrane in live cells. In response to cell adhesion on matrix proteins or stimulation by PDGF, the raft-targeting FAK biosensor showed a stronger FRET response than that at non-rafts. The FAK activation at rafts induced by PDGF is mediated by Src. In contrast, the FAK activation at rafts induced by adhesion is independent of Src activity, but rather is essential for Src activation. Thus, Src is upstream to FAK in response to chemical stimulation (PDGF), but FAK is upstream to Src in response to mechanical stimulation (adhesion). A novel biosensor has been developed to dynamically visualize the activity of membrane type-1-matrix metalloproteinase (MT1-MMP), which proteolytically remodels the extracellular matrix. Epidermal growth factor (EGF) directed active MT1-MMP to the leading edge of migrating live cancer cells with local accumulation of EGF receptor via a process dependent on an intact cytoskeletal network. In summary, FRET-based biosensors enable the elucidation of molecular processes and hierarchies underlying spatiotemporal regulation of biological and pathological processes, thus advancing our knowledge on how cells perceive mechanical/chemical cues in space and time to coordinate molecular/cellular functions.

Nuclear receptor coactivators function in estrogen receptor- and progestin receptor-dependent aspects of sexual behavior in female rats

PubMed Central

Molenda-Figueira, Heather A.; Williams, Casey A.; Griffin, Andreana L.; Rutledge, Eric M.; Blaustein, Jeffrey D.; Tetel, Marc J.

2008-01-01

The ovarian hormones, estradiol (E) and progesterone (P) facilitate the expression of sexual behavior in female rats. E and P mediate many of these behavioral effects by binding to their respective intracellular receptors in specific brain regions. Nuclear receptor coactivators, including Steroid Receptor Coactivator-1 (SRC-1) and CREB Binding Protein (CBP), dramatically enhance ligand-dependent steroid receptor transcriptional activity in vitro. Previously, our lab has shown that SRC-1 and CBP modulate estrogen receptor (ER)-mediated induction of progestin receptor (PR) gene expression in the ventromedial nucleus of the hypothalamus (VMN) and hormone-dependent sexual receptivity in female rats. Female sexual behaviors can be activated by high doses of E alone in ovariectomized rats, and thus are believed to be ER-dependent. However, the full repertoire of female sexual behavior, in particular, proceptive behaviors such as hopping, darting and ear wiggling, are considered to be PR-dependent. In the present experiments, the function of SRC-1 and CBP in distinct ER- (Exp. 1) and PR- (Exp. 2) dependent aspects of female sexual behavior was investigated. In Exp. 1, infusion of antisense oligodeoxynucleotides to SRC-1 and CBP mRNA into the VMN decreased lordosis intensity in rats treated with E alone, suggesting that these coactivators modulate ER-mediated female sexual behavior. In Exp. 2, antisense to SRC-1 and CBP mRNA around the time of P administration reduced PR-dependent ear wiggling and hopping and darting. Taken together, these data suggest that SRC-1 and CBP modulate ER and PR action in brain and influence distinct aspects of hormone-dependent sexual behaviors. These findings support our previous studies and provide further evidence that SRC-1 and CBP function together to regulate ovarian hormone action in behaviorally-relevant brain regions. PMID:16769066

Water use of a multigenotype poplar short-rotation coppice from tree to stand scale.

PubMed

Bloemen, Jasper; Fichot, Régis; Horemans, Joanna A; Broeckx, Laura S; Verlinden, Melanie S; Zenone, Terenzio; Ceulemans, Reinhart

2017-02-01

Short-rotation coppice (SRC) has great potential for supplying biomass-based heat and energy, but little is known about SRC's ecological footprint, particularly its impact on the water cycle. To this end, we quantified the water use of a commercial scale poplar ( Populus ) SRC plantation in East Flanders (Belgium) at tree and stand level, focusing primarily on the transpiration component. First, we used the AquaCrop model and eddy covariance flux data to analyse the different components of the stand-level water balance for one entire growing season. Transpiration represented 59% of evapotranspiration (ET) at stand scale over the whole year. Measured ET and modelled ET were lower as compared to the ET of reference grassland, suggesting that the SRC only used a limited amount of water. Secondly, we compared leaf area scaled and sapwood area scaled sap flow ( F s ) measurements on individual plants vs. stand scale eddy covariance flux data during a 39-day intensive field campaign in late summer 2011. Daily stem diameter variation (∆ D ) was monitored simultaneously with F s to understand water use strategies for three poplar genotypes. Canopy transpiration based on sapwood area or leaf area scaling was 43.5 and 50.3 mm, respectively, and accounted for 74%, respectively, 86%, of total ecosystem ET measured during the intensive field campaign. Besides differences in growth, the significant intergenotypic differences in daily ∆ D (due to stem shrinkage and swelling) suggested different water use strategies among the three genotypes which were confirmed by the sap flow measurements. Future studies on the prediction of SRC water use, or efforts to enhance the biomass yield of SRC genotypes, should consider intergenotypic differences in transpiration water losses at tree level as well as the SRC water balance at stand level.

Self-reported Concussion History and Sensorimotor Tests Predict Head/Neck Injuries.

PubMed

Hides, Julie A; Franettovich Smith, Melinda M; Mendis, M Dilani; Treleaven, Julia; Rotstein, Andrew H; Sexton, Christopher T; Low Choy, Nancy; McCrory, Paul

2017-12-01

Sport-related concussion (SRC) is a risk for players involved in high-impact, collision sports. A history of SRC is a risk factor for future concussions, but the mechanisms underlying this are unknown. Despite evidence that most visible signs and symptoms associated with sports concussion resolve within 7-10 d, it has been proposed that subclinical loss of neuromuscular control and impaired motor functioning may persist and be associated with further injury. Alternatively, indicators of poor sensorimotor performance could be independent risk factors. This study investigated if a history of SRC and/or preseason sensorimotor performance predicted season head/neck injuries. A total of 190 male rugby league, rugby union, and Australian Football League players participated. Preseason assessments included self-report of SRC within the previous 12 months and a suite of measures of sensorimotor function (balance, vestibular function, cervical proprioception, and trunk muscle function). Head/neck injury data were collected in the playing season. Forty-seven players (25%) reported a history of SRC. A history of concussion was related to changes in size and contraction of trunk muscles. Twenty-two (11.6%) players sustained a head/neck injury during the playing season, of which, 14 (63.6%) players had a history of SRC. Predictors of in-season head/neck injuries included history of SRC, trunk muscle function, and cervical proprioceptive errors. Five risk factors were identified, and players with three or more of these had 14 times greater risk of sustaining a season neck/head injury (sensitivity of 75% and specificity of 82.5%) than did players with two or fewer risk factors. The modifiable risk factors identified could be used to screen football players in the preseason and guide the development of exercise programs aimed at injury reduction.

Selective Targeting of SH2 Domain-Phosphotyrosine Interactions of Src Family Tyrosine Kinases with Monobodies.

PubMed

Kükenshöner, Tim; Schmit, Nadine Eliane; Bouda, Emilie; Sha, Fern; Pojer, Florence; Koide, Akiko; Seeliger, Markus; Koide, Shohei; Hantschel, Oliver

2017-05-05

The binding of Src-homology 2 (SH2) domains to phosphotyrosine (pY) sites is critical for the autoinhibition and substrate recognition of the eight Src family kinases (SFKs). The high sequence conservation of the 120 human SH2 domains poses a significant challenge to selectively perturb the interactions of even the SFK SH2 family against the rest of the SH2 domains. We have developed synthetic binding proteins, termed monobodies, for six of the SFK SH2 domains with nanomolar affinity. Most of these monobodies competed with pY ligand binding and showed strong selectivity for either the SrcA (Yes, Src, Fyn, Fgr) or SrcB subgroup (Lck, Lyn, Blk, Hck). Interactome analysis of intracellularly expressed monobodies revealed that they bind SFKs but no other SH2-containing proteins. Three crystal structures of monobody-SH2 complexes unveiled different and only partly overlapping binding modes, which rationalized the observed selectivity and enabled structure-based mutagenesis to modulate inhibition mode and selectivity. In line with the critical roles of SFK SH2 domains in kinase autoinhibition and T-cell receptor signaling, monobodies binding the Src and Hck SH2 domains selectively activated respective recombinant kinases, whereas an Lck SH2-binding monobody inhibited proximal signaling events downstream of the T-cell receptor complex. Our results show that SFK SH2 domains can be targeted with unprecedented potency and selectivity using monobodies. They are excellent tools for dissecting SFK functions in normal development and signaling and to interfere with aberrant SFK signaling networks in cancer cells. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

REVERSE SIGNALING BY GPI-LINKED MANDUCA EPHRIN REQUIRES A SRC FAMILY KINASE TO RESTRICT NEURONAL MIGRATION IN VIVO

PubMed Central

Coate, Thomas M.; Swanson, Tracy L.; Copenhaver, Philip F.

2011-01-01

Reverse signaling via GPI-linked Ephrins may help control cell proliferation and outgrowth within the nervous system, but the mechanisms underlying this process remain poorly understood. In the embryonic enteric nervous system (ENS) of the moth Manduca sexta, migratory neurons forming the enteric plexus (EP cells) express a single Ephrin ligand (GPI-linked MsEphrin), while adjacent midline cells that are inhibitory to migration express the cognate receptor (MsEph). Knocking down MsEph receptor expression in cultured embryos with antisense morpholino oligonucleotides allowed the EP cells to cross the midline inappropriately, consistent with the model that reverse signaling via MsEphrin mediates a repulsive response in the ENS. Src family kinases have been implicated in reverse signaling by type-A Ephrins in other contexts, and MsEphrin colocalizes with activated forms of endogenous Src in the leading processes of the EP cells. Pharmacological inhibition of Src within the developing ENS induced aberrant midline crossovers, similar to the effect of blocking MsEphrin reverse signaling. Hyperstimulating MsEphrin reverse signaling with MsEph-Fc fusion proteins induced the rapid activation of endogenous Src specifically within the EP cells, as assayed by Western blots of single embryonic gut explants and by whole-mount immunostaining of cultured embryos. In longer cultures, treatment with MsEph-Fc caused a global inhibition of EP cell migration and outgrowth, an effect that was prevented by inhibiting Src activation. These results support the model that MsEphrin reverse signaling induces the Src-dependent retraction of EP cell processes away from the enteric midline, thereby helping to confine the neurons to their appropriate pathways. PMID:19295147

Development of Coactivator-Dependent, First-in-Class Therapies for Breast Cancer

DTIC Science & Technology

2014-09-01

star: AMP-activated protein kinase stimulates fat absorption. Cell Metab. 13:1–2 53. Reineke EL, York B, Stashi E, et al. 2012. SRC-2 coactivator...receptor/SRC-3 protein complexes achieved by our group are providing powerful new insights into understanding the conformation of intact, full...length proteins in a complex and should provide valuable new information on the mechanism of action of SRC SMIs as well. 15. SUBJECT TERMS Breast

STEROID RECEPTOR COACTIVATOR 2 (SRC-2) MODULATES STEROID-DEPENDENT MALE SEXUAL BEHAVIOR AND NEUROPLASTICITY IN JAPANESE QUAIL (COTURNIX JAPONICA)

PubMed Central

Niessen, Neville-Andrew; Balthazart, Jacques; Ball, Gregory F.; Charlier, Thierry D.

2011-01-01

Steroid receptor coactivators are necessary for efficient transcriptional regulation by ligand-bound nuclear receptors, including estrogen and androgen receptors. SRC-2 modulates estrogen- and progesterone-dependent sexual behavior in female rats but its implication in the control of male sexual behavior has not been studied to our knowledge. We cloned and sequenced the complete quail SRC-2 transcript and showed by semi-quantitative PCR that SRC-2 expression is nearly ubiquitous, with high levels of expression in the kidney, cerebellum and diencephalon. Real time quantitative PCR did not reveal any differences between intact males and females the medial preoptic nucleus (POM), optic lobes and cerebellum. We next investigated the physiological and behavioral role of this coactivator using in vivo antisense oligonucleotide (AS) techniques. Daily injections in the third ventricle at the level of the POM of locked nucleic acid antisense targeting SRC-2 significantly reduced the expression of testosterone-dependent male-typical copulatory behavior but no inhibition of one aspect of the appetitive sexual behavior was observed. The volume of POM, defined by aromatase-immunoreactive cells, was markedly decreased in animals treated with AS as compared to controls. These results demonstrate that SRC-2 plays a prominent role in the control of steroid-dependent male sexual behavior and its associated neuroplasticity in Japanese quail. PMID:21854393

Effluviibacter roseus gen. nov., sp. nov., isolated from muddy water, belonging to the family "Flexibacteraceae".

PubMed

Suresh, K; Mayilraj, S; Chakrabarti, T

2006-07-01

A Gram-negative bacterial isolate (designated SRC-1(T)) was isolated from an occasional drainage system and characterized by a polyphasic approach to determine its taxonomic position. Phylogenetic analysis based on 16S rRNA gene sequences affiliated strain SRC-1(T) with the family "Flexibacteraceae" of the phylum Bacteroidetes. It showed greatest sequence similarity to Pontibacter actiniarum KMM 6156(T) (95.5 %) followed by Adhaeribacter aquaticus MBRG1.5(T) (89.0 %) and Hymenobacter roseosalivarius DSM 11622(T) (88.9 %), but it differed from these micro-organisms in many phenotypic characteristics. Strain SRC-1(T) was an obligate aerobe and its cells were non-motile, irregular rods. The major fatty acids included mainly unsaturated and hydroxy fatty acids, including 17 : 1 iso I/anteiso B (36.7 %), 15 : 0 iso (15.8 %) and 17 : 0 iso 3-OH (10.3 %), and the DNA G+C content was 59.5 mol%. From the phenotypic and genotypic analyses it was clear that strain SRC-1(T) was quite different from members other genera in the family '"Flexibacteraceae". Therefore we conclude that strain SRC-1(T) represents a novel genus, for which the name Effluviibacter gen. nov., containing a single species Effluviibacter roseus sp. nov., is proposed. The type species of the genus is Effluviibacter roseus, the type strain of which is strain SRC-1(T) (=MTCC 7260(T)=DSM 17521(T)).

Vascular endothelial growth factor receptor-1 mediates migration of human colorectal carcinoma cells by activation of Src family kinases

PubMed Central

Lesslie, D P; Summy, J M; Parikh, N U; Fan, F; Trevino, J G; Sawyer, T K; Metcalf, C A; Shakespeare, W C; Hicklin, D J; Ellis, L M; Gallick, G E

2006-01-01

Vascular endothelial growth factor (VEGF) is the predominant pro-angiogenic cytokine in human malignancy, and its expression correlates with disease recurrence and poor outcomes in patients with colorectal cancer. Recently, expression of vascular endothelial growth factor receptors (VEGFRs) has been observed on tumours of epithelial origin, including those arising in the colon, but the molecular mechanisms governing potential VEGF-driven biologic functioning in these tumours are not well characterised. In this report, we investigated the role of Src family kinases (SFKs) in VEGF-mediated signalling in human colorectal carcinoma (CRC) cell lines. Vascular endothelial growth factor specifically activated SFKs in HT29 and KM12L4 CRC cell lines. Further, VEGF stimulation resulted in enhanced cellular migration, which was effectively blocked by pharmacologic inhibition of VEGFR-1 or Src kinase. Correspondingly, migration studies using siRNA clones with reduced Src expression confirmed the requirement for Src in VEGF-induced migration in these cells. Furthermore, VEGF treatment enhanced VEGFR-1/SFK complex formation and increased tyrosine phosphorylation of focal adhesion kinase, p130 cas and paxillin. Finally, we demonstrate that VEGF-induced migration is not due, at least in part, to VEGF acting as a mitogen. These results suggest that VEGFR-1 promotes migration of tumour cells through a Src-dependent pathway linked to activation of focal adhesion components that regulate this process. PMID:16685275

Coupled motions in the SH2 and kinase domains of Csk control Src phosphorylation.

PubMed

Wong, Lilly; Lieser, Scot A; Miyashita, Osamu; Miller, Meghan; Tasken, Kjetil; Onuchic, Josè N; Adams, Joseph A; Woods, Virgil L; Jennings, Patricia A

2005-08-05

The C-terminal Src kinase (Csk) phosphorylates and down-regulates Src family tyrosine kinases. The Csk-binding protein (Cbp) localizes Csk close to its substrates at the plasma membrane, and increases the specific activity of the kinase. To investigate this long-range catalytic effect, the phosphorylation of Src and the conformation of Csk were investigated in the presence of a high-affinity phosphopeptide derived from Cbp. This peptide binds tightly to the SH2 domain and enhances Src recognition (lowers K(m)) by increasing the apparent phosphoryl transfer rate in the Csk active site, a phenomenon detected in rapid quench flow experiments. Previous studies demonstrated that the regulation of Csk activity is linked to conformational changes in the enzyme that can be probed with hydrogen-deuterium exchange methods. We show that the Cbp peptide impacts deuterium incorporation into its binding partner (the SH2 domain), and into the SH2-kinase linker and several sequences in the kinase domain, including the glycine-rich loop in the active site. These findings, along with computational data from normal mode analyses, suggest that the SH2 domain moves in a cantilever fashion with respect to the small lobe of the kinase domain, ordering the active site for catalysis. The binding of a small Cbp-derived peptide to the SH2 domain of Csk modifies these motions, enhancing Src recognition.

Alcohol approach tendencies in heavy drinkers: comparison of effects in a Relevant Stimulus-Response Compatibility task and an approach/avoidance Simon task.

PubMed

Field, Matt; Caren, Rhiane; Fernie, Gordon; De Houwer, Jan

2011-12-01

Several recent studies suggest that alcohol-related cues elicit automatic approach tendencies in heavy drinkers. A variety of tasks have been used to demonstrate these effects, including Relevant Stimulus-Response Compatibility (R-SRC) tasks and variants of Simon tasks. Previous work with normative stimuli suggests that the R-SRC task may be more sensitive than Simon tasks because the activation of approach tendencies may depend on encoding of the stimuli as alcohol-related, which occurs in the R-SRC task but not in Simon tasks. Our aim was to directly compare these tasks for the first time in the context of alcohol use. We administered alcohol versions of an R-SRC task and a Simon task to 62 social drinkers, who were designated as heavy or light drinkers based on a median split of their weekly alcohol consumption. Results indicated that, compared to light drinkers, heavy drinkers were faster to approach, rather than avoid, alcohol-related pictures in the R-SRC task but not in the Simon task. Theoretical implications and methodological issues are discussed.

Shikonin induces apoptosis and inhibits migration of ovarian carcinoma cells by inhibiting the phosphorylation of Src and FAK

PubMed Central

HAO, ZHENFENG; QIAN, JING; YANG, JISHI

2015-01-01

The present study identified that shikonin, a naphthoquinone extracted from the roots of Lithospermum erythrorhizon, inhibits the migration of ovarian cancer cells and induces their apoptosis by impairing the phosphorylation of two kinases, proto-oncogene tyrosine protein kinase Src (Src) and focal adhesion kinase (FAK). Ovarian carcinoma SKOV-3 cells were treated with various concentrations of shikonin and analyzed for the effects on cell migration, invasion and apoptosis via Transwell assays and flow cytometry. In addition, the effects of shikonin administration on the expression and phosphorylation of Src and FAK in the SKOV-3 cells were analyzed by western blotting. Shikonin appeared to induce apoptosis and decrease cell migration in the SKOV-3 ovarian cells. Furthermore, the present study provides evidence that shikonin may exert these effects on human ovarian carcinoma cells via the inhibition of the protein tyrosine kinases, Src and FAK. Thus, shikonin should be considered for additional investigation as a candidate agent for the prevention and treatment of human ovarian cancer. PMID:25621031

GRP78 confers the resistance to 5-FU by activating the c-Src/LSF/TS axis in hepatocellular carcinoma.

PubMed

Gu, Yan-jiao; Li, Hong-dan; Zhao, Liang; Zhao, Song; He, Wu-bin; Rui, Li; Su, Chang; Zheng, Hua-chuan; Su, Rong-jian

2015-10-20

5-FU is a common first-line chemotherapeutic drug for the treatment of hepatocellular carcinoma. However the development of acquired resistance to 5-FU confines its clinical usages. Although this phenomenon has been the subject of intense investigation, the exact mechanism of acquired resistance to 5-FU remains elusive. Here, we report that over-expression of GRP78 contributes to acquired resistance to 5-FU in HCC by up-regulating the c-Src/LSF/TS axis. Moreover, we found that the resistance to 5-FU conferred by GRP78 is mediated by its ATPase domain. The ATPase domain differentially increased the expression of LSF, TS and promoted the phosphorylation of ERK and Akt. We further identified that GRP78 interacts physically with c-Src through its ATPase domain and promotes the phosphorylation of c-Src, which in turn increases the expression of LSF in the nucleus. Together, GRP78 confers the resistance to 5-FU by up-regulating the c-Src/LSF/TS axis via its ATPase domain.

