bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sphingosine 1-phosphate stimulation of the p42/p44 mitogen-activated protein kinase pathway in airway smooth muscle
Authors: RAKHIT Soma , CONWAY Ann-Marie , TATE Rothwelle , BOWER Tara , PYNE Nigel J. , PYNE Susan 
Published: August 2015

We report here that cultured airway smooth muscle cells contain transcripts of endothelial differentiation gene 1 (EDG-1), a prototypical orphan Gi-coupled receptor whose natural ligand is sphingosine 1-phosphate (S1P). This is consistent with data that showed that S1P activated both c-Src and p42/p44 mitogen-activated protein kinase (p42/p44 MAPK) in a pertussis toxin (PTX)-sensitive manner in these cells. An essential role for c-Src was confirmed by using the c-Src inhibitor, PP1, which markedly decreased p42/p44 MAPK activation. We have also shown that phosphoinositide 3-kinase (PI-3K) inhibitors (wortmannin and LY294002) decreased p42/p44 MAPK activation. An essential role for PI-3K was supported by experiments that showed that PI-3K activity was increased in Grb-2 immunoprecipitates from S1P-stimulated cells. Significantly, Grb-2 associated PI-3K activity was decreased by pretreatment of cells with PTX. Finally, we have shown that the co-stimulation of cells with platelet-derived growth factor (PDGF) and S1P (which failed to stimulate DNA synthesis) elicited a larger p42/p44 MAPK activation over a 30 min stimulation compared with each agonist alone. This was associated with a S1P-dependent increase in PDGF-stimulated DNA synthesis. These results demonstrate that S1P activates c-Src and Grb-2–PI-3K (intermediates in the p42/p44 MAPK cascade) via a PTX-sensitive mechanism. This action of S1P is consistent with the stimulation of EDG-1 receptors. S1P might also function as a co-mitogen with PDGF, producing a more robust activation of a common permissive signal transduction pathway linked to DNA synthesis.
Phospholipid-subclass-specific partitioning of lipophilic ions in membrane–water systems
Authors: ZENG Youchun , HAN Xianlin , GROSS Richard W. 
Published: August 2015

Herein, we systematically investigate phospholipid-subclass-specific alterations in the partitioning of both cationic and anionic amphiphiles to identify the importance of ester, ether and vinyl ether linkages at the sn-1 position of phospholipids in the partitioning of charged amphiphiles. The results demonstrated that the membrane–water partition coefficient of a prototypic cationic amphiphile (i.e. 3,3´-dipropylthiadicarbocyanine iodide) was approximately 2.5 times higher in membranes comprised of plasmenylcholine in comparison with membranes comprised of either phosphatidylcholine or plasmanylcholine. In striking contrast, the membrane–water partition coefficient of a prototypic anionic amphiphile [i.e. bis-(1,3-dibutylbarbituric acid)trimethine oxonol] in membranes comprised of plasmenylcholine was ≈ 2.5 times lower than that manifest in membranes comprised of phosphatidylcholine or plasmanylcholine. Utilizing these experimentally determined partition coefficients,the relative membrane dipole potential of membranes comprised of plasmenylcholine was calculated and found to be ≈ 25 mV lower than in membranes comprised of phosphatidylcholine or plasmanylcholine. This lower membrane dipole potential in membranes comprised of plasmenylcholine is equivalent to the membrane potential induced by incorporation of ≈ 25 mol% of anionic phospholipids in membranes comprised of phosphatidylcholine. Collectively, these results demonstrate that phospholipid-subclass-specific differences in the membrane dipole potential contribute to alterations in the partitioning of lipophilic ions in membrane bilayers comprised of distinct phospholipid subclasses. Moreover, they suggest that these physicochemical differences can be exploited to facilitate the targeting of charged lipophilic drugs to specific cells and subcellular membrane compartments.
Plasmalogens as endogenous antioxidants: somatic cell mutants reveal the importance of the vinyl ether
Authors: ZOELLER Raphael A. , LAKE Andrew C. , NAGAN Narasimhan , GAPOSCHKIN Daniel P. , LEGNER Margaret A. , LIEBERTHAL Wilfred 
Published: August 2015