Aging of SRC liquids

NASA Astrophysics Data System (ADS)

Hara, T.; Jones, L.; Tewari, K. C.; Li, N. C.

1981-02-01

The viscosity of SRC-LL liquid increases when subjected to accelerated aging by bubbling oxygen in the presence of copper strip at 62°C. Precipitates are formed and can be separated from the aged liquid by Soxhlet extraction with pentane. A 30-70 blend of SRC-I with SRC-LL was subjected to oxygen aging in the absence of copper, and the viscosity increased dramatically after 6 days at 62°. The content of preasphaltene and its molecular size increase with time of aging, accompanied by decrease of asphaltene and pentane-soluble contents. For the preasphaltene fraction on aging, gel permeation chromatography shows formation of larger particles. ESR experiments show that with oxygen aging, spin concentration in the preasphaltene fraction decreases. Perhaps some semiquinone, together with di- and tri-substituted phenoxy radicals, generated by oxygen aging of the coal liquid, interact with the free radicals already present in coal to yield larger particles and reduce free radical concentration. We are currently using the very high-field (600-MHz) NMR spectrometer at Mellon Institute to determine changes in structural parameters before and after aging of SRC-II and its chromatographically separated fractions.

Probing short-range correlations in asymmetric nuclei with quasi-free pair knockout reactions

NASA Astrophysics Data System (ADS)

Stevens, Sam; Ryckebusch, Jan; Cosyn, Wim; Waets, Andreas

2018-02-01

Short-range correlations (SRC) in asymmetric nuclei with an unusual neutron-to-proton ratio can be studied with quasi-free two-nucleon knockout processes following the collision between accelerated ions and a proton target. We derive an approximate factorized cross section for those SRC-driven p (A ,p‧N1N2) reactions. Our reaction model hinges on the factorization properties of SRC-driven A (e ,e‧N1N2) reactions for which strong indications are found in theory-experiment comparisons. In order to put our model to the test we compare its predictions with results of 12C (p ,p‧ pn) measurements conducted at Brookhaven National Laboratory (BNL) and find a fair agreement. The model can also reproduce characteristic features of SRC-driven two-nucleon knockout reactions, like back-to-back emission of the correlated nucleons. We study the asymmetry dependence of nuclear SRC by providing predictions for the ratio of proton-proton to proton-neutron knockout cross sections for the carbon isotopes 9-15C thereby covering neutron excess values (N - Z) / Z between -0.5 and +0.5.

Inhibition of c-Src protects paraquat induced microvascular endothelial injury by modulating caveolin-1 phosphorylation and caveolae mediated transcellular permeability.

PubMed

Huang, Yu; He, Qing

2017-06-01

The mechanisms underlying paraquat induced acute lung injury (ALI) is still not clear. C-Src plays an important role in the regulation of microvascular endothelial barrier function and the pathogenesis of ALI. In the present study, we found that paraquat induced cell toxicity and an increase of reactive oxygen species (ROS) in endothelium. Paraquat exposure also induced significant increase of caveolin-1 phosphorylation, caveolae trafficking and albumin permeability in endothelial monolayers. C-Src depletion by siRNA significantly attenuate paraquat induced cell toxicity, caveolin-1 phosphorylation, caveolae formation and endothelial hyperpermeability. N-acetylcysteine (NAC) failed to protect endothelial monolayers against paraquat induced toxicity. Thus, our findings suggest that paraquat exposure increases paracellular endothelial permeability by increasing caveolin-1 phosphorylation in a c-Src dependant manner. The depletion of c-Src might protect microvascular endothelial function by regulating caveolin-1 phosphorylation and caveolae trafficking during paraquat exposure, and might have potential therapeutic effects on paraquat induced ALI. Copyright © 2017 Elsevier B.V. All rights reserved.

Revisiting the ERK/Src cortactin switch

PubMed Central

Kelley, Laura C; Hayes, Karen E; Ammer, Amanda Gatesman; Martin, Karen H

2011-01-01

The filamentous (F)-actin regulatory protein cortactin plays an important role in tumor cell movement and invasion by promoting and stabilizing actin related protein (Arp)2/3-mediated actin networks necessary for plasma membrane protrusion. Cortactin is a substrate for ERK1/2 and Src family kinases, with previous in vitro findings demonstrating ERK1/2 phosphorylation of cortactin as a positive and Src phosphorylation as a negative regulatory event in promoting Arp2/3 activation through neuronal Wiskott Aldrich Syndrome protein (N-WASp). Evidence for this regulatory cortactin “switch” in cells has been hampered due to the lack of phosphorylation-specific antibodies that recognize ERK1/2-phosphorylated cortactin. Our findings with phosphorylation-specific antibodies against these ERK1/2 sites (pS405 and pS418) indicate that cortactin can be co-phosphorylated at 405/418 and tyrosine residues targeted by Src family tyrosine kinases. These results indicate that the ERK/Src cortactin switch is not the sole mechanism by which ERK1/2 and tyrosine phosphorylation events regulate cortactin function in cell systems. PMID:21655441

Examining the psychometric properties of a sport-related concussion survey: a Rasch measurement approach.

PubMed

Hecimovich, Mark; Marais, Ida

2017-06-26

Awareness of sport-related concussion (SRC) is an essential step in increasing the number of athletes or parents who report on SRC. This awareness is important, as there is no established data on medical care at youth-level sports and may be limited to individuals with only first aid training. In this circumstance, aside from the coach, it is the players and their parents who need to be aware of possible signs and symptoms. The aim of this study was to examine the psychometric properties of a parent and player concussion survey intended for use before and after an education campaign regarding SRC. 1441 questionnaires were received from parents and 284 questionnaires from players. The responses to the sixteen-item section of the questionnaire's 'recognition of signs and symptoms' were submitted to psychometric analysis using the dichotomous and polytomous Rasch model via the Rasch Unidimensional Measurement Model software RUMM2030. The Rasch model of Modern Test Theory can be considered a refinement of, or advance on, traditional analyses of an instrument's psychometric properties. The main finding is that these sixteen items measure two factors: items that are symptoms of concussion and items that are not symptoms of concussion. Parents and athletes were able to identify most or all of the symptoms, but were not as good at distinguishing symptoms that are not symptoms of concussion. Analyzing these responses revealed differential item functioning for parents and athletes on non-symptom items. When the DIF was resolved a significant difference was found between parents and athletes. The main finding is that the items measure two 'dimensions' in concussion symptom recognition. The first dimension consists of those items that are symptoms of concussion and the second dimension of those items that are not symptoms of concussion. Parents and players were able to identify most or all of the symptoms of concussion, so one would not expect to pick up any positive change on these items after an education campaign. Parents and players were not as good at distinguishing symptoms that are not symptoms of concussion. It is on these items that one may possibly expect improvement to manifest, so to evaluate the effectiveness of an education campaign it would pay to look for improvement in distinguishing symptoms that are not symptoms of concussion.

Identification of N-methyl-D-aspartic acid (NMDA) receptor subtype-specific binding sites that mediate direct interactions with scaffold protein PSD-95.

PubMed

Cousins, Sarah L; Stephenson, F Anne

2012-04-13

N-methyl-D-aspartate (NMDA) neurotransmitter receptors and the postsynaptic density-95 (PSD-95) membrane-associated guanylate kinase (MAGUK) family of scaffolding proteins are integral components of post-synaptic macromolecular signaling complexes that serve to propagate glutamate responses intracellularly. Classically, NMDA receptor NR2 subunits associate with PSD-95 MAGUKs via a conserved ES(E/D)V amino acid sequence located at their C termini. We previously challenged this dogma to demonstrate a second non-ES(E/D)V PSD-95-binding site in both NMDA receptor NR2A and NR2B subunits. Here, using a combination of co-immunoprecipitations from transfected mammalian cells, yeast two-hybrid interaction assays, and glutathione S-transferase (GST) pulldown assays, we show that NR2A subunits interact directly with PSD-95 via the C-terminal ESDV motif and additionally via an Src homology 3 domain-binding motif that associates with the Src homology 3 domain of PSD-95. Peptide inhibition of co-immunoprecipitations of NR2A and PSD-95 demonstrates that both the ESDV and non-ESDV sites are required for association in native brain tissue. Furthermore, we refine the non-ESDV site within NR2B to residues 1149-1157. These findings provide a molecular basis for the differential association of NMDA receptor subtypes with PSD-95 MAGUK scaffold proteins. These selective interactions may contribute to the organization, lateral mobility, and ultimately the function of NMDA receptor subtypes at synapses. Furthermore, they provide a more general molecular mechanism by which the scaffold, PSD-95, may discriminate between potential interacting partner proteins.

Identification of N-Methyl-d-aspartic Acid (NMDA) Receptor Subtype-specific Binding Sites That Mediate Direct Interactions with Scaffold Protein PSD-95*

PubMed Central

Cousins, Sarah L.; Stephenson, F. Anne

2012-01-01

N-methyl-d-aspartate (NMDA) neurotransmitter receptors and the postsynaptic density-95 (PSD-95) membrane-associated guanylate kinase (MAGUK) family of scaffolding proteins are integral components of post-synaptic macromolecular signaling complexes that serve to propagate glutamate responses intracellularly. Classically, NMDA receptor NR2 subunits associate with PSD-95 MAGUKs via a conserved ES(E/D)V amino acid sequence located at their C termini. We previously challenged this dogma to demonstrate a second non-ES(E/D)V PSD-95-binding site in both NMDA receptor NR2A and NR2B subunits. Here, using a combination of co-immunoprecipitations from transfected mammalian cells, yeast two-hybrid interaction assays, and glutathione S-transferase (GST) pulldown assays, we show that NR2A subunits interact directly with PSD-95 via the C-terminal ESDV motif and additionally via an Src homology 3 domain-binding motif that associates with the Src homology 3 domain of PSD-95. Peptide inhibition of co-immunoprecipitations of NR2A and PSD-95 demonstrates that both the ESDV and non-ESDV sites are required for association in native brain tissue. Furthermore, we refine the non-ESDV site within NR2B to residues 1149–1157. These findings provide a molecular basis for the differential association of NMDA receptor subtypes with PSD-95 MAGUK scaffold proteins. These selective interactions may contribute to the organization, lateral mobility, and ultimately the function of NMDA receptor subtypes at synapses. Furthermore, they provide a more general molecular mechanism by which the scaffold, PSD-95, may discriminate between potential interacting partner proteins. PMID:22375001

Iron catalyzed coal liquefaction process

DOEpatents

Garg, Diwakar; Givens, Edwin N.

1983-01-01

A process is described for the solvent refining of coal into a gas product, a liquid product and a normally solid dissolved product. Particulate coal and a unique co-catalyst system are suspended in a coal solvent and processed in a coal liquefaction reactor, preferably an ebullated bed reactor. The co-catalyst system comprises a combination of a stoichiometric excess of iron oxide and pyrite which reduce predominantly to active iron sulfide catalysts in the reaction zone. This catalyst system results in increased catalytic activity with attendant improved coal conversion and enhanced oil product distribution as well as reduced sulfide effluent. Iron oxide is used in a stoichiometric excess of that required to react with sulfur indigenous to the feed coal and that produced during reduction of the pyrite catalyst to iron sulfide.

Estimation of trace amounts of benzene in solvent-extracted vegetable oils and oil seed cakes.

PubMed

Masohan, A; Parsad, G; Khanna, M K; Chopra, S K; Rawat, B S; Garg, M O

2000-09-01

A new method is presented for the qualitative and quantitative estimation of trace amounts (up to 0.15 ppm) of benzene in crude as well as refined vegetable oils obtained by extraction with food grade hexane (FGH), and in the oil seed cakes left after extraction. The method involves the selection of two solvents; cyclohexanol, for thinning of viscous vegetable oil, and heptane, for azeotroping out trace benzene as a concentrate from the resulting mixture. Benzene is then estimated in the resulting azeotrope either by UV spectroscopy or by GC-MS subject to availability and cost effectiveness of the latter. Repeatability and reproducibility of the method is within 1-3% error. This method is suitable for estimating benzene in vegetable oils and oil seed cakes.

Magnesium Electrorefining in Non-Aqueous Electrolyte at Room Temperature

NASA Astrophysics Data System (ADS)

Kwon, Kyungjung; Park, Jesik; Kusumah, Priyandi; Dilasari, Bonita; Kim, Hansu; Lee, Churl Kyoung

Magnesium, of which application is often limited by its poor corrosion resistance, is more vulnerable to corrosion with existence of metal impurities such as Fe. Therefore, for the refining and recycling of magnesium, high temperature electrolysis using molten salts has been frequently adopted. In this report, the purification of magnesium scrap by electrolysis at room temperature is investigated with non-aqueous electrolytes. An aprotic solvent of tetrahydrofuran (THF) was used as a solvent of the electrolyte. Magnesium scrap was used as anode materials and ethyl magnesium bromide (EtMgBr) was dissolved in THF for magnesium source. The purified magnesium can be uniformly electrodeposited on copper electrode under potentiostatic conditions. The deposits were confirmed by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS) analysis.

Src Kinase: A Novel Target of Early-Stage ER-Negative Breast Cancer

DTIC Science & Technology

2012-03-01

patients with early stage ErbB2-overexpressing biopsies and ER- atypia . 13 REFERENCES: 1. Jordan VC. Tamoxifen for breast cancer prevention. Proc Soc...Summary01-03-2012 Src Kinase: A Novel Target of Early-Stage ER-Negative Breast Cancer Shalini Jain University of Texas M.D. Anderson Cancer Center Houston...SUBTITLE “Src Kinase: A Novel Target of Early-Stage ER-Negative Breast Cancer” 5a. CONTRACT NUMBER W81XWH-11-1-0004 5b. GRANT NUMBER

Recent Developments in the Classification, Evaluation, Pathophysiology, and Management of Scleroderma Renal Crisis.

PubMed

Ghossein, Cybele; Varga, John; Fenves, Andrew Z

2016-01-01

Scleroderma renal crisis (SRC) is an uncommon complication of systemic sclerosis. Despite the advent of angiotensin-converting inhibitor therapy, SRC remains a life-threatening complication. Recent studies have contributed to a better understanding of SRC, but much remains unknown regarding its pathophysiology, risk factors, and optimal management. Genetic studies provide evidence that immune dysregulation might be a contributing factor, providing hope that further research in this direction might illuminate pathogenesis and provide novel predictors for this complication.

Vestibulo-ocular dysfunction in pediatric sports-related concussion.

PubMed

Ellis, Michael J; Cordingley, Dean; Vis, Sara; Reimer, Karen; Leiter, Jeff; Russell, Kelly

2015-09-01

The objective of this study was 2-fold: 1) to examine the prevalence of vestibulo-ocular dysfunction (VOD) among children and adolescents with acute sports-related concussion (SRC) and postconcussion syndrome (PCS) who were referred to a multidisciplinary pediatric concussion program; and 2) to determine if VOD is associated with the development of PCS in this cohort. The authors conducted a retrospective review of all patients with acute SRC (presenting 30 days or less postinjury) and PCS (3 or more symptoms for at least 1 month) referred to a multidisciplinary pediatric concussion program between September 2013 and July 2014. Initial assessment included clinical history, physical examination, and Post-Concussion Symptom Scale assessment. Patients were also assessed for VOD, which was defined as more than one subjective vestibular and oculomotor complaint (dizziness, blurred vision, and so on) and more than one objective physical examination finding (abnormal smooth pursuits, saccades, vestibulo-ocular reflex, and so on). This study was approved by the local institutional ethics review board. A total of 101 patients (mean age 14.2 years, SD 2.3 years; 63 male and 38 female patients) participated, including 77 (76.2%) with acute SRC and 24 (23.8%) with PCS. Twenty-two of the 77 patients (28.6%) with acute SRC and 15 of the 24 (62.5%) with PCS met the clinical criteria for VOD. The median duration of symptoms was 40 days (interquartile range [IQR] 28.5-54 days) for patients with acute SRC who had VOD compared with 21 days (IQR 13-32 days) for those without VOD (p = 0.0001). There was a statistically significant increase in the adjusted odds of developing PCS among patients with acute SRC who had VOD compared with those without VOD (adjusted OR 4.10; 95% CI 1.04-16.16). Evidence of VOD was detected in a significant proportion of children and adolescents with acute SRC and PCS who were referred to a multidisciplinary pediatric concussion program. This clinical feature was a significant risk factor for the subsequent development of PCS in this pediatric acute SRC cohort.

Acute Sport-Related Concussion Screening for Collegiate Athletes Using an Instrumented Balance Assessment.

PubMed

Baracks, Joshua; Casa, Douglas J; Covassin, Tracey; Sacko, Ryan; Scarneo, Samantha E; Schnyer, David; Yeargin, Susan W; Neville, Christopher

2018-06-13

  Without a true criterion standard assessment, the sport-related concussion (SRC) diagnosis remains subjective. Inertial balance sensors have been proposed to improve acute SRC assessment, but few researchers have studied their clinical utility.   To determine if group differences exist when using objective measures of balance in a sample of collegiate athletes with recent SRCs and participants serving as the control group and to calculate sensitivity and specificity to determine the diagnostic utility of the inertial balance sensor for acute SRC injuries.   Cohort study.   Multicenter clinical trial.   We enrolled 48 participants with SRC (age = 20.62 ± 1.52 years, height = 179.76 ± 10.00 cm, mass = 83.92 ± 23.22 kg) and 45 control participants (age = 20.85 ± 1.42 years, height = 177.02 ± 9.59 cm, mass = 74.61 ± 14.92 kg) at 7 clinical sites in the United States. All were varsity or club collegiate athletes, and all participants with SRC were tested within 72 hours of SRC.   Balance performance was assessed using an inertial balance sensor. Two measures (root mean square [RMS] sway and 95% ellipse sway area) were analyzed to represent a range of general balance measures. Balance assessments were conducted in double-legged, single-legged, and tandem stances.   A main effect for group was associated with the root mean square sway measure ( F 1,91 = 11.75, P = .001), with the SRC group demonstrating balance deficits compared with the control group. We observed group differences in the 95% ellipse sway area measure for the double-legged ( F 1,91 = 11.59, P = .001), single-legged ( F 1,91 = 6.91, P = .01), and tandem ( F 1,91 = 7.54, P = .007) stances. Sensitivity was greatest using a cutoff value of 0.5 standard deviations (54% [specificity = 71%]), whereas specificity was greatest using a cutoff value of 2 standard deviations (98% [sensitivity = 33%]).   Inertial balance sensors may be useful tools for objectively measuring balance during acute SRC evaluation. However, low sensitivity suggests that they may be best used in conjunction with other assessments to form a comprehensive screening that may improve sensitivity.

Ableson Kinases Negatively Regulate Invadopodia Function and Invasion in Head and Neck Squamous Cell Carcinoma by Inhibiting an HB-EGF Autocrine Loop

PubMed Central

Hayes, Karen E.; Walk, Elyse L.; Ammer, Amanda Gatesman; Kelley, Laura C.; Martin, Karen H.; Weed, Scott A.

2014-01-01

Head and neck squamous cell carcinoma (HNSCC) has a proclivity for locoregional invasion. HNSCC mediates invasion in part through invadopodia-based proteolysis of the extracellular matrix (ECM). Activation of Src, Erk1/2, Abl and Arg downstream of epidermal growth factor receptor (EGFR) modulates invadopodia activity through phosphorylation of the actin regulatory protein cortactin. In MDA-MB-231 breast cancer cells, Abl and Arg function downstream of Src to phosphorylate cortactin, promoting invadopodia ECM degradation activity and thus assigning a pro-invasive role for Ableson kinases. We report that Abl kinases have an opposite, negative regulatory role in HNSCC where they suppress invadopodia and tumor invasion. Impairment of Abl expression or Abl kinase activity with imatinib mesylate enhanced HNSCC matrix degradation and 3D collagen invasion, functions that were impaired in MDA-MB-231. HNSCC lines with elevated EGFR and Src activation did not contain increased Abl or Arg kinase activity, suggesting Src could bypass Abl/Arg to phosphorylate cortactin and promote invadopodia ECM degradation. Src transformed Abl−/−/Arg−/− fibroblasts produced ECM degrading invadopodia containing pY421 cortactin, indicating that Abl/Arg are dispensable for invadopodia function in this system. Imatinib treated HNSCC cells had increased EGFR, Erk1/2 and Src activation, enhancing cortactin pY421 and pS405/418 required for invadopodia function. Imatinib stimulated shedding of the EGFR ligand heparin-binding EGF-like growth factor (HB-EGF) from HNSCC cells, where soluble HB-EGF enhanced invadopodia ECM degradation in HNSCC but not in MDA-MB-231. HNSCC cells treated with inhibitors of the EGFR invadopodia pathway indicated that EGFR and Src are required for invadopodia function. Collectively our results indicate that Abl kinases negatively regulate HNSCC invasive processes through suppression of an HB-EGF autocrine loop responsible for activating a EGFR-Src-cortactin cascade, in contrast to the invasion promoting functions of Abl kinases in breast and other cancer types. Our results provide mechanistic support for recent failed HNSCC clinical trials utilizing imatinib. PMID:23146907

Properties and Corrosion Performance of Self-reinforced Composite PEEK for Proposed Use as a Modular Taper Gasket.

PubMed

Ouellette, Eric S; Gilbert, Jeremy L

2016-11-01

Fretting corrosion in medical alloys is a persistent problem, and the need for biomaterials that can effectively suppress mechanically assisted crevice corrosion in modular taper junctions or otherwise insulate metal-on-metal interfaces in mechanically demanding environments is as yet unmet. The purpose of this study is to characterize a novel material, self-reinforced composite polyetheretherketone (SRC-PEEK) and to evaluate its ability to inhibit fretting corrosion in a pin-on-disk metal-on-metal interface test. SRC-PEEK was fabricated by hot compaction of in-house-made PEEK fibers by compacting uniaxial layups at 344°C under a load of 18,000 N for 10 minutes. SRC-PEEK, bulk isotropic PEEK, and the in-house-made PEEK fibers were analyzed for thermal transitions (T g , T m ) through differential scanning calorimetry, crystallinity, crystal size, crystalline orientation (Hermanns orientation parameter) through wide-angle x-ray scattering, and modulus, tensile strength, yield stress, and strain to failure through monotonic tensile testing. SRC-insulated pin-on-disk samples were compared with metal-on-metal control samples in pin-on-disk fretting corrosion experiments using fretting current and fretting mechanics measurements. Fifty-micron cyclic motion at 2.5 Hz was applied to the interface, first over a range of loads (0.5-35 N) while held at -0.05 V versus Ag/AgCl and then over a range of voltages (-0.5 to 0.5 V) at a constant contact stress of 73 ± 19 MPa for SRC-PEEK and 209 ± 41 MPa for metal-on-metal, which were different for each group as a result of changes in true contact area due to variations in modulus between sample groups. Pins, disks, and SRC samples were imaged for damage (on alloy and SRC surfaces) and evidence of corrosion (on alloy pin and disk surfaces). SRC specimens were analyzed for traces of alloy transferred to the surface using energy dispersive spectroscopy after pin-on-disk testing. SRC-PEEK showed improved mechanical properties to bulk PEEK (modulus = 5.0 ± 0.3 GPa, 2.8 ± 0.1 GPa, respectively, p < 0.001) and higher crystallinity to bulk PEEK (44.2% ± 3%, 39.5% ± 0.5%, respectively, p = 0.039), but had comparable crystalline orientation as compared with the initial PEEK fibers. SRC-PEEK reduced fretting currents compared with metal-on-metal controls by two to three orders of magnitude in both variable load (4.0E-5 ± 3.8E-5 μA versus 2.9E-3 ± 7.1E-4 μA, respectively, p = 0.018) and variable potential (7.5E-6 ± 4.7E-6 μA versus 5.3E-3 ± 1.4E-3 μA, respectively, p = 0.022) fretting corrosion testing. Minimal damage was observed on surfaces insulated with SRC-PEEK, whereas control surfaces showed considerable fretting corrosion damage and metal transfer. The SRC-PEEK gaskets in this study demonstrated higher crystallinity and crystalline orientation and improved monotonic tensile properties compared with bulk PEEK with the ability to effectively insulate Ti6Al4V and CoCrMo alloy surfaces and prevent the initiation of fretting corrosion under high contact-stress conditions. This novel SRC-PEEK material may offer potential as a thin film gasket material for modular tapers. Pending further in vitro and in vivo analyses, this approach may be able to preserve the advantages of modular junctions for surgeons while potentially limiting the downside risks associated with mechanically assisted crevice corrosion.

Three-dimensional quantitative structure-activity relationship studies on c-Src inhibitors based on different docking methods.

PubMed

Bairy, Santhosh Kumar; Suneel Kumar, B V S; Bhalla, Joseph Uday Tej; Pramod, A B; Ravikumar, Muttineni

2009-04-01

c-Src kinase play an important role in cell growth and differentiation and its inhibitors can be useful for the treatment of various diseases, including cancer, osteoporosis, and metastatic bone disease. Three dimensional quantitative structure-activity relationship (3D-QSAR) studies were carried out on quinazolin derivatives inhibiting c-Src kinase. Molecular field analysis (MFA) models with four different alignment techniques, namely, GLIDE, GOLD, LIGANDFIT and Least squares based methods were developed. glide based MFA model showed better results (Leave one out cross validation correlation coefficient r(2)(cv) = 0.923 and non-cross validation correlation coefficient r(2)= 0.958) when compared with other models. These results help us to understand the nature of descriptors required for activity of these compounds and thereby provide guidelines to design novel and potent c-Src kinase inhibitors.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhengfu, He; Hu, Zhang; Huiwen, Miao

The search for safe, effective and affordable therapeutics against non-small cell lung cancer (NSCLC) and other lung cancers is important. Here we explored the potential effect of 1-o-acetylbritannilactone (ABL), a novel extract from Inula britannica-F, on angiogenesis and lung cancer cell growth. We demonstrated that ABL dose-dependently inhibited vascular endothelial growth factor (VEGF)-induced proliferation, migration, and capillary structure formation of cultured human umbilical vascular endothelial cells (HUVECs). In vivo, ABL administration suppressed VEGF-induced new vasculature formation in Matrigel plugs. For the mechanism investigations, we found that ABL largely inhibited VEGF-mediated activation of Src kinase and focal adhesion kinase (FAK) in HUVECs.more » Furthermore, treatment of A549 NSCLC cells with ABL resulted in cell growth inhibition and Src-FAK in-activation. Significantly, administration of a single dose of ABL (12 mg/kg/day) remarkably suppressed growth of A549 xenografts in nude mice. In vivo microvessels formation and Src activation were also significantly inhibited in ABL-treated xenograft tumors. Taken together, our findings suggest that ABL suppresses angiogenesis and lung cancer cell growth possibly via regulating the VEGFR-Src-FAK signaling. - Highlights: • 1-o-acetylbritannilactone (ABL) inhibits VEGF-induced angiogenesis in vivo. • ABL inhibits VEGF-induced HUVEC migration, proliferation, capillary tube formation. • ABL inhibits VEGF-mediated activation of Src and FAK in HUVECs. • ABL inhibits growth and Src-FAK activation in A549 cells. • ABL administration inhibits A549 tumor angiogenesis and growth in nude mice.« less

Ror2-Src signaling in metastasis of mouse melanoma cells is inhibited by NRAGE.

PubMed

Lai, Shan-Shan; Xue, Bin; Yang, Yang; Zhao, Li; Chu, Chao-Shun; Hao, Jia-Yin; Wen, Chuan-Jun

2012-11-01

The receptor tyrosine kinase (RTK) Ror2 plays important roles in developmental morphogenesis and mediates the filopodia formation in Wnt5a-induced cell migration. However, the function of Ror2 in noncanonical Wnt signaling resulting in cancer metastasis is largely unknown. Here, we show that Ror2 expression is higher in the highly metastatic murine B16-BL6 melanoma cells than in the low metastatic variant B16 cells. Overexpression of Ror2 increases the metastasis ability of B16 cells, and knockdown of Ror2 reduces the migration ability of B16-BL6 cells. Furthermore, the inhibition of Src kinase activity is critical for the Ror2-mediated cell migration upon Wnt5a treatment. The C-terminus of Ror2, which is deleted in brachydactyly type B (BDB), is essential for the mutual interaction with the SH1 domain of Src. Intriguingly, the Neurotrophin receptor-interacting MAGE homologue (NRAGE), which, as we previously reported, can remodel the cellular skeleton and inhibit cell-cell adhesion and metastasis of melanoma and pancreatic cancer, sharply blocks the interaction between Src and Ror2 and inhibits Ror2-mediated B16 cell migration by decreasing the activity of Src and focal adhesion kinase (FAK). Our data show that Ror2 is a potential factor in the tumorigenesis and metastasis in a Src-dependent manner that is negatively regulated by NRAGE. Copyright © 2012. Published by Elsevier Inc.

Optimal Couple Projections for Domain Adaptive Sparse Representation-based Classification.

PubMed

Zhang, Guoqing; Sun, Huaijiang; Porikli, Fatih; Liu, Yazhou; Sun, Quansen

2017-08-29

In recent years, sparse representation based classification (SRC) is one of the most successful methods and has been shown impressive performance in various classification tasks. However, when the training data has a different distribution than the testing data, the learned sparse representation may not be optimal, and the performance of SRC will be degraded significantly. To address this problem, in this paper, we propose an optimal couple projections for domain-adaptive sparse representation-based classification (OCPD-SRC) method, in which the discriminative features of data in the two domains are simultaneously learned with the dictionary that can succinctly represent the training and testing data in the projected space. OCPD-SRC is designed based on the decision rule of SRC, with the objective to learn coupled projection matrices and a common discriminative dictionary such that the between-class sparse reconstruction residuals of data from both domains are maximized, and the within-class sparse reconstruction residuals of data are minimized in the projected low-dimensional space. Thus, the resulting representations can well fit SRC and simultaneously have a better discriminant ability. In addition, our method can be easily extended to multiple domains and can be kernelized to deal with the nonlinear structure of data. The optimal solution for the proposed method can be efficiently obtained following the alternative optimization method. Extensive experimental results on a series of benchmark databases show that our method is better or comparable to many state-of-the-art methods.

Steroid receptor coactivators, HER-2 and HER-3 expression is stimulated by tamoxifen treatment in DMBA-induced breast cancer.

PubMed

Moi, Line L Haugan; Flågeng, Marianne Hauglid; Gjerde, Jennifer; Madsen, Andre; Røst, Therese Halvorsen; Gudbrandsen, Oddrun Anita; Lien, Ernst A; Mellgren, Gunnar

2012-06-15

Steroid receptor coactivators (SRCs) may modulate estrogen receptor (ER) activity and the response to endocrine treatment in breast cancer, in part through interaction with growth factor receptor signaling pathways. In the present study the effects of tamoxifen treatment on the expression of SRCs and human epidermal growth factor receptors (HERs) were examined in an animal model of ER positive breast cancer. Sprague-Dawley rats with DMBA-induced breast cancer were randomized to 14 days of oral tamoxifen 40 mg/kg bodyweight/day or vehicle only (controls). Tumors were measured throughout the study period. Blood samples and tumor tissue were collected at sacrifice and tamoxifen and its main metabolites were quantified using LC-MS/MS. The gene expression in tumor of SRC-1, SRC-2/transcription intermediary factor-2 (TIF-2), SRC-3/amplified in breast cancer 1 (AIB1), ER, HER-1, -2, -3 and HER-4, as well as the transcription factor Ets-2, was measured by real-time RT-PCR. Protein levels were further assessed by Western blotting. Tamoxifen and its main metabolites were detected at high concentrations in serum and accumulated in tumor tissue in up to tenfolds the concentration in serum. Mean tumor volume/rat decreased in the tamoxifen treated group, but continued to increase in controls. The mRNA expression levels of SRC-1 (P = 0.035), SRC-2/TIF-2 (P = 0.002), HER-2 (P = 0.035) and HER-3 (P = 0.006) were significantly higher in tamoxifen treated tumors compared to controls, and the results were confirmed at the protein level using Western blotting. SRC-3/AIB1 protein was also higher in tamoxifen treated tumors. SRC-1 and SRC-2/TIF-2 mRNA levels were positively correlated with each other and with HER-2 (P ≤ 0.001), and the HER-2 mRNA expression correlated with the levels of the other three HER family members (P < 0.05). Furthermore, SRC-3/AIB1 and HER-4 were positively correlated with each other and Ets-2 (P < 0.001). The expression of SRCs and HER-2 and -3 is stimulated by tamoxifen treatment in DMBA-induced breast cancer. Stimulation and positive correlation of coactivators and HERs may represent an early response to endocrine treatment. The role of SRCs and HER-2 and -3 should be further studied in order to evaluate their effects on response to long-term tamoxifen treatment.

Steroid receptor coactivators, HER-2 and HER-3 expression is stimulated by tamoxifen treatment in DMBA-induced breast cancer

PubMed Central

2012-01-01

Background Steroid receptor coactivators (SRCs) may modulate estrogen receptor (ER) activity and the response to endocrine treatment in breast cancer, in part through interaction with growth factor receptor signaling pathways. In the present study the effects of tamoxifen treatment on the expression of SRCs and human epidermal growth factor receptors (HERs) were examined in an animal model of ER positive breast cancer. Methods Sprague-Dawley rats with DMBA-induced breast cancer were randomized to 14 days of oral tamoxifen 40 mg/kg bodyweight/day or vehicle only (controls). Tumors were measured throughout the study period. Blood samples and tumor tissue were collected at sacrifice and tamoxifen and its main metabolites were quantified using LC-MS/MS. The gene expression in tumor of SRC-1, SRC-2/transcription intermediary factor-2 (TIF-2), SRC-3/amplified in breast cancer 1 (AIB1), ER, HER-1, -2, -3 and HER-4, as well as the transcription factor Ets-2, was measured by real-time RT-PCR. Protein levels were further assessed by Western blotting. Results Tamoxifen and its main metabolites were detected at high concentrations in serum and accumulated in tumor tissue in up to tenfolds the concentration in serum. Mean tumor volume/rat decreased in the tamoxifen treated group, but continued to increase in controls. The mRNA expression levels of SRC-1 (P = 0.035), SRC-2/TIF-2 (P = 0.002), HER-2 (P = 0.035) and HER-3 (P = 0.006) were significantly higher in tamoxifen treated tumors compared to controls, and the results were confirmed at the protein level using Western blotting. SRC-3/AIB1 protein was also higher in tamoxifen treated tumors. SRC-1 and SRC-2/TIF-2 mRNA levels were positively correlated with each other and with HER-2 (P ≤ 0.001), and the HER-2 mRNA expression correlated with the levels of the other three HER family members (P < 0.05). Furthermore, SRC-3/AIB1 and HER-4 were positively correlated with each other and Ets-2 (P < 0.001). Conclusions The expression of SRCs and HER-2 and -3 is stimulated by tamoxifen treatment in DMBA-induced breast cancer. Stimulation and positive correlation of coactivators and HERs may represent an early response to endocrine treatment. The role of SRCs and HER-2 and -3 should be further studied in order to evaluate their effects on response to long-term tamoxifen treatment. PMID:22703232

Role of STATs as downstream signal transducers in Src family kinase-mediated tumorigenesis.

PubMed

Silva, Corinne M

2004-10-18

The signal transducers and activators of transcription (STATs) were originally identified in the signaling pathway activated by the nontyrosine kinase containing cytokine receptors. The role of these STATs in hematopoietic cell signaling has been well described. In the case of cytokine receptors, activation of STAT tyrosine phosphorylation occurs through ligand-induced recruitment, and activation of the intracellular JAK kinases. However, STATs can also be activated by growth factor receptors, particularly the EGFR; as well as by members of the Src Family of Kinases (SFKs), particularly c-Src. In many cases, there is a differential activation of the STATs by these tyrosine kinases as compared to activation by the cytokine receptors. This difference provides for the potential of unique actions of STATs in response to growth factor receptor and SFK activation. Since there are many cancers in which SFKs and c-Src in particular, are co-overexpressed with growth factor receptors, it is not surprising that STATs play an important role in the tumorigenesis process induced by c-Src. The activation paradigm and role of STATs in these cancers, with particular emphasis on breast cancer models, is discussed.

Nck adaptor proteins link Tks5 to invadopodia actin regulation and ECM degradation.

PubMed

Stylli, Stanley S; Stacey, T T I; Verhagen, Anne M; Xu, San San; Pass, Ian; Courtneidge, Sara A; Lock, Peter

2009-08-01

Invadopodia are actin-based projections enriched with proteases, which invasive cancer cells use to degrade the extracellular matrix (ECM). The Phox homology (PX)-Src homology (SH)3 domain adaptor protein Tks5 (also known as SH3PXD2A) cooperates with Src tyrosine kinase to promote invadopodia formation but the underlying pathway is not clear. Here we show that Src phosphorylates Tks5 at Y557, inducing it to associate directly with the SH3-SH2 domain adaptor proteins Nck1 and Nck2 in invadopodia. Tks5 mutants unable to bind Nck show reduced matrix degradation-promoting activity and recruit actin to invadopodia inefficiently. Conversely, Src- and Tks5-driven matrix proteolysis and actin assembly in invadopodia are enhanced by Nck1 or Nck2 overexpression and inhibited by Nck1 depletion. We show that clustering at the plasma membrane of the Tks5 inter-SH3 region containing Y557 triggers phosphorylation at this site, facilitating Nck recruitment and F-actin assembly. These results identify a Src-Tks5-Nck pathway in ECM-degrading invadopodia that shows parallels with pathways linking several mammalian and pathogen-derived proteins to local actin regulation.

Activated d16HER2 homodimers and SRC kinase mediate optimal efficacy for trastuzumab.

PubMed

Castagnoli, Lorenzo; Iezzi, Manuela; Ghedini, Gaia C; Ciravolo, Valentina; Marzano, Giulia; Lamolinara, Alessia; Zappasodi, Roberta; Gasparini, Patrizia; Campiglio, Manuela; Amici, Augusto; Chiodoni, Claudia; Palladini, Arianna; Lollini, Pier Luigi; Triulzi, Tiziana; Menard, Sylvie; Nanni, Patrizia; Tagliabue, Elda; Pupa, Serenella M

2014-11-01

A splice isoform of the HER2 receptor that lacks exon 16 (d16HER2) is expressed in many HER2-positive breast tumors, where it has been linked with resistance to the HER2-targeting antibody trastuzumab, but the impact of d16HER2 on tumor pathobiology and therapeutic response remains uncertain. Here, we provide genetic evidence in transgenic mice that expression of d16HER2 is sufficient to accelerate mammary tumorigenesis and improve the response to trastuzumab. A comparative analysis of effector signaling pathways activated by d16HER2 and wild-type HER2 revealed that d16HER2 was optimally functional through a link to SRC activation (pSRC). Clinically, HER2-positive breast cancers from patients who received trastuzumab exhibited a positive correlation in d16HER2 and pSRC abundance, consistent with the mouse genetic results. Moreover, patients expressing high pSRC or an activated "d16HER2 metagene" were found to derive the greatest benefit from trastuzumab treatment. Overall, our results establish the d16HER2 signaling axis as a signature for decreased risk of relapse after trastuzumab treatment. ©2014 American Association for Cancer Research.

The targeted delivery of the c-Src peptide complexed with schizophyllan to macrophages inhibits polymicrobial sepsis and ulcerative colitis in mice.