Exposure of plasmalogen-deficient variants of the murine cell line RAW 264.7 to short-term (0–100 min) treatment with electron transport inhibitors antimycin A or cyanide (chemical hypoxia) resulted in a more rapid loss of viability than in the parent strain. Results suggested that plasmalogen-deficient cells were more sensitive to reactive oxygen species (ROS) generated during chemical hypoxia; the mutants could be rescued from chemical hypoxia by using the antioxidant Trolox, an α-tocopherol analogue, and they were more sensitive to ROS generation by plumbagin or by rose bengal treatment coupled with irradiation. In addition, the use of buffers containing 2H2O greatly enhanced the cytotoxic effect of chemical hypoxia, suggesting the involvement of singlet oxygen. We used the unique enzymic deficiencies displayed by the mutants to differentially restore either plasmenylethanolamine (the major plasmalogen species normally found in this cell line) or its biosynthetic precursor, plasmanylethanolamine. Restoration of plasmenylethanolamine, which contains the vinyl ether, resulted in wild-type-like resistance to chemical hypoxia and ROS generators, whereas increasing levels of its precursor, which bears the saturated ether, had no effect on cell survival. These findings identify the vinyl ether double bond as a crucial element in cellular protection under these conditions and support the hypothesis that plasmalogens, through the vinyl ether, act as antioxidants to protect cells against ROS. These phospholipids might protect cells from ROS-mediated damage during events such as chemical hypoxia.
AMP-activated kinase reciprocally regulates triacylglycerol synthesis and fatty acid oxidation in liver and muscle: evidence that sn-glycerol-3-phosphate acyltransferase is a novel target
Authors: MUOIO Deborah M. , SEEFELD Kimberly , WITTERS Lee A. , COLEMAN Rosalind A. 
Published: August 2015

AMP-activated kinase (AMPK) is activated in response to metabolic stresses that deplete cellular ATP, and in both liver and skeletal muscle, activated AMPK stimulates fatty acid oxidation. To determine whether AMPK might reciprocally regulate glycerolipid synthesis, we studied liver and skeletal-muscle lipid metabolism in the presence of 5-amino-4-imidazolecarboxamide (AICA) riboside, a cell-permeable compound whose phosphorylated metabolite activates AMPK. Adding AICA riboside to cultured rat hepatocytes for 3 h decreased [14C]oleate and [3H]glycerol incorporation into triacylglycerol (TAG) by 50% and 38% respectively, and decreased oleate labelling of diacylglycerol by 60%. In isolated mouse soleus, a highly oxidative muscle, incubation with AICA riboside for 90 min decreased [14C]oleate incorporation into TAG by 37% and increased 14CO2 production by 48%. When insulin was present, [14C]oleate oxidation was 49% lower and [14C]oleate incorporation into TAG was 62% higher than under basal conditions. AICA riboside blocked insulin's antioxidative and lipogenic effects, increasing fatty acid oxidation by 78% and decreasing labelled TAG 43%. Similar results on fatty acid oxidation and acylglycerol synthesis were observed in C2C12 myoblasts, and in differentiated C2C12 myotubes, AICA riboside also inhibited the hydrolysis of intracellular TAG. These data suggest that AICA riboside might inhibit sn-glycerol-3-phosphate acyltransferase (GPAT), which catalyses the committed step in the pathway of glycerolipid biosynthesis. Incubating rat hepatocytes with AICA riboside for both 15 and 30 min decreased mitochondrial GPAT activity 22–34% without affecting microsomal GPAT, diacylglycerol acyltransferase or acyl-CoA synthetase activities. Finally, purified recombinant AMPKα1 and AMPKα2 inhibited hepatic mitochondrial GPAT in a time-and ATP-dependent manner. These data show that AMPK reciprocally regulates acyl-CoA channelling towards β-oxidation and away from glycerolipid biosynthesis, and provide strong evidence that AMPK phosphorylates and inhibits mitochondrial GPAT.
Lignocellulose degradation by Phanerochaete chrysosporium: purification and characterization of the main α-galactosidase
Authors: BRUMER Harry , SIMS Paul F. G. , SINNOTT Michael L. 
Published: August 2015

The main α-galactosidase was purified to homogeneity, in 30% yield, from a solid culture of Phanerochaete chrysosporium on 1 part wheat bran/2 parts thermomechanical softwood pulp. It is a glycosylated tetramer of 50 kDa peptide chains, which gives the N-terminal sequence ADNGLAITPQMG(?W)NT(?W)NHFG(?W)DIS(?W)DTI. It is remarkably stable, with crude extracts losing no activity over 3 h at 80 °C, and the purified enzyme retaining its activity over several months at 4 °C. The kinetics of hydrolysis at 25 °C of various substrates by this retaining enzyme were measured, absolute parameters being obtained by active-site titration with 2´,4´,6´-trinitrophenyl 2-deoxy-2,2-difluoro-α-d-galactopyranoside. The variation of kcat/Km for 1-naphthyl-α-d-galactopyranoside with pH is bell-shaped, with pK1 = 1.91 and pK2 = 5.54. The αD(V/K) value for p-nitrophenyl-α-d-glucopyranoside is 1.031±0.007 at the optimal pH of 3.75 and 1.114±0.006 at pH 7.00, indicating masking of the intrinsic effect at optimal pH. There is no α-2H effect on binding galactose [αD(Ki) = 0.994±0.013]. The enzyme hydrolyses p-nitrophenyl β-l-arabinopyranoside ∼ 510 times slower than the galactoside, but has no detectable activity on the α-d-glucopyranoside or α-d-mannopyranoside. Hydrolysis of α-galactosides with poor leaving groups is Michaelian, but that of substrates with good leaving groups exhibits pronounced apparent substrate inhibition, with Kis values similar to Km values. We attribute this to the binding of the second substrate molecule to a β-galactopyranosyl-enzyme intermediate, forming an E•βGal•αGalX complex which turns over slowly, if at all. 1-Fluoro-α-d-galactopyranosyl fluoride, unlike α-d-galactopyranosyl fluoride, is a Michaelian substrate, indicating that the effect of 1-fluorine substitution is greater on the first than on the second step of the enzyme reaction.
SNAP-25a and -25b isoforms are both expressed in insulin-secreting cells and can function in insulin secretion
Authors: GONELLE-GISPERT Carmen , HALBAN Philippe A. , NIEMANN Heiner , PALMER Michael , CATSICAS Stefan , SADOUL Karin 
Published: August 2015