PubMed

Kim, Ye-Ram; Hwang, Jangsun; Koh, Hyun-Jung; Jang, Kiseok; Lee, Jong-Dae; Choi, Jonghoon; Yang, Chul-Su

2016-05-01

Hyper-inflammatory responses triggered by intracellular reactive oxygen species (ROS) can lead to a variety of diseases, including sepsis and colitis. However, the regulators of this process remain poorly defined. In this study, we demonstrate that c-Src is a negative regulator of cellular ROS generation through its binding to p47phox. This molecule also competitively inhibits the NADPH oxidase complex (NOX) assembly. Furthermore, we developed the schizophyllan (SPG)-c-Src SH3 peptide, which is a β-1,3-glucan conjugated c-Src SH3-derived peptide composed of amino acids 91-108 and 121-140 of c-Src. The SPG-SH3 peptide has a significant therapeutic effect on mouse ROS-mediated inflammatory disease models, cecal-ligation-puncture-induced sepsis, and dextran sodium sulfate-induced colitis. It does so by inhibiting the NOX subunit assembly and proinflammatory mediator production. Therefore, the SPG-SH3 peptide is a potential therapeutic agent for ROS-associated lethal inflammatory diseases. Our findings provide clues for the development of new peptide-base drugs that will target p47phox. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mapping Heart Development in Flies: Src42A Acts Non-Autonomously to Promote Heart Tube Formation in Drosophila

PubMed Central

Vanderploeg, Jessica; Jacobs, J. Roger

2017-01-01

Congenital heart defects, clinically identified in both small and large animals, are multifactorial and complex. Although heritable factors are known to have a role in cardiovascular disease, the full genetic aetiology remains unclear. Model organism research has proven valuable in providing a deeper understanding of the essential factors in heart development. For example, mouse knock-out studies reveal a role for the Integrin adhesion receptor in cardiac tissue. Recent research in Drosophila melanogaster (the fruit fly), a powerful experimental model, has demonstrated that the link between the extracellular matrix and the cell, mediated by Integrins, is required for multiple aspects of cardiogenesis. Here we test the hypothesis that Integrins signal to the heart cells through Src42A kinase. Using the powerful genetics and cell biology analysis possible in Drosophila, we demonstrate that Src42A acts in early events of heart tube development. Careful examination of mutant heart tissue and genetic interaction data suggests that Src42A’s role is independent of Integrin and the Integrin-related Focal Adhesion Kinase. Rather, Src42A acts non-autonomously by promoting programmed cell death of the amnioserosa, a transient tissue that neighbors the developing heart. PMID:29056682

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Chunfa, E-mail: chunfa.huang@case.edu; Department of Medicine, Case Western Reserve University; Rammelkamp Center for Research and Education, MetroHealth System Campus, Cleveland, OH 44106

The glomerular capillary wall, composed of endothelial cells, the glomerular basement membrane and the podocytes, is continually subjected to hemodynamic force arising from tractional stress due to blood pressure and shear stress due to blood flow. Exposure of glomeruli to abnormal hemodynamic force such as hyperfiltration is associated with glomerular injury and progressive renal disease, and the conversion of mechanical stimuli to chemical signals in the regulation of the process is poorly understood in podocytes. By examining DNA fragmentation, apoptotic nuclear changes and cytochrome c release, we found that shear stress induced cell apoptosis in cultured podocytes. Meanwhile, podocytes exposedmore » to shear stress also stimulated c-Src phosphorylation, phospholipase D (PLD) activation and mammalian target of rapamycin (mTOR) signaling. Using the antibodies against c-Src, PLD{sub 1}, and PLD{sub 2} to perform reciprocal co-immunoprecipitations and in vitro PLD activity assay, our data indicated that c-Src interacted with and activated PLD{sub 1} but not PLD{sub 2}. The inhibition of shear stress-induced c-Src phosphorylation by PP{sub 2} (a specific inhibitor of c-Src kinase) resulted in reduced PLD activity. Phosphatidic acid, produced by shear stress-induced PLD activation, stimulated mTOR signaling, and caused podocyte hypertrophy and apoptosis.« less

Potential of mean force for ion pairs in non-aqueous solvents. Comparison of polarizable and non-polarizable MD simulations

NASA Astrophysics Data System (ADS)

Odinokov, A. V.; Leontyev, I. V.; Basilevsky, M. V.; Petrov, N. Ch.

2011-01-01

Potentials of mean force (PMF) are calculated for two model ion pairs in two non-aqueous solvents. Standard non-polarizable molecular dynamics simulation (NPMD) and approximate polarizable simulation (PMD) are implemented and compared as tools for monitoring PMF profiles. For the polar solvent (dimethylsulfoxide, DMSO) the PMF generated in terms of the NPMD reproduces fairly well the refined PMD-PMF profile. For the non-polar solvent (benzene) the conventional NPMD computation proves to be deficient. The validity of the correction found in terms of the approximate PMD approach is verified by its comparison with the result of the explicit PMD computation in benzene. The shapes of the PMF profiles in DMSO and in benzene are quite different. In DMSO, owing to dielectric screening, the PMF presents a flat plot with a shallow minimum positioned in the vicinity of the van der Waals contact of the ion pair. For the benzene case, the observed minimum proves to be unexpectedly deep, which manifests the formation of a tightly-binded contact ion pair. This remarkable effect arises owing to the strong electrostatic interaction that is incompletely screened by a non-polar medium. The PMFs for the binary benzene/DMSO mixtures display intermediate behaviour depending on the DMSO content.

77 FR 4580 - Alaska Region's Subsistence Resource Commission (SRC) Program

Federal Register 2010, 2011, 2012, 2013, 2014

2012-01-30

... canceled due to a lack of quorum caused by inclement Arctic weather conditions. The NPS has rescheduled... weather or exceptional circumstances. Kobuk Valley National Park SRC Meeting Date and Location: The Kobuk...

The Role of C-SRC Activation in Prostate Tumor Progression

DTIC Science & Technology

2006-07-01

cancer cell line PANC -1 and prostrate cancer cell line PC-3 (B2-fold increase relative to control in both cell lines), while the Src inhibitory PP2 blocks...at normoxia in PANC -1 and PC-3 cells, its levels significantly increase in response to hypoxia (B4.5–8-fold induction). Inhibition of endo- genous c...Src activation in PANC -1 and PC-3 cells by PP2 drastically reduced HIF-1a levels to below those levels observed at normoxia (Figure 1a). STAT3 has

An Efficient, Lossless Database for Storing and Transmitting Medical Images

NASA Technical Reports Server (NTRS)

Fenstermacher, Marc J.

1998-01-01

This research aimed in creating new compression methods based on the central idea of Set Redundancy Compression (SRC). Set Redundancy refers to the common information that exists in a set of similar images. SRC compression methods take advantage of this common information and can achieve improved compression of similar images by reducing their Set Redundancy. The current research resulted in the development of three new lossless SRC compression methods: MARS (Median-Aided Region Sorting), MAZE (Max-Aided Zero Elimination) and MaxGBA (Max-Guided Bit Allocation).

Molecular-Scale Investigation of Heavy Metal Ions at a Charged Langmuir Monolayer

NASA Astrophysics Data System (ADS)

Rock, William; Qiao, Baofu; Uysal, Ahmet; Bu, Wei; Lin, Binhua

Solvent extraction - the surfactant-aided preferential transfer of a species from an aqueous to an organic phase - is an important technique used in heavy and precious metal refining and reprocessing. Solvent extraction requires transfer through an oil/water interface, and interfacial interactions are expected to control transfer kinetics and phase stability, yet these key interactions are poorly understood. Langmuir monolayers with charged headgroups atop concentrated salt solutions containing heavy metal ions act as a model of solvent extraction interfaces; studies of ions at a charged surface are also fundamentally important to many other phenomena including protein solvation, mineral surface chemistry, and electrochemistry. We probe these charged interfaces using a variety of surface-sensitive techniques - vibrational sum frequency generation (VSFG) spectroscopy, x-ray reflectivity (XRR), x-ray fluorescence near total reflection (XFNTR), and grazing incidence diffraction (GID). We integrate experiments with Molecular Dynamics (MD) simulations to uncover the molecular-level interfacial structure. This work is supported by the U.S. DOE, BES, Contract DE-AC02-06CH11357. ChemMatCARS is supported by NSF/CHE-1346572.

Applications of supercritical fluid extraction (SFE) of palm oil and oil from natural sources.

PubMed

Akanda, Mohammed Jahurul Haque; Sarker, Mohammed Zaidul Islam; Ferdosh, Sahena; Manap, Mohd Yazid Abdul; Ab Rahman, Nik Norulaini Nik; Ab Kadir, Mohd Omar

2012-02-10

Supercritical fluid extraction (SFE), which has received much interest in its use and further development for industrial applications, is a method that offers some advantages over conventional methods, especially for the palm oil industry. SC-CO₂ refers to supercritical fluid extraction (SFE) that uses carbon dioxide (CO₂) as a solvent which is a nontoxic, inexpensive, nonflammable, and nonpolluting supercritical fluid solvent for the extraction of natural products. Almost 100% oil can be extracted and it is regarded as safe, with organic solvent-free extracts having superior organoleptic profiles. The palm oil industry is one of the major industries in Malaysia that provides a major contribution to the national income. Malaysia is the second largest palm oil and palm kernel oil producer in the World. This paper reviews advances in applications of supercritical carbon dioxide (SC-CO₂) extraction of oils from natural sources, in particular palm oil, minor constituents in palm oil, producing fractionated, refined, bleached, and deodorized palm oil, palm kernel oil and purified fatty acid fractions commendable for downstream uses as in toiletries and confectionaries.

Localization of caveolin-1 and c-src in mature and differentiating photoreceptors: raft proteins co-distribute with rhodopsin during development

PubMed Central

Berta, Ágnes I.; Boesze-Battaglia, Kathleen; Magyar, Attila; Szél, Ágoston; Kiss, Anna L.

2014-01-01

Numerous biochemical and morphological studies have provided insight into the distribution pattern of caveolin-1 and the presence of membrane rafts in the vertebrate retina. To date however, studies have not addressed the localization profile of raft specific proteins during development. Therefore the purpose of our studies was to follow the localization pattern of caveolin-1, phosphocaveolin-1 and c-src in the developing retina and compare it to that observed in adults. Specific antibodies were used to visualize the distribution of caveolin-1, c-src, a kinase phosphorylating caveolin-1, and phospho-caveolin-1. The labeling pattern of this scaffolded complex was compared to those of rhodopsin and rhodopsin kinase. Samples were analyzed at various time points during postnatal development and compared to adult retinas. The immunocytochemical studies were complemented with immunoblots and immunoprecipitation studies. In the mature retina caveolin-1 and c-src localized mainly to the cell body and IS of photoreceptors, with only very weakly labeled OS. In contrast, phospho-caveolin-1 was only detectable in the OS of photoreceptors. During development we followed the expression and distribution profile of these proteins in a temporal sequence with special attention to the period when OS formation is most robust. Double labeling immunocytochemistry and immunoprecipitation showed rhodopsin to colocalize and co-immunoprecipitate with caveolin-1 and c-src. Individual punctate structures between the outer limiting membrane and the outer plexiform layer were seen at P10 to be labeled by both rhodopsin and caveolin-1 as well as by rhodopsin and c-src, respectively. These studies suggest that membrane raft specific proteins are co-distributed during development, thereby pointing to a role for such complexes in OS formation. In addition, the presence of small punctate structures containing caveolin-1, c-src and rhodopsin raise the possibility that these proteins may transport together to OS during development and that caveolin-1 exists predominantly in a phosphorylated form in the OS. PMID:21938483

Trends in serum relaxin concentration among elite collegiate female athletes

PubMed Central

Dragoo, Jason L; Castillo, Tiffany N; Korotkova, Tatiana A; Kennedy, Ashleigh C; Kim, Hyeon Joo; Stewart, Dennis R

2011-01-01

Purpose: This study was designed to investigate the relationship between serum relaxin concentration (SRC) and menstrual history and hormonal contraceptive use among elite collegiate female athletes. Evaluation of SRC in athletes is necessary, because relaxin has been associated with increased knee joint laxity and decreased anterior cruciate ligament (ACL) strength in animal models. Methods: National Collegiate Athletic Association Division I female athletes participating in sports at high risk for ACL tears – basketball, field hockey, gymnastics, lacrosse, soccer, and volleyball – were invited to participate. All participants completed a questionnaire about their menstrual history and hormonal contraceptive use. Venipuncture was performed to obtain samples of serum progesterone and relaxin. Samples were obtained during the mid-luteal phase from ovulating participants, and between the actual or projected cycle days 21 to 24, from anovulatory participants. Serum concentration of relaxin and progesterone was determined by ELISA and the data were analyzed using SPSS statistical software with significance set at P = 0.05. Results: 169 female athletes participated. The mean SRC among all participants was 3.08 ± 6.66 pg/mL). The mean SRC differed significantly between those participants using hormonal contraceptives (1.41 pg/mL) and those not using hormonal contraceptives (3.08 pg/mL, P = 0.002). Mean SRC was lowest among amenorrheic participants (1.02 pg/mL) and highest among oligomenorrheic participants (3.71 pg/mL) and eumenorrheic participants (3.06 pg/mL); these differences were not significant (P = 0.53). Mean serum progesterone concentration (SPC) differed significantly between those participants using hormonal contraceptives (2.80 ng/mL), and those not using hormonal contraceptives (6.99 ng/mL, P < 0.0001). Conclusions: There is a positive correlation between serum progesterone and SRC and an attenuation of SRC with hormonal contraceptive use. Our results underscore the significant role that hormonal contraceptives can play in decreasing relaxin levels, if future investigations establish a link between relaxin levels and ligamentous injury among female athletes. PMID:21339934

The Effect of Solar Reflective Cover on Soak Air Temperature and Thermal Comfort of Car Parked under the Sun

NASA Astrophysics Data System (ADS)

Lahimer, A. A.; Alghoul, M. A.; Sopian, K.; Khrit, N. G.

2017-11-01

Parking a vehicle under the sun for a short period of time can rapidly increase the interior air cabin temperature no matter in clear sky days or even in partially cloudy days. These circumstances can be anxieties to car occupants upon entry. The aim of this paper is to evaluate experimentally the effect of solar reflective cover (SRC) on vehicle air temperature and cabin thermal comfort. Experimental measurements of parked cars were conducted in UKM, Bangi city, Malaysia (latitude of 2.9° N and longitude of 101.78° E) under partially cloudy day where average ambient temperature is 33°C. The experimental measurements cover the following cases: case (I): car with/ without SRC (at different measurement time); Case (II): using two identical cars concurrently (SRC versus baseline); Case (III): using two identical cars concurrently (solar reflective film (SRF) versus baseline) and Case (IV): using two identical cars concurrently (SRF versus SRC). Experimental results dedicated to case (I) revealed that the maximum cabin air temperature with SRC (39.6°C) is significantly lower than that of baseline case (57.3°C). This leads to temperature reduction improvement of 31% and the difference between the cabin and the ambient air temperature was minimized by approximately 73%. In addition, the results revealed that the air temperature at breath level of car with SRC dropped to comfort temperature (27°C) after 7 min while baseline car reached comfort temperature after 14 min. Results of the other cases are discussed inside the paper. Overall, it is learned that SRC is found superior as an efficient thermal insulation system limits solar radiation transmission into the cabin through the glass; keeps cabin air temperature close to the ambient temperature; and provide acceptable thermal environment to the occupants as they settle into their parked car.

Genesis failure investigation report : JPL Failure Review Board, Avionics Sub-Team

NASA Technical Reports Server (NTRS)

Klein, John; Manning, Rob; Barry, Ed; Donaldson, Jim; Rivellini, Tom; Battel, Steven; Savino, Joe; Lee, Wayne; Dalton, Jerry; Underwood, Mark;

Restoration Of MEX SRC Images For Improved Topography: A New Image Product

NASA Astrophysics Data System (ADS)

Duxbury, T. C.

2012-12-01

Surface topography is an important constraint when investigating the evolution of solar system bodies. Topography is typically obtained from stereo photogrammetric or photometric (shape from shading) analyses of overlapping / stereo images and from laser / radar altimetry data. The ESA Mars Express Mission [1] carries a Super Resolution Channel (SRC) as part of the High Resolution Stereo Camera (HRSC) [2]. The SRC can build up overlapping / stereo coverage of Mars, Phobos and Deimos by viewing the surfaces from different orbits. The derivation of high precision topography data from the SRC raw images is degraded because the camera is out of focus. The point spread function (PSF) is multi-peaked, covering tens of pixels. After registering and co-adding hundreds of star images, an accurate SRC PSF was reconstructed and is being used to restore the SRC images to near blur free quality. The restored images offer a factor of about 3 in improved geometric accuracy as well as identifying the smallest of features to significantly improve the stereo photogrammetric accuracy in producing digital elevation models. The difference between blurred and restored images provides a new derived image product that can provide improved feature recognition to increase spatial resolution and topographic accuracy of derived elevation models. Acknowledgements: This research was funded by the NASA Mars Express Participating Scientist Program. [1] Chicarro, et al., ESA SP 1291(2009) [2] Neukum, et al., ESA SP 1291 (2009). A raw SRC image (h4235.003) of a Martian crater within Gale crater (the MSL landing site) is shown in the upper left and the restored image is shown in the lower left. A raw image (h0715.004) of Phobos is shown in the upper right and the difference between the raw and restored images, a new derived image data product, is shown in the lower right. The lower images, resulting from an image restoration process, significantly improve feature recognition for improved derived topographic accuracy.

Sex Differences in Time to Return-to-Play Progression After Sport-Related Concussion.

PubMed

Stone, Sarah; Lee, Bobby; Garrison, J Craig; Blueitt, Damond; Creed, Kalyssa

2016-10-03

Recently, female sports participation has increased, and there is a tendency for women to experience more symptoms and variable presentation after sport-related concussion (SRC). The purpose of this study was to determine whether sex differences exist in time to begin a return-to-play (RTP) progression after an initial SRC. After initial SRC, female athletes (11-20 years old) would take longer to begin an RTP progression compared with age-matched male athletes. Retrospective cohort study. Level 3. A total of 579 participants (365 males [mean age, 15.0 ± 1.7 years], 214 females [mean age, 15.2 ± 1.5 years]), including middle school, high school, and collegiate athletes who participated in various sports and experienced an initial SRC were included and underwent retrospective chart review. The following information was collected: sex, age at injury, sport, history of prior concussion, date of injury, and date of initiation of RTP progression. Participants with a history of more than 1 concussion or injury sustained from non-sport-related activity were excluded. Despite American football having the greatest percentage (49.2%) of sport participation, female athletes took significantly longer to start an RTP progression after an initial SRC (29.1 ± 26.3 days) compared with age-matched male athletes (22.7 ± 18.3 days; P = 0.002). On average, female athletes took approximately 6 days longer to begin an RTP progression compared with age-matched male athletes. This suggests that sex differences exist between athletes, ages 11 to 20 years, with regard to initiation of an RTP progression after SRC. Female athletes may take longer to recover after an SRC, and therefore, may take longer to return to sport. Sex should be considered as part of the clinical decision-making process when determining plan of care for this population. © 2016 The Author(s).

Integrin {alpha}{beta}1, {alpha}{sub v}{beta}, {alpha}{sub 6}{beta} effectors p130Cas, Src and talin regulate carcinoma invasion and chemoresistance

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sansing, Hope A.; Sarkeshik, Ali; Yates, John R.

2011-03-11

Research highlights: {yields} Proteomics of clustered integrin {alpha}{beta}1, {alpha}{sub v}{beta}, {alpha}{sub 6}{beta} receptors in oral carcinoma. {yields} p130Cas, Dek, Src and talin regulate oral carcinoma invasion. {yields} p130Cas, talin, Src and zyxin regulate oral carcinoma resistance to cisplatin. -- Abstract: Ligand engagement by integrins induces receptor clustering and formation of complexes at the integrin cytoplasmic face that controls cell signaling and cytoskeletal dynamics critical for adhesion-dependent processes. This study searches for a subset of integrin effectors that coordinates both tumor cell invasion and resistance to the chemotherapeutic drug cisplatin in oral carcinomas. Candidate integrin effectors were identified in a proteomicsmore » screen of proteins recruited to clustered integrin {alpha}{beta}1, {alpha}{sub v}{beta} or {alpha}{sub 6}{beta} receptors in oral carcinomas. Proteins with diverse functions including microtubule and actin binding proteins, and factors involved in trafficking, transcription and translation were identified in oral carcinoma integrin complexes. Knockdown of effectors in the oral carcinoma HN12 cells revealed that p130Cas, Dek, Src and talin were required for invasion through Matrigel. Disruption of talin or p130Cas by RNA interference increased resistance to cisplatin, whereas targeting Dek, Src or zyxin reduced HN12 resistance to cisplatin. Analysis of the spreading of HN12 cells on collagen I and laminin I revealed that a decrease in p130Cas or talin expression inhibited spreading on both matrices. Interestingly, a reduction in zyxin expression enhanced spreading on laminin I and inhibited spreading on collagen I. Reduction of Dek, Src, talin or zyxin expression reduced HN12 proliferation by 30%. Proliferation was not affected by a reduction in p130Cas expression. We conclude that p130Cas, Src and talin function in both oral carcinoma invasion and resistance to cisplatin.« less

Activation pathway of Src kinase reveals intermediate states as novel targets for drug design

PubMed Central

Shukla, Diwakar; Meng, Yilin; Roux, Benoît; Pande, Vijay S.

2014-01-01

Unregulated activation of Src kinases leads to aberrant signaling, uncontrolled growth, and differentiation of cancerous cells. Reaching a complete mechanistic understanding of large scale conformational transformations underlying the activation of kinases could greatly help in the development of therapeutic drugs for the treatment of these pathologies. In principle, the nature of conformational transition could be modeled in silico via atomistic molecular dynamics simulations, although this is very challenging due to the long activation timescales. Here, we employ a computational paradigm that couples transition pathway techniques and Markov state model-based massively distributed simulations for mapping the conformational landscape of c-src tyrosine kinase. The computations provide the thermodynamics and kinetics of kinase activation for the first time, and help identify key structural intermediates. Furthermore, the presence of a novel allosteric site in an intermediate state of c-src that could be potentially utilized for drug design is predicted. PMID:24584478

Apatinib inhibits cellular invasion and migration by fusion kinase KIF5B-RET via suppressing RET/Src signaling pathway

PubMed Central

Xie, Weiwei; Zheng, Rongliang; Gan, Yu; Chang, Jianhua

2016-01-01

The Rearranged during transfection (RET) fusion gene is a newly identified oncogenic mutation in non-small cell lung cancer (NSCLC). The aim of this study is to explore the biological functions of the gene in tumorigenesis and metastasis in RET gene fusion-driven preclinical models. We also investigate the anti-tumor activity of Apatinib, a potent inhibitor of VEGFR-2, PDGFR-β, c-Src and RET, in RET-rearranged lung adenocarcinoma, together with the mechanisms underlying. Our results suggested that KIF5B-RET fusion gene promoted cell invasion and migration, which were probably mediated through Src signaling pathway. Apatinib exerted its anti-cancer effect not only via cytotoxicity, but also via inhibition of migration and invasion by suppressing RET/Src signaling pathway, supporting a potential role for Apatinib in the treatment of KIF5B-RET driven tumors. PMID:27494860

Apatinib inhibits cellular invasion and migration by fusion kinase KIF5B-RET via suppressing RET/Src signaling pathway.

PubMed

Lin, Chen; Wang, Shanshan; Xie, Weiwei; Zheng, Rongliang; Gan, Yu; Chang, Jianhua

2016-09-13

The Rearranged during transfection (RET) fusion gene is a newly identified oncogenic mutation in non-small cell lung cancer (NSCLC). The aim of this study is to explore the biological functions of the gene in tumorigenesis and metastasis in RET gene fusion-driven preclinical models. We also investigate the anti-tumor activity of Apatinib, a potent inhibitor of VEGFR-2, PDGFR-β, c-Src and RET, in RET-rearranged lung adenocarcinoma, together with the mechanisms underlying. Our results suggested that KIF5B-RET fusion gene promoted cell invasion and migration, which were probably mediated through Src signaling pathway. Apatinib exerted its anti-cancer effect not only via cytotoxicity, but also via inhibition of migration and invasion by suppressing RET/Src signaling pathway, supporting a potential role for Apatinib in the treatment of KIF5B-RET driven tumors.

Fetal-to-maternal signaling to initiate parturition

PubMed Central

Reinl, Erin L.; England, Sarah K.

2015-01-01

Multiple processes are capable of activating the onset of parturition; however, the specific contributions of the mother and the fetus to this process are not fully understood. In this issue of the JCI, Gao and colleagues present evidence that steroid receptor coactivators 1 and 2 (SRC-1 and SRC-2) regulate surfactant protein-A (SP-A) and platelet-activating factor (PAF) expression, which increases in the developing fetal lung. WT dams crossed with males deficient for both SRC-1 and SRC-2 had suppressed myometrial inflammation, increased serum progesterone, and delayed parturition, which could be reconciled by injection of either SP-A or PAF into the amnion. Together, the results of this study demonstrate that the fetal lungs produce signals to initiate labor in the mouse. This work underscores the importance of the fetus as a contributor to the onset of murine, and potentially human, parturition. PMID:26098207

Effect of anodic treatment on the electrocatalytic activity of superficial Raney nickel catalyst in cathodic hydrogen

DOE Office of Scientific and Technical Information (OSTI.GOV)

Korovin, N.V.; Kozlova, N.I.; Kumenko, M.V.

This work is concerned with the effect of oxidation on the activity of Raney nickel catalyst in cathodic hydrogen evolution. The superficial Raney nickel catalyst (nickel SRC) was prepared by a previously described procedure. The surface of the nickel SRC was oxidized by applying an anodic sweep over the potential range from 0.25 to 1.00 V with a potential sweep rate of 1 mV/sec. The rate of cathodic hydrogen evolution increases after pretreatment of the surface of nickel SRC by application of an anodic pulse. A significant increase in the reaction rate most probably is due to oxygen adsorption onmore » the nickel SRC surface. The largest increase in the amount of weakly bound hydrogen corresponds to the most active electrode. Oxidation of the nickel surface by an anodic pulse causes both an acceleration and a retardation of the cathodic hydrogen evolution reaction.« less

Tropomyosin Tm5NM1 Spatially Restricts Src Kinase Activity through Perturbation of Rab11 Vesicle Trafficking

PubMed Central

Bach, Cuc T.; Murray, Rachael Z.; Owen, Dylan; Gaus, Kat

2014-01-01

In order for cells to stop moving, they must synchronously stabilize actin filaments and their associated focal adhesions. How these two structures are coordinated in time and space is not known. We show here that the actin association protein Tm5NM1, which induces stable actin filaments, concurrently suppresses the trafficking of focal-adhesion-regulatory molecules. Using combinations of fluorescent biosensors and fluorescence recovery after photobleaching (FRAP), we demonstrate that Tm5NM1 reduces the level of delivery of Src kinase to focal adhesions, resulting in reduced phosphorylation of adhesion-resident Src substrates. Live imaging of Rab11-positive recycling endosomes that carry Src to focal adhesions reveals disruption of this pathway. We propose that tropomyosin synchronizes adhesion dynamics with the cytoskeleton by regulating actin-dependent trafficking of essential focal-adhesion molecules. PMID:25288639

Signaling by ectopically expressed Drosophila Src64 requires the protein-tyrosine phosphatase corkscrew and the adapter downstream of receptor kinases.

PubMed

Cooper, J A; Simon, M A; Kussick, S J

1996-11-01

Vertebrate Src can be activated by specific mutations to become oncogenic. Analogous mutations in Drosophila Src64 (DSrc) induce abnormal differentiation of photoreceptor cells when expressed ectopically in the developing Drosophila adult eye. We have investigated the roles that the adapter protein, Downstream of receptor kinases (Drk), and the SH2 domain-containing tyrosine phosphatase, Corkscrew (Csw), play in this process. We find that dominant-negative mutations in either the drk or csw genes ameliorate the developmental abnormalities induced by activated DSrc. This suggests that Drk and Csw are required downstream of, or parallel to, DSrc. Csw does not act solely as an upstream activator of DSrc. The results are discussed in relation to potential roles for the vertebrate homologues of Drk and Csw (Grb2 and SHP2, respectively) in the transformation of fibroblasts by vertebrate Src.

Biodiesel production using waste frying oil

DOE Office of Scientific and Technical Information (OSTI.GOV)

Charpe, Trupti W.; Rathod, Virendra K., E-mail: vk.rathod@ictmumbai.edu.in

2011-01-15

Research highlights: {yields} Waste sunflower frying oil is successfully converted to biodiesel using lipase as catalyst. {yields} Various process parameters that affects the conversion of transesterification reaction such as temperature, enzyme concentration, methanol: oil ratio and solvent are optimized. {yields} Inhibitory effect of methanol on lipase is reduced by adding methanol in three stages. {yields} Polar solvents like n-hexane and n-heptane increases the conversion of tranesterification reaction. - Abstract: Waste sunflower frying oil is used in biodiesel production by transesterification using an enzyme as a catalyst in a batch reactor. Various microbial lipases have been used in transesterification reaction tomore » select an optimum lipase. The effects of various parameters such as temperature, methanol:oil ratio, enzyme concentration and solvent on the conversion of methyl ester have been studied. The Pseudomonas fluorescens enzyme yielded the highest conversion. Using the P. fluorescens enzyme, the optimum conditions included a temperature of 45 deg. C, an enzyme concentration of 5% and a methanol:oil molar ratio 3:1. To avoid an inhibitory effect, the addition of methanol was performed in three stages. The conversion obtained after 24 h of reaction increased from 55.8% to 63.84% because of the stage-wise addition of methanol. The addition of a non-polar solvent result in a higher conversion compared to polar solvents. Transesterification of waste sunflower frying oil under the optimum conditions and single-stage methanol addition was compared to the refined sunflower oil.« less

HCl, KCl and KOH solvation resolved solute-solvent interactions and solution surface stress

NASA Astrophysics Data System (ADS)

Zhang, Xi; Xu, Yan; Zhou, Yong; Gong, Yinyan; Huang, Yongli; Sun, Chang Q.

2017-11-01

An incorporation of the hydrogen bond (O:Hsbnd O or HB) cooperativity notion, contact angle detection, and the differential phonon spectrometrics (DPS) has enabled us to gain refined information on the HCl, KCl and KOH solvation resolved solute-solvent molecular interactions and the solution surface stresses. Results show that ionic polarization stiffens the solvent Hsbnd O bond phonon from 3200 to 3480 cm-1 in the hydration shells. The HO- in alkaline solution, however, shares not only the same Hsbnd O phonon redshift of compressed water from 3200 to < 3100 cm-1 but also the dangling bonds of H2O surface featured at 3610 cm-1. Salt and alkaline solvation enhances the solution surface stress by K+ and Cl- ionic polarization. The excessive H+ proton in acid solution forms a H↔H anti-HB that depresses the solution surface stress, instead. The solute capability of transforming the fraction of the O:Hsbnd O bonds of the solvent matrix is featured by: fH = 0 and fx ∝ 1-exp(-C/C0) (x = HO-, K+ and Cl-) towards saturation. Exercises not only confirm the presence of the H↔H anti-HB point fragilization, the O:⇔:O super-HB point compression, and ionic polarization dominating the performance of the respective HCl, KOH, and KCl solutions, but also demonstrate the power of the DPS that enables high resolution of solute-solute-solvent interactions and correlation between HB relaxation and solution surface stress.

Structure of the ThDP-dependent enzyme benzaldehyde lyase refined to 1.65 Å resolution

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maraite, Andy; Schmidt, Thomas; Ansörge-Schumacher, Marion B.

2007-07-01

The X-ray crystal structure of the ThDP-dependent enzyme benzaldehyde lyase has been refined to 1.65 Å. Benzaldehyde lyase (BAL; EC 4.1.2.38) is a thiamine diphosphate (ThDP) dependent enzyme that catalyses the enantioselective carboligation of two molecules of benzaldehyde to form (R)-benzoin. BAL has hence aroused interest for its potential in the industrial synthesis of optically active benzoins and derivatives. The structure of BAL was previously solved to a resolution of 2.6 Å using MAD experiments on a selenomethionine derivative [Mosbacher et al. (2005 ▶), FEBS J.272, 6067–6076]. In this communication of parallel studies, BAL was crystallized in an alternative spacemore » group (P2{sub 1}2{sub 1}2{sub 1}) and its structure refined to a resolution of 1.65 Å, allowing detailed observation of the water structure, active-site interactions with ThDP and also the electron density for the co-solvent 2-methyl-2,4-pentanediol (MPD) at hydrophobic patches of the enzyme surface.« less

Oxidative stability of refined olive and sunflower oils supplemented with lycopene-rich oleoresin from tomato peels industrial by-product, during accelerated shelf-life storage.

PubMed

Kehili, Mouna; Choura, Sirine; Zammel, Ayachi; Allouche, Noureddine; Sayadi, Sami

2018-04-25

Tomato peels by-product from a Tunisian industry was used for the extraction of lycopene-rich oleoresin using hexane solvent maceration. Tomato peels oleoresin, TPO, exhibited competitive free radicals scavenging activity with synthetic antioxidants. The efficacy of TPO in stabilizing refined olive (ROO) and sunflower (RSO) oils was investigated for five months, under accelerated shelf-life, compared to the synthetic antioxidant, butylated hydroxytoluene (BHT). TPO was added to ROO and RSO at four different concentrations, namely 250, 500, 1000 and 2000 µg/g and BHT standard at 200 µg/g. Lipid oxidation was tracked by measuring the peroxide value, acidity, conjugated dienes and trienes. Results suggested the highest efficiency of 250 µg/g and 2000 µg/g of TPO, referring to 5 µg/g and 40 µg/g of lycopene, for the oxidative stabilization of ROO and RSO, respectively. The protective effect of TPO against the primary oxidation of these refined oils was significantly correlated to their lycopene contents. Copyright © 2017 Elsevier Ltd. All rights reserved.

Neutron study of B 12 coenzyme at 15 K

NASA Astrophysics Data System (ADS)

Bouquiere, J. P.

1992-06-01

This high resolution and low temperature study, at 15 K, of the vitamin B 12 coenzyme ( C72H100N18O17PCo) was undertaken to confirm and clarify the water networks identified at 279 K by Savage [1]. Details of the data collection and refinement of the low temperature structure are described, a comparison of the coenzyme molecule structures at 15 and 279 K is made, and that of solvent structures outlined.

A refined 2010-based VOC emission inventory and its improvement on modeling regional ozone in the Pearl River Delta Region, China.

PubMed

Yin, Shasha; Zheng, Junyu; Lu, Qing; Yuan, Zibing; Huang, Zhijiong; Zhong, Liuju; Lin, Hui

2015-05-01

Accurate and gridded VOC emission inventories are important for improving regional air quality model performance. In this study, a four-level VOC emission source categorization system was proposed. A 2010-based gridded Pearl River Delta (PRD) regional VOC emission inventory was developed with more comprehensive source coverage, latest emission factors, and updated activity data. The total anthropogenic VOC emission was estimated to be about 117.4 × 10(4)t, in which on-road mobile source shared the largest contribution, followed by industrial solvent use and industrial processes sources. Among the industrial solvent use source, furniture manufacturing and shoemaking were major VOC emission contributors. The spatial surrogates of VOC emission were updated for major VOC sources such as industrial sectors and gas stations. Subsector-based temporal characteristics were investigated and their temporal variations were characterized. The impacts of updated VOC emission estimates and spatial surrogates were evaluated by modeling O₃ concentration in the PRD region in the July and October of 2010, respectively. The results indicated that both updated emission estimates and spatial allocations can effectively reduce model bias on O₃ simulation. Further efforts should be made on the refinement of source classification, comprehensive collection of activity data, and spatial-temporal surrogates in order to reduce uncertainty in emission inventory and improve model performance. Copyright © 2015 Elsevier B.V. All rights reserved.

Near-Channel Versus Watershed Controls on Sediment Rating Curves

NASA Astrophysics Data System (ADS)

Vaughan, Angus A.; Belmont, Patrick; Hawkins, Charles P.; Wilcock, Peter

2017-10-01

Predicting riverine suspended sediment flux is a fundamental problem in geomorphology, with important implications for water quality, land and water resource management, and aquatic ecosystem health. To advance understanding, we evaluated environmental and landscape factors that influence sediment rating curves (SRCs). We generated SRCs with recent total suspended solids (TSSs) and discharge data from 45 gages on 36 rivers throughout the state of Minnesota, USA. Watersheds range from 32 to 14,600 km2 and represent distinct settings regarding topography, land cover, and geologic history. Rivers exhibited three distinct SRC shapes: simple power functions, threshold power functions, and peaked or negative-slope functions. We computed SRC exponents and coefficients (describing the steepness of the relation and the TSS concentration at median flows, respectively). In addition to quantifying watershed topography, climate/hydrology, geology, soil type, and land cover, we used lidar topography to characterize the near-channel environment upstream of gages. We used random forest models to analyze relations between SRC parameters and attributes of the watershed and the near-channel environment. The models correctly classify 78% of SRC shapes and explain 37%-60% of variance in SRC parameters. We find that SRC steepness (exponent) is strongly related to near-channel morphological characteristics including near-channel relief, channel gradient, and presence of lakes along the local channel network, but not to land use. In contrast, land use influences TSS concentrations at moderate and low flow. These findings suggest that the near-channel environment controls changes in TSS as flows increase, whereas land use drives median and low flow TSS conditions.

Tyrosine phosphorylation of LRP6 by Src and Fer inhibits Wnt/β-catenin signalling

PubMed Central

Chen, Qing; Su, Yi; Wesslowski, Janine; Hagemann, Anja I; Ramialison, Mirana; Wittbrodt, Joachim; Scholpp, Steffen; Davidson, Gary

2014-01-01

Low-density lipoprotein receptor-related proteins 5 and 6 (LRP5/6) function as transmembrane receptors to transduce Wnt signals. A key mechanism for signalling is Wnt-induced serine/threonine phosphorylation at conserved PPPSPxS motifs in the LRP6 cytoplasmic domain, which promotes pathway activation. Conserved tyrosine residues are positioned close to all PPPSPxS motifs, which suggests they have a functional significance. Using a cell culture-based cDNA expression screen, we identified the non-receptor tyrosine kinases Src and Fer as novel LRP6 modifiers. Both Src and Fer associate with LRP6 and phosphorylate LRP6 directly. In contrast to the known PPPSPxS Ser/Thr kinases, tyrosine phosphorylation by Src and Fer negatively regulates LRP6-Wnt signalling. Epistatically, they function upstream of β-catenin to inhibit signalling and in agreement with a negative role in regulating LRP6, MEF cells lacking these kinases show enhanced Wnt signalling. Wnt3a treatment of cells enhances tyrosine phosphorylation of endogenous LRP6 and, mechanistically, Src reduces cell surface LRP6 levels and disrupts LRP6 signalosome formation. Interestingly, CK1γ inhibits Fer-induced LRP6 phosphorylation, suggesting a mechanism whereby CK1γ acts to de-represses inhibitory LRP6 tyrosine phosphorylation. We propose that LRP6 tyrosine phosphorylation by Src and Fer serves a negative regulatory function to prevent over-activation of Wnt signalling at the level of the Wnt receptor, LRP6. Subject Categories Membrane & Intracellular Transport; Post-translational Modifications, Proteolysis & Proteomics PMID:25391905

Policies, Procedures, and Practices Regarding Sport-Related Concussion in Community College Athletes.

PubMed

Paddack, Michael; DeWolf, Ryan; Covassin, Tracey; Kontos, Anthony

2016-01-01

College sport organizations and associations endorse concussion-management protocols and policies. To date, little information is available on concussion policies and practices at community college institutions. To assess and describe current practices and policies regarding the assessment, management, and return-to-play criteria for sport-related concussion (SRC) among member institutions of the California Community College Athletic Association (CCCAA). Cross-sectional study. Web-based survey. A total of 55 head athletic trainers (ATs) at CCCAA institutions. Data about policies, procedures, and practices regarding SRC were collected over a 3-week period in March 2012 and analyzed using descriptive statistics, the Fisher exact test, and the Spearman test. Almost half (47%) of ATs stated they had a policy for SRC assessment, management, and return to play at their institution. They reported being in compliance with baseline testing guidelines (25%), management guidelines (34.5%), and return-to-play guidelines (30%). Nearly 31% of ATs described having an SRC policy in place for academic accommodations. Conference attendance was positively correlated with institutional use of academic accommodations after SRC (r = 0.44, P = .01). The number of meetings ATs attended and their use of baseline testing were also positively correlated (r = 0.38, P = .01). At the time of this study, nearly half of CCCAA institutions had concussion policies and 31% had academic-accommodation policies. However, only 18% of ATs at CCCAA institutions were in compliance with all of their concussion policies. Our findings demonstrate improvements in the management of SRCs by ATs at California community colleges compared with previous research but a need for better compliance with SRC policies.

Correlation between E-cadherin interactions, survivin expression, and apoptosis in MDCK and ts-Src MDCK cell culture models.

PubMed

Capra, Janne; Eskelinen, Sinikka

2017-12-01

Survivin, a member of inhibitor of apoptosis (IAP) protein family, is a multifunctional protein expressed in most cancers. In addition to inhibition of apoptosis, it regulates proliferation and promotes migration. Its presence and function in cells is strongly regulated via transcription factors, intracellular localization, and degradation. We analyzed the presence of survivin at protein level in various culture environments and under activation of Src tyrosine kinase in epithelial canine kidney MDCK cells in order to elucidate factors controlling survivin 'lifespan'. We used untransformed and temperature sensitive ts-Src MDCK cells as a model and forced them to grow in suspension (1D), in 2D on hard and soft surfaces and in soft 3D Matrigel environment with or without EGTA. In addition, we tested the effect of stressful conditions by cultivating the cells in the presence of an anti-cancer drug and a generator of reactive oxygen species (ROS), piperlongumine (PL) with or without an antioxidant, N-acetylcysteine (NAC). We could confirm that inhibition of apoptosis and simultaneous downregulation of survivin in MDCK cells required both intact cell-cell junctions, trans-interactions of E-cadherin and soft 3D matrix environment. In ts-Src-transformed MDCK cells, survivin was upregulated as soon as the cell-cell junctions were disintegrated. ROS generation with PL-induced cell death of ts-Src MDCK cells concomitantly with survivin downregulation. NAC rescued the ts-Src MDCK cells from ROS-induced apoptosis without upregulation of survivin resulting in a situation resembling untransformed MDCK cells in 3D environment and E-cadherin delineating the lateral cell walls.

The use of assistive technology in the everyday lives of young people living with dementia and their caregivers. Can a simple remote control make a difference?

PubMed

Jentoft, Rita; Holthe, Torhild; Arntzen, Cathrine

2014-12-01

This study was a part of a larger study exploring the impact of assistive technology on the lives of young people living with dementia (YPD). This paper focuses on one of the most useful devices, the simple remote control (SRC). The objective was to explore the reason why the SRC is significant and beneficial in the everyday lives of YPD and their caregivers. This qualitative longitudinal study had a participatory design. Eight participants received an SRC. The range for using it was 0-15 months. In-depth interviews and observations were conducted at baseline and repeated every third month up to 18 months. A situated learning approach was used in the analysis to provide a deeper understanding of the significance and use of SRC. Young people having dementia spend a substantial amount of time alone. Watching television was reported to be important, but handling remote controls was challenging and created a variety of problems. YPD learned to use SRC, which made important differences in the everyday lives of all family members. Comprehensive support from caregivers and professionals was important for YPD in the learning process. The SRC was deemed a success because it solved challenges regarding the use of television in everyday lives of families. The design was recognizable and user-friendly, thus allowing YPD to learn its operation. Access to professional support and advice regarding assistive technology is vital for establishing a system for follow-up and continued collaboration to make future adaptations and adjustments.

Sustained expression of steroid receptor coactivator SRC-2/TIF-2 is associated with better prognosis in malignant pleural mesothelioma.

PubMed

Jennings, Cormac J; O'Grady, Anthony; Cummins, Robert; Murer, Bruno; Al-Alawi, Mazen; Madden, Stephen F; Mutti, Luciano; Harvey, Brian J; Thomas, Warren; Kay, Elaine W

2012-01-01

Estrogen receptor beta (ERβ) overexpression by malignant pleural mesothelioma (MPM) tumor cells correlates with enhanced patient survival. ER-regulated transcription is mediated by the p160 family of steroid receptor coactivators (SRCs), and SRC isoform overexpression is associated with worse prognosis in many steroid-related malignancies. The aim of this study was to establish whether SRC isoform expression varied between individual MPM tumors with positive or negative prognostic significance. Immunohistochemical analysis of tumor biopsies from 89 subjects with confirmed histological diagnosis of MPM and biopsies from 3 normal control subjects was performed to detect the expression of SRC-1, SRC-2 (TIF-2), SRC-3 (AIB-1), and ERβ. Allred scores for expression of ERβ and each of the SRCs were determined, and Kaplan-Meier survival curves were calculated to correlate biomarker expression, gender, and histology type with postdiagnosis survival. ERβ and all the SRCs were expressed at high levels in normal pleural mesothelium, and expression of each biomarker was reduced or lost in a subset of the MPM subjects; however, postdiagnosis survival only significantly correlated with TIF-2 expression. Low or intermediate expression of TIF-2 correlated with reduced median postdiagnosis survival (9 months) compared with those subjects whose tumors highly expressed TIF-2 (20 months) (p = 0.036, log-rank test). Maintained high expression of TIF-2 in tumor cells is a positive prognostic indicator for postdiagnosis survival in patients with confirmed MPM. This is the first clinical study to correlate high TIF-2 expression with improved patient prognosis in any malignancy.

Deregulated Expression of SRC, LYN and CKB Kinases by DNA Methylation and Its Potential Role in Gastric Cancer Invasiveness and Metastasis

PubMed Central

Rey, Juan Antonio; Pinto, Giovanny Rebouças; Lamarão, Leticia Martins; Montenegro, Raquel Carvalho; Alves, Ana Paula Negreiros Nunes; Assumpção, Paulo Pimentel; Borges, Barbara do Nascimento; Smith, Marília Cardoso; Burbano, Rommel Rodriguez

2015-01-01

Kinases are downstream modulators and effectors of several cellular signaling cascades and play key roles in the development of neoplastic disease. In this study, we aimed to evaluate SRC, LYN and CKB protein and mRNA expression, as well as their promoter methylation, in gastric cancer. We found elevated expression of SRC and LYN kinase mRNA and protein but decreased levels of CKB kinase, alterations that may have a role in the invasiveness and metastasis of gastric tumors. Expression of the three studied kinases was also associated with MYC oncogene expression, a possible biomarker for gastric cancer. To understand the mechanisms that regulate the expression of these genes, we evaluated the DNA promoter methylation of the three kinases. We found that reduced SRC and LYN methylation and increased CKB methylation was associated with gastric cancer. The reduced SRC and LYN methylation was associated with increased levels of mRNA and protein expression, suggesting that DNA methylation is involved in regulating the expression of these kinases. Conversely, reduced CKB methylation was observed in samples with reduced mRNA and protein expression, suggesting CKB expression was found to be only partly regulated by DNA methylation. Additionally, we found that alterations in the DNA methylation pattern of the three studied kinases were also associated with the gastric cancer onset, advanced gastric cancer, deeper tumor invasion and the presence of metastasis. Therefore, SRC, LYN and CKB expression or DNA methylation could be useful markers for predicting tumor progression and targeting in anti-cancer strategies. PMID:26460485

Deregulated Expression of SRC, LYN and CKB Kinases by DNA Methylation and Its Potential Role in Gastric Cancer Invasiveness and Metastasis.

PubMed

Mello, Adriano Azevedo; Leal, Mariana Ferreira; Rey, Juan Antonio; Pinto, Giovanny Rebouças; Lamarão, Leticia Martins; Montenegro, Raquel Carvalho; Alves, Ana Paula Negreiros Nunes; Assumpção, Paulo Pimentel; Borges, Barbara do Nascimento; Smith, Marília Cardoso; Burbano, Rommel Rodriguez

2015-01-01

Kinases are downstream modulators and effectors of several cellular signaling cascades and play key roles in the development of neoplastic disease. In this study, we aimed to evaluate SRC, LYN and CKB protein and mRNA expression, as well as their promoter methylation, in gastric cancer. We found elevated expression of SRC and LYN kinase mRNA and protein but decreased levels of CKB kinase, alterations that may have a role in the invasiveness and metastasis of gastric tumors. Expression of the three studied kinases was also associated with MYC oncogene expression, a possible biomarker for gastric cancer. To understand the mechanisms that regulate the expression of these genes, we evaluated the DNA promoter methylation of the three kinases. We found that reduced SRC and LYN methylation and increased CKB methylation was associated with gastric cancer. The reduced SRC and LYN methylation was associated with increased levels of mRNA and protein expression, suggesting that DNA methylation is involved in regulating the expression of these kinases. Conversely, reduced CKB methylation was observed in samples with reduced mRNA and protein expression, suggesting CKB expression was found to be only partly regulated by DNA methylation. Additionally, we found that alterations in the DNA methylation pattern of the three studied kinases were also associated with the gastric cancer onset, advanced gastric cancer, deeper tumor invasion and the presence of metastasis. Therefore, SRC, LYN and CKB expression or DNA methylation could be useful markers for predicting tumor progression and targeting in anti-cancer strategies.

Leap-dynamics: efficient sampling of conformational space of proteins and peptides in solution.

PubMed

Kleinjung, J; Bayley, P; Fraternali, F

2000-03-31

A molecular simulation scheme, called Leap-dynamics, that provides efficient sampling of protein conformational space in solution is presented. The scheme is a combined approach using a fast sampling method, imposing conformational 'leaps' to force the system over energy barriers, and molecular dynamics (MD) for refinement. The presence of solvent is approximated by a potential of mean force depending on the solvent accessible surface area. The method has been successfully applied to N-acetyl-L-alanine-N-methylamide (alanine dipeptide), sampling experimentally observed conformations inaccessible to MD alone under the chosen conditions. The method predicts correctly the increased partial flexibility of the mutant Y35G compared to native bovine pancreatic trypsin inhibitor. In particular, the improvement over MD consists of the detection of conformational flexibility that corresponds closely to slow motions identified by nuclear magnetic resonance techniques.

Biological production of acetic acid from waste gases with Clostridium ljungdahlii

DOEpatents

Gaddy, James L.

1998-01-01

A method and apparatus for converting waste gases from industrial processes such as oil refining, carbon black, coke, ammonia, and methanol production, into useful products. The method includes introducing the waste gases into a bioreactor where they are fermented to various organic acids or alcohols by anaerobic bacteria within the bioreactor. These valuable end products are then recovered, separated and purified. In an exemplary recovery process, the bioreactor raffinate is passed through an extraction chamber into which one or more non-inhibitory solvents are simultaneously introduced to extract the product. Then, the product is separated from the solvent by distillation. Gas conversion rates can be maximized by use of centrifuges, hollow fiber membranes, or other means of ultrafiltration to return entrained anaerobic bacteria from the bioreactor raffinate to the bioreactor itself, thus insuring the highest possible cell concentration.

Clostridium stain which produces acetic acid from waste gases

DOEpatents

Gaddy, James L.

1997-01-01

A method and apparatus for converting waste gases from industrial processes such as oil refining, carbon black, coke, ammonia, and methanol production, into useful products. The method includes introducing the waste gases into a bioreactor where they are fermented to various organic acids or alcohols by anaerobic bacteria within the bioreactor. These valuable end products are then recovered, separated and purified. In an exemplary recovery process, the bioreactor raffinate is passed through an extraction chamber into which one or more non-inhibitory solvents are simultaneously introduced to extract the product. Then, the product is separated from the solvent by distillation. Gas conversion rates can be maximized by use of centrifuges, hollow fiber membranes, or other means of ultrafiltration to return entrained anaerobic bacteria from the bioreactor raffinate to the bioreactor itself, thus insuring the highest possible cell concentration.

Clostridium strain which produces acetic acid from waste gases

DOEpatents

Gaddy, J.L.

1997-01-14

A method and apparatus are disclosed for converting waste gases from industrial processes such as oil refining, carbon black, coke, ammonia, and methanol production, into useful products. The method includes introducing the waste gases into a bioreactor where they are fermented to various organic acids or alcohols by anaerobic bacteria within the bioreactor. These valuable end products are then recovered, separated and purified. In an exemplary recovery process, the bioreactor raffinate is passed through an extraction chamber into which one or more non-inhibitory solvents are simultaneously introduced to extract the product. Then, the product is separated from the solvent by distillation. Gas conversion rates can be maximized by use of centrifuges, hollow fiber membranes, or other means of ultrafiltration to return entrained anaerobic bacteria from the bioreactor raffinate to the bioreactor itself, thus insuring the highest possible cell concentration. 4 figs.

Royal Society, Discussion on New Coal Chemistry, London, England, May 21, 22, 1980, Proceedings

NASA Astrophysics Data System (ADS)

1981-03-01

A discussion of new coal chemistry is presented. The chemical and physical structure of coal is examined in the first section, including structural studies of coal extracts, metal and metal complexes in coal and coal microporosity. The second section presents new advances in applied coal technology. The development of liquid fuels and chemicals from coal is given especial emphasis, with papers on the Sasol Synthol process, the Shell-Koppers gasification process, liquefaction and gasification in Germany, the Solvent Refined Coal process, the Exxon Donor Solvent liquefaction process and the Mobil Methanol-to-Gasoline process. Finally, some developments that will be part of the future of coal chemistry in the year 2000 are examined in the third section, including coal-based chemical complexes and the use of coal as an alternative source to oil for chemical feedstocks.

A multiwavelength frequency-domain near-infrared cerebral oximeter

NASA Astrophysics Data System (ADS)

Kurth, C. Dean; Thayer, William S.

1999-03-01

This study tests a multiwavelength frequency-domain near-infrared oximeter (fdNIRS) in an in vitro model of the human brain. The model is a solid plastic structure containing a vascular network perfused with blood in which haemoglobin oxygen saturation was measured by co-oximetry, providing a standard for comparison. Plastic shells of varying thickness (0.5-2 cm), with a vascular network of their own and encircling the brain model, were also added to simulate extracranial tissues of the infant, child and adult. The fdNIRS oximeter utilizes frequency-domain technology to monitor phaseshifts at 754 nm, 785 nm and 816 nm relative to a 780 nm reference to derive through photon transport and Beer-Lambert equations. We found a linear relationship between fdNIRS and co-oximetry with excellent correlation that fitted the line of identity in all experiments ( n = 7). The bias of fdNIRS oximetry was -2% and the precision was 6%. Blood temperature and fdNIRS source-detector distance did not affect fdNIRS oximetry. Low haemoglobin concentration altered the fdNIRS versus co-oximetry line slope and intercept, producing a 15% error at the extremes of . The infant- and child-like shells overlying the brain model did not alter fdNIRS oximetry, whereas the adult-like shell yielded an error as high as 32%. In conclusion, fdNIRS accurately measures in an in vitro brain model, although low haemoglobin concentration and extracranial tissue of adult thickness influence accuracy.

Role of c-Src in cellular events associated with colony-stimulating factor-1-induced spreading in osteoclasts.

PubMed

Insogna, K; Tanaka, S; Neff, L; Horne, W; Levy, J; Baron, R

1997-01-01

We and others have observed that in response to treatment with Colony Stimulating Factor-1 (CSF-1) neonatal rat osteoclasts demonstrate rapid cytoplasmic spreading. The receptor for CSF-1, c-Fms, is expressed in osteoclasts, possesses intrinsic tyrosine-kinase activity, and signals via rapid phosphorylation of selected proteins. It has been reported previously that c-Src becomes tyrosine phosphorylated following CSF-1 treatment of fibroblasts overexpressing c-Fms. We therefore examined the cellular events associated with CSF-1-induced spreading in osteoclasts and what role, if any, c-Src played in these processes. Confocal microscopic studies using phosphotyrosine (P-tyr) monoclonal antibodies demonstrated that CSF-1 induced a significant dose- and time-dependent increase in P-tyr labeling of neonatal rat osteoclasts. Phalloidin staining was consistent with partial to complete disassembly of the actin attachment ring with redistribution of actin to the spreading cytoplasmic edge of the cell. Quantitation of cellular F-actin using NBD-phallicidin confirmed a decrease in polymerized actin following exposure to CSF-1. In contrast, CSF-1 failed to induce any cytoplasmic spreading in osteoclasts isolated from mice with targeted disruption of the src gene. Further, in src- osteoclasts no well defined attachment ring could be identified. To investigate cell-signaling events associated with osteoclast spreading, detergent lysates were made from purified multinucleated osteoclast-like cells (OCLs) obtained by coculturing murine bone marrow and osteoblasts with calcitriol. Western blot analyses of lysates from control and CSF-1-treated normal cells indicated that several proteins were specifically phosphorylated in response to CSF-1, most notably proteins of 165, 60, and 85-90 kDa. Immunoprecipitation studies revealed that the 165 and 60 kDa proteins were, respectively, c-Fms and c-Src. The c-Src kinase activity was increased 2.9-fold following CSF-1 treatment. The 85-90 kDa protein is as yet unidentified. Since activated receptor tyrosine kinases may induce spreading in part by reducing phosphoinositol 4,5-bisphosphate (PIP2) binding to actin-associated proteins, a monoclonal antibody to PIP2 was used to assess the nature of PIP2 binding proteins in OCLs. Proteins of 85-90 kDa, 43 kDa, and 30 kDa were consistently demonstrated to bind PIP2. Further, the PIP2 content of the 85-90 kDa protein appeared to decrease with CSF-1 treatment. Whether this protein represents the phosphoprotein of the same M.W. is unclear. We also examined the effect of CSF-1 on the PIP2 content of alpha-actinin. Alpha-actinin showed low-level PIP2 binding, which was demonstrable only after immuno-precipitation and did not change with CSF-1 treatment. However, CSF-1 did cause a significant decline in the phosphotyrosine content of alpha-actinin. In contrast, in src- OCLs, CSF-1 induced more prolonged phosphorylation of c-Fms, and the 85-90 kDa protein was markedly hypophosphorylated. Further, alpha-actinin did not dephosphorylate in src- cells. We conclude that CSF-1-induced osteoclast spreading is accompanied by rapid reorganization of the actin cytoskeleton and phosphorylation of several cellular substrates, including c-Fms and c-Src. PIP2 binding to at least one protein appears to decrease with CSF-1 treatment, which may favor actin depolymerization. The reduced tyrosine phosphorylation of alpha-actinin could effect its ability to bind to actin. Thus c-Src may play an important role in these cellular events since in its absence, osteoclasts do not spread and signaling events downstream are altered. Whether these changes relate in part to the basal abnormalities in the cytoskeletal organization of src- osteoclasts remains to be determined.

78 FR 17428 - Notice of Open Public Meetings for the National Park Service Alaska Region's Subsistence Resource...

Federal Register 2010, 2011, 2012, 2013, 2014

2013-03-21

... Purpose 6. Commission Membership Status 7. SRC Chair and SRC Members' Reports 8. Superintendent's Report 9... a. Red Dog Road Study Update b. Marine Resources (Seals/Walrus) 11. Federal Subsistence Board Update...

AmeriFlux US-SRC Santa Rita Creosote

DOE Data Explorer

Kurc, Shirley [University of Arizona

2016-01-01

This is the AmeriFlux version of the carbon flux data for the site US-SRC Santa Rita Creosote. Site Description - Part of the Santa Rita Experimental Range since 1901; Site vegetation has been dominated by Creosote bush since at least 1934

Role of SRC-3delta4 in the Progression and Metastasis of Castration-Resistant Prostate Cancer

DTIC Science & Technology

2013-10-01

Expression of SRC-3∆4, GAPDH, and AR target genes including PSA, KLK2, IGFBP5, Cyclin A2, and UBE2C was determined by RT-qPCR analysis . Data are...Expression of AR (B), GAPDH, and TMPRSS2- ERG (C) was determined by RT-qPCR analysis . Data are presented using the comparative Ct method, in which GAPDH...input. An irrelevant region (1800 bp downstream of transcription start site) was served as a negative control. (E) and (F). ChIP analysis of SRC-3∆4’s

Iron depletion results in Src kinase inhibition with associated cell cycle arrest in neuroblastoma cells

PubMed Central

Siriwardana, Gamini; Seligman, Paul A

2015-01-01

Iron is required for cellular proliferation. Recently, using systematic time studies of neuroblastoma cell growth, we better defined the G1 arrest caused by iron chelation to a point in mid-G1, where cyclin E protein is present, but the cyclin E/CDK2 complex kinase activity is inhibited. In this study, we again used the neuroblastoma SKNSH cells lines to pinpoint the mechanism responsible for this G1 block. Initial studies showed in the presence of DFO, these cells have high levels of p27 and after reversal of iron chelation p27 is degraded allowing for CDK2 kinase activity. The initial activation of CDK2 kinase allows cells to exit G1 and enter S phase. Furthermore, we found that inhibition of p27 degradation by DFO is directly associated with inhibition of Src kinase activity measured by lack of phosphorylation of Src at the 416 residue. Activation of Src kinase occurs very early after reversal from the DFO G1 block and is temporally associated with initiation of cellular proliferation associated with entry into S phase. For the first time therefore we show that iron chelation inhibits Src kinase activity and this activity is a requirement for cellular proliferation. PMID:25825542

Attentional Modulation of Emotional Conflict Processing with Flanker Tasks

PubMed Central

Zhou, Pingyan; Liu, Xun

2013-01-01

Emotion processing has been shown to acquire priority by biasing allocation of attentional resources. Aversive images or fearful expressions are processed quickly and automatically. Many existing findings suggested that processing of emotional information was pre-attentive, largely immune from attentional control. Other studies argued that attention gated the processing of emotion. To tackle this controversy, the current study examined whether and to what degrees attention modulated processing of emotion using a stimulus-response-compatibility (SRC) paradigm. We conducted two flanker experiments using color scale faces in neutral expressions or gray scale faces in emotional expressions. We found SRC effects for all three dimensions (color, gender, and emotion) and SRC effects were larger when the conflicts were task relevant than when they were task irrelevant, suggesting that conflict processing of emotion was modulated by attention, similar to those of color and face identity (gender). However, task modulation on color SRC effect was significantly greater than that on gender or emotion SRC effect, indicating that processing of salient information was modulated by attention to a lesser degree than processing of non-emotional stimuli. We proposed that emotion processing can be influenced by attentional control, but at the same time salience of emotional information may bias toward bottom-up processing, rendering less top-down modulation than that on non-emotional stimuli. PMID:23544155

Src kinase signaling mediates estrous behavior induced by 5β-reduced progestins, GnRH, prostaglandin E2 and vaginocervical stimulation in estrogen-primed rats.

PubMed

Lima-Hernández, Francisco J; Beyer, Carlos; Gómora-Arrati, Porfirio; García-Juárez, Marcos; Encarnación-Sánchez, José L; Etgen, Anne M; González-Flores, Oscar

2012-11-01

The progesterone receptor (PR) is a dual function protein that acts in the nucleus as a transcriptional factor and at the cytoplasm as a scaffold for the Src-MAPK signaling pathway. Several agents lacking affinity for the PR, such as 5β-reduced progestins, GnRH or prostaglandin E(2) (PGE(2)) facilitate estrous behavior in ovariectomized (ovx), estrogen-primed rats yet their action is blocked by the antiprogestin RU486. We hypothesize that these agents act by using the PR-Src-mitogen activated protein kinase alternative pathway. To test this hypothesis we used PP2, a specific inhibitor of the Src kinase family. Intraventricular infusion of 30 μg of PP2, 30 min before behavioral testing, significantly attenuated estrous behaviors induced in estradiol benzoate (E(2)B)-primed rats by 5β-dihydroprogesterone (5β-DHP), 5β-pregnan-3β-ol-20-one (5β,3β-Pgl), GnRH, PGE(2) and by manual flank/vaginocervical stimulation. These results suggest that the Src signaling system, by activating mitogen-activated protein kinases, participates in the facilitation of estrous behavior in E(2)B-primed rats induced by agents lacking affinity for the PR. Copyright © 2012 Elsevier Inc. All rights reserved.

Dynamin Forms a Src Kinase–sensitive Complex with Cbl and Regulates Podosomes and Osteoclast Activity

PubMed Central

Bruzzaniti, Angela; Neff, Lynn; Sanjay, Archana; Horne, William C.; De Camilli, Pietro; Baron, Roland

2005-01-01

Podosomes are highly dynamic actin-containing adhesion structures found in osteoclasts, macrophages, and Rous sarcoma virus (RSV)-transformed fibroblasts. After integrin engagement, Pyk2 recruits Src and the adaptor protein Cbl, forming a molecular signaling complex that is critical for cell migration, and deletion of any molecule in this complex disrupts podosome ring formation and/or decreases osteoclast migration. Dynamin, a GTPase essential for endocytosis, is also involved in actin cytoskeleton remodeling and is localized to podosomes where it has a role in actin turnover. We found that dynamin colocalizes with Cbl in the actin-rich podosome belt of osteoclasts and that dynamin forms a complex with Cbl in osteoclasts and when overexpressed in 293VnR or SYF cells. The association of dynamin with Cbl in osteoclasts was decreased by Src tyrosine kinase activity and we found that destabilization of the dynamin-Cbl complex involves the recruitment of Src through the proline-rich domain of Cbl. Overexpression of dynamin increased osteoclast bone resorbing activity and migration, whereas overexpression of dynK44A decreased osteoclast resorption and migration. These studies suggest that dynamin, Cbl, and Src coordinately participate in signaling complexes that are important in the assembly and remodeling of the actin cytoskeleton, leading to changes in osteoclast adhesion, migration, and resorption. PMID:15872089

Decipher the dynamic coordination between enzymatic activity and structural modulation at focal adhesions in living cells

NASA Astrophysics Data System (ADS)

Lu, Shaoying; Seong, Jihye; Wang, Yi; Chang, Shiou-Chi; Eichorst, John Paul; Ouyang, Mingxing; Li, Julie Y.-S.; Chien, Shu; Wang, Yingxiao

2014-07-01

Focal adhesions (FAs) are dynamic subcellular structures crucial for cell adhesion, migration and differentiation. It remains an enigma how enzymatic activities in these local complexes regulate their structural remodeling in live cells. Utilizing biosensors based on fluorescence resonance energy transfer (FRET), we developed a correlative FRET imaging microscopy (CFIM) approach to quantitatively analyze the subcellular coordination between the enzymatic Src activation and the structural FA disassembly. CFIM reveals that the Src kinase activity only within the microdomain of lipid rafts at the plasma membrane is coupled with FA dynamics. FA disassembly at cell periphery was linearly dependent on this raft-localized Src activity, although cells displayed heterogeneous levels of response to stimulation. Within lipid rafts, the time delay between Src activation and FA disassembly was 1.2 min in cells seeded on low fibronectin concentration ([FN]) and 4.3 min in cells on high [FN]. CFIM further showed that the level of Src-FA coupling, as well as the time delay, was regulated by cell-matrix interactions, as a tight enzyme-structure coupling occurred in FA populations mediated by integrin αvβ3, but not in those by integrin α5β1. Therefore, different FA subpopulations have distinctive regulation mechanisms between their local kinase activity and structural FA dynamics.

Chlorogenic acid inhibits hypoxia-induced pulmonary artery smooth muscle cells proliferation via c-Src and Shc/Grb2/ERK2 signaling pathway.

PubMed

Li, Qun-Yi; Zhu, Ying-Feng; Zhang, Meng; Chen, Li; Zhang, Zhen; Du, Yong-Li; Ren, Guo-Qiang; Tang, Jian-Min; Zhong, Ming-Kang; Shi, Xiao-Jin

2015-03-15

Chlorogenic acid (CGA), abundant in coffee and particular fruits, can modulate hypertension and vascular dysfunction. Hypoxia-induced pulmonary artery smooth muscle cells (PASMCs) proliferation has been tightly linked to vascular remodeling in pulmonary arterial hypertension (PAH). Thus, the present study was designed to investigate the effect of CGA on hypoxia-induced proliferation in cultured rat PASMCs. The data showed that CGA potently inhibited PASMCs proliferation and DNA synthesis induced by hypoxia. These inhibitory effects were associated with G1 cell cycle arrest and down-regulation of cell cycle proteins. Treatment with CGA reduced hypoxia-induced hypoxia inducible factor 1α (HIF-1α) expression and trans-activation. Furthermore, hypoxia-evoked c-Src phosphorylation was inhibited by CGA. In vitro ELISA-based tyrosine kinase assay indicated that CGA was a direct inhibitor of c-Src. Moreover, CGA attenuated physical co-association of c-Src/Shc/Grb2 and ERK2 phosphorylation in PASMCs. These results suggest that CGA inhibits hypoxia-induced proliferation in PASMCs via regulating c-Src-mediated signaling pathway. In vivo investigation showed that chronic CGA treatment inhibits monocrotaline-induced PAH in rats. These findings presented here highlight the possible therapeutic use of CGA in hypoxia-related PAH. Copyright © 2015 Elsevier B.V. All rights reserved.

FAK/Src family of kinases: protective or aggravating factor for ischemia reperfusion injury in nervous system?

PubMed

Bikis, Christos; Moris, Demetrios; Vasileiou, Ioanna; Patsouris, Eustratios; Theocharis, Stamatios

2015-04-01

The focal adhesion kinase (FAK) and the Src families of kinases are subfamilies of the non-receptor protein tyrosine kinases. FAK activity is regulated by gene amplification, alternative splicing and phosporylation/dephosphorylation. FAK/Src complex has been found to participate through various pathways in neuronal models of ischemia-reperfusion injury (IRI) with conflicting results. The aim of the present review is to summarize the currently available data on this subject. The MEDLINE/PubMed database was searched for publications with the medical subject heading IRI and FAK and/or Src, nervous system. We restricted our search till 2014. We identified 93 articles that were available in English as abstracts or/and full-text articles that were deemed appropriate for our review. FAK has been found to have a beneficial preconditioning effect on IRI through activation via the protein kinase C (PKC) pathway by anesthetic agents. Of great importance are the interactions between FAK/Src and VEGF that has been already detected as a protective mean for IRI. The effect of VEGF administration might depend on dose as well as on time of administration. A Ca(2+)/calmodulin-dependent protein kinase II or PKC inhibitors seem to have protective effects on IRI by inhibiting ion channels activation.

Annatto Tocotrienol Induces a Cytotoxic Effect on Human Prostate Cancer PC3 Cells via the Simultaneous Inhibition of Src and Stat3.

PubMed

Sugahara, Ryosuke; Sato, Ayami; Uchida, Asuka; Shiozawa, Shinya; Sato, Chiaki; Virgona, Nantiga; Yano, Tomohiro

2015-01-01

Prostate cancer is one of the most frequently occurring cancers and often acquires the potential of androgen-independent growth as a malignant phenotype. Androgen-independent prostate cancer has severe chemoresistance towards conventional chemotherapeutic agents, so a new treatment approach is required for curing such prostate cancer. In this context, the present study was undertaken to check if annatto tocotrienol (main component δ-tocotrienol) could suppress cell growth in human prostate cancer (PC3, androgen-independent type) cells via the inhibition of Src and Stat3. The tocotrienol showed cytotoxic effects on PC3 cells in a dose-dependent manner, and the effect depended on G1 arrest in the cell cycle and subsequent induction of apoptosis. In a cytotoxic dose, the tocotrienol suppressed cellular growth via the simultaneous inhibition of Src and Stat3. Similarly, the treatment combination of both Src and Stat3 inhibitors induced cytotoxic effects in PC3 cells in an additive manner compared to each by itself. With respect to cell cycle regulation and the induction of apoptosis, the combination treatment showed a similar effect to that of the tocotrienol treatment. These results suggest that annatto tocotrienol effectively induces cytotoxicity in androgen-independent prostate cancer cells via the suppression of Src and Stat3.

N-(5-chloro-1,3-benzodioxol-4-yl)-7-[2-(4-methylpiperazin-1-yl)ethoxy]-5- (tetrahydro-2H-pyran-4-yloxy)quinazolin-4-amine, a novel, highly selective, orally available, dual-specific c-Src/Abl kinase inhibitor.

PubMed

Hennequin, Laurent F; Allen, Jack; Breed, Jason; Curwen, Jon; Fennell, Michael; Green, Tim P; Lambert-van der Brempt, Christine; Morgentin, Rémy; Norman, Richard A; Olivier, Annie; Otterbein, Ludovic; Plé, Patrick A; Warin, Nicolas; Costello, Gerard

2006-11-02

Src family kinases (SFKs) are nonreceptor tyrosine kinases that are reported to be critical for cancer progression. We report here a novel subseries of C-5-substituted anilinoquinazolines that display high affinity and specificity for the tyrosine kinase domain of the c-Src and Abl enzymes. These compounds exhibit high selectivity for SFKs over a panel of recombinant protein kinases, excellent pharmacokinetics, and in vivo activity following oral dosing. N-(5-Chloro-1,3-benzodioxol-4-yl)-7-[2-(4-methylpiperazin-1-yl)ethoxy]-5-(tetrahydro-2H-pyran-4-yloxy)quinazolin-4-amine (AZD0530) inhibits c-Src and Abl enzymes at low nanomolar concentrations and is highly selective over a range of kinases. AZD0530 displays excellent pharmacokinetic parameters in animal preclinically and in man (t(1/2) = 40 h). AZD0530 is a potent inhibitor of tumor growth in a c-Src-transfected 3T3-fibroblast xenograft model in vivo and led to a significant increase in survival in a highly aggressive, orthotopic model of human pancreatic cancer when dosed orally once daily. AZD0530 is currently undergoing clinical evaluation in man.

Inappropriate activation of the androgen receptor by nonsteroids: involvement of the Src kinase pathway and its therapeutic implications.

PubMed

Desai, Sonal J; Ma, Ai-Hong; Tepper, Clifford G; Chen, Hong-Wu; Kung, Hsing-Jien

2006-11-01

The inappropriate activation of androgen receptor (AR) by nonsteroids is considered a potential mechanism in the emergence of hormone-refractory prostate tumors, but little is known about the properties of these "pseudoactivated" AR. Here, we present the first comprehensive analysis closely examining the properties of AR activated by the neuropeptide bombesin that distinguish it from androgen-activated AR. We show that bombesin-activated AR (a) is required for bombesin-induced growth of LNCaP cells, (b) has a transcriptional profile overlapping with, but not identical to, androgen-activated AR, (c) activates prostate-specific antigen by preferentially binding to its proximal promoter, and (d) assembles a distinct coactivator complex. Significantly, we found that Src kinase is critical for bombesin-induced AR-mediated activity and is required for translocation and transactivation of AR. Additionally, we identify c-Myc, a Src target gene, to be activated by bombesin and a potential coactivator of AR-mediated activity specific to bombesin-induced signaling. Because Src kinase is often activated by other nonsteroids, such as other neuropeptides, growth factors, chemokines, and cytokines, our findings have general applicability and provide rationale for investigating the efficacy of the Src kinase pathway as a target for the prevention of relapsed prostate cancers.

EF24 Suppresses Invasion and Migration of Hepatocellular Carcinoma Cells In Vitro via Inhibiting the Phosphorylation of Src

PubMed Central

Tin, Lamtin; Wu, Yiqi; Jin, Yinji; Jin, Xiaoming; Zhang, Fengmin

2016-01-01

Diphenyl difluoroketone (EF24), a curcumin analog, is a promising anticancer compound that exerts its effects by inhibiting cell proliferation and inducing apoptosis. However, the efficacy of EF24 against cancer metastasis, particularly in hepatocellular carcinoma (HCC), remains elusive. In this study, the effect of EF24 on HCCLM-3 and HepG2 cell migration and invasion was detected by wound healing and transwell assay, respectively. The results revealed that EF24 suppressed the migration and invasion of both HCCLM-3 and HepG2 cells. Furthermore, EF24 treatment decreased the formation of filopodia on the cell surface and inhibited the phosphorylation of Src in both cell lines, which may help contribute towards understanding the mechanism underlying the suppressive effect of EF24 on HCC migration and invasion. Additionally, the expression of total- and phosphorylated-Src in primary HCC tissues and their paired lymph node metastatic tissues was detected, and phosphorylated-Src was found to be associated with HCC lymph node metastasis. The results of this study suggest that Src is a novel and promising therapeutic target in HCC and provide evidence to support the hypothesis that EF24 may be a useful therapeutic agent for the treatment of HCC. PMID:27999817

EF24 Suppresses Invasion and Migration of Hepatocellular Carcinoma Cells In Vitro via Inhibiting the Phosphorylation of Src.

PubMed

Zhao, Ran; Tin, Lamtin; Zhang, Yuhua; Wu, Yiqi; Jin, Yinji; Jin, Xiaoming; Zhang, Fengmin; Li, Xiaobo

2016-01-01

Diphenyl difluoroketone (EF24), a curcumin analog, is a promising anticancer compound that exerts its effects by inhibiting cell proliferation and inducing apoptosis. However, the efficacy of EF24 against cancer metastasis, particularly in hepatocellular carcinoma (HCC), remains elusive. In this study, the effect of EF24 on HCCLM-3 and HepG2 cell migration and invasion was detected by wound healing and transwell assay, respectively. The results revealed that EF24 suppressed the migration and invasion of both HCCLM-3 and HepG2 cells. Furthermore, EF24 treatment decreased the formation of filopodia on the cell surface and inhibited the phosphorylation of Src in both cell lines, which may help contribute towards understanding the mechanism underlying the suppressive effect of EF24 on HCC migration and invasion. Additionally, the expression of total- and phosphorylated-Src in primary HCC tissues and their paired lymph node metastatic tissues was detected, and phosphorylated-Src was found to be associated with HCC lymph node metastasis. The results of this study suggest that Src is a novel and promising therapeutic target in HCC and provide evidence to support the hypothesis that EF24 may be a useful therapeutic agent for the treatment of HCC.

Attentional modulation of emotional conflict processing with flanker tasks.

PubMed

Zhou, Pingyan; Liu, Xun

2013-01-01

Emotion processing has been shown to acquire priority by biasing allocation of attentional resources. Aversive images or fearful expressions are processed quickly and automatically. Many existing findings suggested that processing of emotional information was pre-attentive, largely immune from attentional control. Other studies argued that attention gated the processing of emotion. To tackle this controversy, the current study examined whether and to what degrees attention modulated processing of emotion using a stimulus-response-compatibility (SRC) paradigm. We conducted two flanker experiments using color scale faces in neutral expressions or gray scale faces in emotional expressions. We found SRC effects for all three dimensions (color, gender, and emotion) and SRC effects were larger when the conflicts were task relevant than when they were task irrelevant, suggesting that conflict processing of emotion was modulated by attention, similar to those of color and face identity (gender). However, task modulation on color SRC effect was significantly greater than that on gender or emotion SRC effect, indicating that processing of salient information was modulated by attention to a lesser degree than processing of non-emotional stimuli. We proposed that emotion processing can be influenced by attentional control, but at the same time salience of emotional information may bias toward bottom-up processing, rendering less top-down modulation than that on non-emotional stimuli.

Post-traumatic stress avoidance is attenuated by corticosterone and associated with brain levels of steroid receptor co-activator-1 in rats.

PubMed

Whitaker, Annie M; Farooq, Muhammad A; Edwards, Scott; Gilpin, Nicholas W

2016-01-01

Individuals with post-traumatic stress disorder (PTSD) avoid trauma-related stimuli and exhibit blunted hypothalamic-pituitary-adrenal (HPA) axis activation at the time of stress. Our rodent model of stress mimics the avoidance symptom cluster of PTSD. Rats are classified as "Avoiders" or "Non-Avoiders" post-stress based on the avoidance of a predator-odor paired context. Previously, we found Avoiders exhibit an attenuated HPA stress response to predator odor. We hypothesized that corticosterone administration before stress would reduce the magnitude and incidence of stress-paired context avoidance. Furthermore, we also predicted that Avoiders would exhibit altered expression of glucocorticoid receptor (GR) signaling machinery elements, including steroid receptor co-activator (SRC)-1. Male Wistar rats (n = 16) were pretreated with corticosterone (25 mg/kg) or saline and exposed to predator-odor stress paired with a context and tested for avoidance 24 h later. A second group of corticosterone-naïve rats (n = 24) were stressed (or not), indexed for avoidance 24 h later, and killed 48 h post-odor exposure to measure phosphorylated GR, FKBP51 and SRC-1 levels in the paraventricular nucleus (PVN), central amygdala (CeA) and ventral hippocampus (VH), all brain sites that highly express GRs and regulate HPA function. Corticosterone pretreatment reduced the magnitude and incidence of avoidance. In Avoiders, predator-odor exposure led to lower SRC-1 expression in the PVN and CeA, and higher SRC-1 expression in the VH. SRC-1 expression in PVN, CeA and VH was predicted by prior avoidance behavior. Hence, a blunted HPA stress response may contribute to stress-induced neuroadaptations in central SRC-1 levels and behavioral dysfunction in Avoider rats.

What is the difference in concussion management in children as compared with adults? A systematic review.

PubMed

Davis, Gavin A; Anderson, Vicki; Babl, Franz E; Gioia, Gerard A; Giza, Christopher C; Meehan, William; Moser, Rosemarie Scolaro; Purcell, Laura; Schatz, Philip; Schneider, Kathryn J; Takagi, Michael; Yeates, Keith Owen; Zemek, Roger

2017-06-01

To evaluate the evidence regarding the management of sport-related concussion (SRC) in children and adolescents. The eight subquestions included the effects of age on symptoms and outcome, normal and prolonged duration, the role of computerised neuropsychological tests (CNTs), the role of rest, and strategies for return to school and return to sport (RTSp). Systematic review. MEDLINE (OVID), Embase (OVID) and PsycInfo (OVID). Studies were included if they were original research on SRC in children aged 5 years to 18 years, and excluded if they were review articles, or did not focus on childhood SRC. A total of 5853 articles were identified, and 134 articles met the inclusion criteria. Some articles were common to multiple subquestions. Very few studies examined SRC in young children, aged 5-12 years. This systematic review recommends that in children: child and adolescent age-specific paradigms should be applied; child-validated symptom rating scales should be used; the widespread routine use of baseline CNT is not recommended; the expected duration of symptoms associated with SRC is less than 4 weeks; prolonged recovery be defined as symptomatic for greater than 4 weeks; a brief period of cognitive and physical rest should be followed with gradual symptom-limited physical and cognitive activity; all schools be encouraged to have a concussion policy and should offer appropriate academic accommodations and support to students recovering from SRC; and children and adolescents should not RTSp until they have successfully returned to school, however early introduction of symptom-limited physical activity is appropriate. PROSPERO 2016:CRD42016039184. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Activation of Src and release of intracellular calcium by phosphatidic acid during Xenopus laevis fertilization

PubMed Central

Bates, Ryan C.; Fees, Colby P.; Holland, William L.; Winger, Courtney C.; Batbayar, Khulan; Ancar, Rachel; Bergren, Todd; Petcoff, Douglas; Stith, Bradley J.

2014-01-01

We report a new step in the fertilization in Xenopus laevis which has been found to involve activation of Src tyrosine kinase to stimulate phospholipase C-γ (PLC- γ) which increases inositol 1,4,5-trisphosphate (IP3) to release intracellular calcium ([Ca]i). Molecular species analysis and mass measurements suggested that sperm activate phospholipase D (PLD) to elevate phosphatidic acid (PA). We now report that PA mass increased 2.7 fold by 1 minute after insemination and inhibition of PA production by two methods inhibited activation of Src and PLCγ, increased [Ca]i and other fertilization events. As compared to 14 other lipids, PA strongly bound Xenopus Src but not PLCγ. Addition of synthetic PA activated egg Src (an action requiring intact lipid rafts) and PLCγ as well as doubling the amount of PLCγ in rafts. In the absence of elevated [Ca]i, PA addition elevated IP3 mass to levels equivalent to that induced by sperm (but twice that achieved by calcium ionophore). Finally, PA induced [Ca]i release that was blocked by an IP3 receptor inhibitor. As only PLD1b message was detected, and Western blotting did not detect PLD2, we suggest that sperm activate PLD1b to elevate PA which then binds to and activates Src leading to PLCγ stimulation, IP3 elevation and [Ca]i release. Due to these and other studies, PA may also play a role in membrane fusion events such as sperm-egg fusion, cortical granule exocytosis, the elevation of phosphatidylinositol 4,5-bisphosphate and the large, late increase in sn 1,2-diacylglycerol in fertilization. PMID:24269904

EphA2 and Src regulate equatorial cell morphogenesis during lens development

PubMed Central

Cheng, Catherine; Ansari, Moham M.; Cooper, Jonathan A.; Gong, Xiaohua

2013-01-01

High refractive index and transparency of the eye lens require uniformly shaped and precisely aligned lens fiber cells. During lens development, equatorial epithelial cells undergo cell-to-cell alignment to form meridional rows of hexagonal cells. The mechanism that controls this morphogenesis from randomly packed cuboidal epithelial cells to highly organized hexagonal fiber cells remains unknown. In Epha2-/- mouse lenses, equatorial epithelial cells fail to form precisely aligned meridional rows; moreover, the lens fulcrum, where the apical tips of elongating epithelial cells constrict to form an anchor point before fiber cell differentiation and elongation at the equator, is disrupted. Phosphorylated Src-Y424 and cortactin-Y466, actin and EphA2 cluster at the vertices of wild-type hexagonal epithelial cells in organized meridional rows. However, phosphorylated Src and phosphorylated cortactin are not detected in disorganized Epha2-/- cells with altered F-actin distribution. E-cadherin junctions, which are normally located at the basal-lateral ends of equatorial epithelial cells and are diminished in newly differentiating fiber cells, become widely distributed in the apical, lateral and basal sides of epithelial cells and persist in differentiating fiber cells in Epha2-/- lenses. Src-/- equatorial epithelial cells also fail to form precisely aligned meridional rows and lens fulcrum. These results indicate that EphA2/Src signaling is essential for the formation of the lens fulcrum. EphA2 also regulates Src/cortactin/F-actin complexes at the vertices of hexagonal equatorial cells for cell-to-cell alignment. This mechanistic information explains how EphA2 mutations lead to disorganized lens cells that subsequently contribute to altered refractive index and cataracts in humans and mice. PMID:24026120

Augmentation of the anticancer activity of CYT997 in human prostate cancer by inhibiting Src activity.

PubMed

Teng, Yong; Cai, Yafei; Pi, Wenhu; Gao, Lixia; Shay, Chloe

2017-06-12

Abnormalities of tubulin polymerization and microtubule assembly are often seen in cancer, which make them very suitable targets for the development of therapeutic approach against rapidly dividing and aggressive cancer cells. CYT997 is a novel microtubule-disrupting agent with anticancer activity in multiple cancer types including prostate cancer. However, the molecular mechanisms of action of CYT997 in prostate cancer have not been well characterized. Src knockdown cells were achieved by lentiviral-mediated interference. The drug effects on cell proliferation were measured by MTS. The drug effects on cell viability and death were determined by Cell Titer-Glo® Luminescent cell viability kit and flow cytometry with Zombie Aqua™ staining. The drug effects on apoptosis were assessed by Cell Death Detection Elisa kit and Western blot with a cleaved PARP antibody. The drug effects on cell invasion were examined by Matrigel-coated Boyden chambers. Oxidative stress was detected by DCFH-DA staining and electrochemical biosensor. Mouse models generated by subcutaneous or intracardiac injection were used to investigate the in vivo drug efficacy in tumor growth and metastasis. CYT997 effectively inhibited proliferation, survival, and invasion of prostate cancer cells via blocking multiple oncogenic signaling cascades but not the Src pathway. Inhibition of Src expression by small hairpin RNA or inactivation of Src by dasatinib increased the CYT997-induced cytotoxicity of in vitro. Moreover, the combination of dasatinib and CYT997 exhibited a superior inhibitory effect on tumor growth and metastasis compared with either of the drugs alone. Our findings demonstrate that blockage of Src augments the anticancer effect of CYT997 on prostate cancer and suggest that co-treatment of dasatinib and CYT997 may represent an effective therapeutic regimen for limiting prostate cancer.

Bio-Energy Retains Its Mitigation Potential Under Elevated CO2

PubMed Central

Bellassen, Valentin; Njakou Djomo, Sylvestre; Lukac, Martin; Calfapietra, Carlo; Janssens, Ivan A.; Hoosbeek, Marcel R.; Viovy, Nicolas; Churkina, Galina; Scarascia-Mugnozza, Giuseppe; Ceulemans, Reinhart

2010-01-01

Background If biofuels are to be a viable substitute for fossil fuels, it is essential that they retain their potential to mitigate climate change under future atmospheric conditions. Elevated atmospheric CO2 concentration [CO2] stimulates plant biomass production; however, the beneficial effects of increased production may be offset by higher energy costs in crop management. Methodology/Main Findings We maintained full size poplar short rotation coppice (SRC) systems under both current ambient and future elevated [CO2] (550 ppm) and estimated their net energy and greenhouse gas balance. We show that a poplar SRC system is energy efficient and produces more energy than required for coppice management. Even more, elevated [CO2] will increase the net energy production and greenhouse gas balance of a SRC system with 18%. Managing the trees in shorter rotation cycles (i.e., 2 year cycles instead of 3 year cycles) will further enhance the benefits from elevated [CO2] on both the net energy and greenhouse gas balance. Conclusions/Significance Adapting coppice management to the future atmospheric [CO2] is necessary to fully benefit from the climate mitigation potential of bio-energy systems. Further, a future increase in potential biomass production due to elevated [CO2] outweighs the increased production costs resulting in a northward extension of the area where SRC is greenhouse gas neutral. Currently, the main part of the European terrestrial carbon sink is found in forest biomass and attributed to harvesting less than the annual growth in wood. Because SRC is intensively managed, with a higher turnover in wood production than conventional forest, northward expansion of SRC is likely to erode the European terrestrial carbon sink. PMID:20657833

Activation of Src and release of intracellular calcium by phosphatidic acid during Xenopus laevis fertilization.

PubMed

Bates, Ryan C; Fees, Colby P; Holland, William L; Winger, Courtney C; Batbayar, Khulan; Ancar, Rachel; Bergren, Todd; Petcoff, Douglas; Stith, Bradley J

2014-02-01

We report a new step in the fertilization in Xenopus laevis which has been found to involve activation of Src tyrosine kinase to stimulate phospholipase C-γ (PLC-γ) which increases inositol 1,4,5-trisphosphate (IP3) to release intracellular calcium ([Ca](i)). Molecular species analysis and mass measurements suggested that sperm activate phospholipase D (PLD) to elevate phosphatidic acid (PA). We now report that PA mass increased 2.7 fold by 1 min after insemination and inhibition of PA production by two methods inhibited activation of Src and PLCγ, increased [Ca](i) and other fertilization events. As compared to 14 other lipids, PA specifically bound Xenopus Src but not PLCγ. Addition of synthetic PA activated egg Src (an action requiring intact lipid rafts) and PLCγ as well as doubling the amount of PLCγ in rafts. In the absence of elevated [Ca](i), PA addition elevated IP3 mass to levels equivalent to that induced by sperm (but twice that achieved by calcium ionophore). Finally, PA induced [Ca](i) release that was blocked by an IP3 receptor inhibitor. As only PLD1b message was detected, and Western blotting did not detect PLD2, we suggest that sperm activate PLD1b to elevate PA which then binds to and activates Src leading to PLCγ stimulation, IP3 elevation and [Ca](i) release. Due to these and other studies, PA may also play a role in membrane fusion events such as sperm-egg fusion, cortical granule exocytosis, the elevation of phosphatidylinositol 4,5-bisphosphate and the large, late increase in sn 1,2-diacylglycerol in fertilization. © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Confirmation of the E(sup src)(sub Peak)-E(sub iso) (Amati) relation from the x-ray flash XRF 050416A observed by the Swift burst alert telescope

NASA Technical Reports Server (NTRS)

Sakamoti, T.; Barbier, L.; Barthelmy, S. D.; Cummings, J. R.; Fenimore, E. E.; Gehrels, N.; Hullinger, D.; Krimm, H. A.; Markwardt, C. B.

2006-01-01

We report Swift Burst Alert Telescope (BAT) observations of the X-ray flash (XRF) XRF 050416A. The fluence ratio between the 15-25 and 25-50 keV energy bands of this event is 1.5, thus making it the softest gamma-ray burst (GRB) observed by BAT so far. The spectrum is well fitted by a Band function with E(sup obs)(sub peak) of 15.0(sup +2.3)(sub -2.7) keV. Assuming the redshift of the host galaxy (z = 0.6535), the isotropic equivalent radiated energy E(sub iso) and the peak energy at the GRB rest frame (E(sup src)(sub peak)) of XRF 050416A are not only consistent with the correlation found by Amati et al. and extended to XRFs by Sakamoto et al. but also fill in the gap of this relation around the 30-80 keV range of E(sup src)(sub peak). This result tightens the validity of the E(sup src)(sub Peak)-E(sup src)(sub peak) relation from XRFs to GRBs. We also find that the jet break time estimated using the empirical relation between E(sup src)(sub peak) and the collimation corrected energy E(sub gamma), is inconsistent with the afterglow observation by the Swift X-Ray Telescope. This could be due to the extra external shock emission overlaid around the jet break time or to the nonexistence of a jet break feature for XRFs, which might be a further challenge for GRB jet emission models and XRF/GRB unification scenarios.

Development and experimental test of support vector machines virtual screening method for searching Src inhibitors from large compound libraries.

PubMed

Han, Bucong; Ma, Xiaohua; Zhao, Ruiying; Zhang, Jingxian; Wei, Xiaona; Liu, Xianghui; Liu, Xin; Zhang, Cunlong; Tan, Chunyan; Jiang, Yuyang; Chen, Yuzong

2012-11-23

Src plays various roles in tumour progression, invasion, metastasis, angiogenesis and survival. It is one of the multiple targets of multi-target kinase inhibitors in clinical uses and trials for the treatment of leukemia and other cancers. These successes and appearances of drug resistance in some patients have raised significant interest and efforts in discovering new Src inhibitors. Various in-silico methods have been used in some of these efforts. It is desirable to explore additional in-silico methods, particularly those capable of searching large compound libraries at high yields and reduced false-hit rates. We evaluated support vector machines (SVM) as virtual screening tools for searching Src inhibitors from large compound libraries. SVM trained and tested by 1,703 inhibitors and 63,318 putative non-inhibitors correctly identified 93.53%~ 95.01% inhibitors and 99.81%~ 99.90% non-inhibitors in 5-fold cross validation studies. SVM trained by 1,703 inhibitors reported before 2011 and 63,318 putative non-inhibitors correctly identified 70.45% of the 44 inhibitors reported since 2011, and predicted as inhibitors 44,843 (0.33%) of 13.56M PubChem, 1,496 (0.89%) of 168 K MDDR, and 719 (7.73%) of 9,305 MDDR compounds similar to the known inhibitors. SVM showed comparable yield and reduced false hit rates in searching large compound libraries compared to the similarity-based and other machine-learning VS methods developed from the same set of training compounds and molecular descriptors. We tested three virtual hits of the same novel scaffold from in-house chemical libraries not reported as Src inhibitor, one of which showed moderate activity. SVM may be potentially explored for searching Src inhibitors from large compound libraries at low false-hit rates.

Advancing bioluminescence imaging technology for the evaluation of anticancer agents in the MDA-MB-435-HAL-Luc mammary fat pad and subrenal capsule tumor models.

PubMed

Zhang, Cathy; Yan, Zhengming; Arango, Maria E; Painter, Cory L; Anderes, Kenna

2009-01-01

Tumors grafted s.c. or under the mammary fat pad (MFP) rarely develop efficient metastasis. By applying bioluminescence imaging (BLI) technology, the MDA-MB-435-HAL-Luc subrenal capsule (SRC) model was compared with the MFP model for disease progression, metastatic potential, and response to therapy. The luciferase-expressing MDA-MB-435-HAL-Luc cell line was used in both MFP and SRC models. BLI technology allowed longitudinal assessment of disease progression and the therapeutic response to PD-0332991, Avastin, and docetaxel. Immunohistochemical analysis of Ki67 and CD31 staining in the primary tumors was compared in these models. Caliper measurement was used in the MFP model to validate the BLI quantification of primary tumors. The primary tumors in MDA-MB-435-HAL-Luc MFP and SRC models displayed comparable growth rates and vascularity. However, tumor-bearing mice in the SRC model developed lung metastases much earlier (4 weeks) than in the MFP model (>7 weeks), and the metastatic progression contributed significantly to the survival time. In the MFP model, BLI and caliper measurements were comparable for quantifying palpable tumors, but BLI offered an advantage for detecting the primary tumors that fell below a palpable threshold and for visualizing metastases. In the SRC model, BLI allowed longitudinal assessment of the antitumor and antimetastatic effects of PD-0332991, Avastin, and docetaxel, and the results correlated with the survival benefits of these agents. The MDA-MB-435-HAL-Luc SRC model and the MFP model displayed differences in disease progression. BLI is an innovative approach for developing animal models and creates opportunities for improving preclinical evaluations of anticancer agents.

What domains of clinical function should be assessed after sport-related concussion? A systematic review.

PubMed

Feddermann-Demont, Nina; Echemendia, Ruben J; Schneider, Kathryn J; Solomon, Gary S; Hayden, K Alix; Turner, Michael; Dvořák, Jiří; Straumann, Dominik; Tarnutzer, Alexander A

2017-06-01

Sport-related concussion (SRC) is a clinical diagnosis made after a sport-related head trauma. Inconsistency exists regarding appropriate methods for assessing SRC, which focus largely on symptom-scores, neurocognitive functioning and postural stability. Systematic literature review. MEDLINE, EMBASE, PsycINFO, Cochrane-DSR, Cochrane CRCT, CINAHL, SPORTDiscus (accessed July 9, 2016). Original (prospective) studies reporting on postinjury assessment in a clinical setting and evaluation of diagnostic tools within 2 weeks after an SRC. Forty-six studies covering 3284 athletes were included out of 2170 articles. Only the prospective studies were considered for final analysis (n=33; 2416 athletes). Concussion diagnosis was typically made on the sideline by an (certified) athletic trainer (55.0%), mainly on the basis of results from a symptom-based questionnaire. Clinical domains affected included cognitive, vestibular and headache/migraine. Headache, fatigue, difficulty concentrating and dizziness were the symptoms most frequently reported. Neurocognitive testing was used in 30/33 studies (90.9%), whereas balance was assessed in 9/33 studies (27.3%). The overall quality of the studies was considered low. The absence of an objective, gold standard criterion makes the accurate diagnosis of SRC challenging. Current approaches tend to emphasise cognition, symptom assessment and postural stability with less of a focus on other domains of functioning. We propose that the clinical assessment of SRC should be symptom based and interdisciplinary. Whenever possible, the SRC assessment should incorporate neurological, vestibular, ocular motor, visual, neurocognitive, psychological and cervical aspects. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Are Canadian clinicians providing consistent sport-related concussion management advice?

PubMed

Carson, James D; Rendely, Alexandra; Garel, Alisha; Meaney, Christopher; Stoller, Jacqueline; Kaicker, Jatin; Hayden, Leigh; Moineddin, Rahim; Frémont, Pierre

2016-06-01

To compare the knowledge and use of recommendations for the management of sport-related concussion (SRC) among sport and exercise medicine physicians (SEMPs) and emergency department physicians (EDPs) to assess the success of SRC knowledge transfer across Canada. A self-administered, multiple-choice survey accessed via e-mail by SEMPs and EDPs. The survey had been assessed for content validity. Canada. The survey was completed between May and July 2012 by SEMPs who had passed the diploma examination of the Canadian Academy of Sport and Exercise Medicine and by EDPs who did not hold this diploma. Knowledge and identification of sources of concussion management information, use of concussion diagnosis strategies, and whether physicians use common and consistent terminology when explaining cognitive rest strategies to patients after an SRC. There was a response rate of 28% (305 of 1085). The SEMP and EDP response rates were 41% (147 of 360) and 22% (158 of 725), respectively. Of the responses, 41% of EDPs and 3% of SEMPs were unaware of any consensus statements on concussion in sport; 74% of SEMPs used the Sport Concussion Assessment Tool, version 2 (SCAT2), "usually or always," whereas 88% of EDPs never used the SCAT2. When queried about how cognitive rest could best be achieved after an SRC, no consistent answer was documented. Differences and a lack of consistency in the implementation of recommendations for SRC patients were identified for SEMPs and EDPs. It appears that the SCAT2 is used more in the SEMP setting than in the emergency context. Further knowledge transfer efforts and research should address the barriers to achieving more consistent advice given by physicians who attend SRC patients. Copyright© the College of Family Physicians of Canada.

The Diaphanous-related Formin FHOD1 associates with ROCK1 and promotes Src-dependent plasma membrane blebbing.

PubMed

Hannemann, Sebastian; Madrid, Ricardo; Stastna, Jana; Kitzing, Thomas; Gasteier, Judith; Schönichen, André; Bouchet, Jerome; Jimenez, Alberto; Geyer, Matthias; Grosse, Robert; Benichou, Serge; Fackler, Oliver T

2008-10-10

Diaphanous-related formins (DRFs) mediate GTPase-triggered actin rearrangements to regulate central cellular processes, such as cell motility and cytokinesis. The DRF FHOD1 interacts with the Rho-GTPase Rac1 and mediates formation of actin stress fibers in its deregulated form; the physiologically relevant activities and molecular mechanisms of endogenous FHOD1, however, are still unknown. Here we report that FHOD1 physically associates via the N-terminal part of its FH2 domain with the central domain of ROCK1. Although FHOD1 does not affect the kinase activity of ROCK1, the DRF is an efficient substrate for phosphorylation by ROCK1. Co-expression of FHOD1 and ROCK1 results in the generation of nonapoptotic plasma membrane (PM) blebs, to which the DRF is efficiently recruited. Blebbing induced by FHOD1 and ROCK1 depends on F-actin integrity, the Rho-ROCK cascade, and Src activity and is reminiscent of the recently described PM blebs triggered by expression of Src homology 4 (SH4) domain PM targeting signals. Consistently, endogenous FHOD1 is required in SH4 domain expressing cells for efficient PM blebbing and rounded cell morphology in two-dimensional cultures or three-dimensional matrices, respectively. Efficient association of FHOD1 with ROCK1, as well as recruitment of the DRF to blebs, depends on Src activity, suggesting that the functional interaction between both proteins is regulated by Src. These results define a role for endogenous FHOD1 in SH4 domain-induced blebbing and suggest that its activity is regulated by ROCK1 in a Src-dependent manner.

Cowden syndrome-associated germline SDHD variants alter PTEN nuclear translocation through SRC-induced PTEN oxidation

PubMed Central

Yu, Wanfeng; He, Xin; Ni, Ying; Ngeow, Joanne; Eng, Charis

2015-01-01

Germline mutations in the PTEN tumor-suppressor gene and germline variations in succinate dehydrogenase subunit D gene (SDHD-G12S, SDHD-H50R) are associated with a subset of Cowden syndrome and Cowden syndrome-like individuals (CS/CSL) and confer high risk of breast, thyroid and other cancers. However, very little is known about the underlying crosstalk between SDHD and PTEN in CS-associated thyroid cancer. Here, we show SDHD-G12S and SDHD-H50R lead to impaired PTEN function through alteration of its subcellular localization accompanied by resistance to apoptosis and induction of migration in both papillary and follicular thyroid carcinoma cell lines. Other studies have shown elevated proto-oncogene tyrosine kinase (SRC) activity in invasive thyroid cancer cells; so, we explore bosutinib, a specific inhibitor for SRC, to explore SRC as a mediator of SDH-PTEN crosstalk in this context. We show that SRC inhibition could rescue SDHD dysfunction-induced cellular phenotype and tumorigenesis only when wild-type PTEN is expressed, in thyroid cancer lines. Patient lymphoblast cells carrying either SDHD-G12S or SDHD-H50R also show increased nuclear PTEN and more oxidized PTEN after hydrogen peroxide treatment. Like in thyroid cells, bosutinib decreases oxidative PTEN in patient lymphoblast cells carrying SDHD variants, but not in patients carrying both SDHD variants and PTEN truncating mutations. In summary, our data suggest a novel mechanism whereby SDHD germline variants SDHD-G12S or SDHD-H50R induce thyroid tumorigenesis mediated by PTEN accumulation in the nucleus and may shed light on potential treatment with SRC inhibitors like bosutinib in PTEN-wild-type SDHD-variant/mutation positive CS/CSL patients and sporadic thyroid neoplasias. PMID:25149476

THE BINDING OF COCAINE TO CYCLODEXTRINS. (R826653)

EPA Science Inventory

Abstract

Cocaine binds into -cyclodextrin, but not detectably into