The tSNARE (the target-membrane soluble NSF-attachment protein receptor, where NSF is N-ethylmaleimide-sensitive fusion protein) synaptosomal-associated protein of 25 kDa (SNAP-25) is expressed in pancreatic B-cells and its cleavage by botulinum neurotoxin E (BoNT/E) abolishes stimulated secretion of insulin. In the nervous system, two SNAP-25 isoforms (a and b) have been described that are produced by alternative splicing. Here it is shown, using reverse transcriptase PCR, that messages for both SNAP-25 isoforms are expressed in primary pancreatic B and non-B cells as well as in insulin-secreting cell lines. After transfection, both isoforms can be detected at the plasma membrane as well as in an intracellular perinuclear region in the insulin-secreting cell line, HIT. To test for the functional role of the two isoforms in insulin secretion, mutant forms of SNAP-25a and b resistant against cleavage by BoNT/E were generated. Such mutant SNAP-25, when expressed in HIT cells, is not inactivated by BoNT/E and its ability to restore insulin secretion can thus be investigated. To obtain the toxin-resistant mutant isoforms, the sequence around the BoNT/E cleavage site (R176QIDRIM182) was changed to P176QIKRIT182. This is the sequence of the equivalent region of human SNAP-23 (P187–T194), which has been shown to be resistant to BoNT/E. The mutant SNAP-25 was resistant to BoNT/E in vitro and in vivo and both mutant isoforms were able to reconstitute insulin secretion from toxin-treated HIT cells.
Functional expression, quantification and cellular localization of the Hxt2 hexose transporter of Saccharomyces cerevisiae tagged with the green fluorescent protein
Authors: KRUCKEBERG Arthur L. , YE Ling , BERDEN Jan A. , DAM Karel van 
Published: August 2015

The Hxt2 glucose transport protein of Saccharomyces cerevisiae was genetically fused at its C-terminus with the green fluorescent protein (GFP). The Hxt2-GFP fusion protein is a functional hexose transporter: it restored growth on glucose to a strain bearing null mutations in the hexose transporter genes GAL2 and HXT1 to HXT7. Furthermore, its glucose transport activity in this null strain was not markedly different from that of the wild-type Hxt2 protein. We calculated from the fluorescence level and transport kinetics that induced cells had 1.4×105 Hxt2-GFP molecules per cell, and that the catalytic-centre activity of the Hxt2-GFP molecule in vivo is 53 s-1 at 30 °C. Expression of Hxt2-GFP was induced by growth at low concentrations of glucose. Under inducing conditions the Hxt2-GFP fluorescence was localized to the plasma membrane. In a strain impaired in the fusion of secretory vesicles with the plasma membrane, the fluorescence accumulated in the cytoplasm. When induced cells were treated with high concentrations of glucose, the fluorescence was redistributed to the vacuole within 4 h. When endocytosis was genetically blocked, the fluorescence remained in the plasma membrane after treatment with high concentrations of glucose.
Human ornithine transcarbamylase: crystallographic insights into substrate recognition and conformational changes
Authors: SHI Dashuang , MORIZONO Hiroki , YU Xiaolin , TONG Liang , ALLEWELL Norma M. , TUCHMAN Mendel 
Published: August 2015

Two crystal structures of human ornithine transcarbamylase (OTCase) complexed with the substrate carbamoyl phosphate (CP) have been solved. One structure, whose crystals were prepared by substituting N-phosphonacetyl-L-ornithine (PALO) liganded crystals with CP, has been refined at 2.4 Å (1 Å = 0.1nm) resolution to a crystallographic R factor of 18.4%. The second structure, whose crystals were prepared by co-crystallization with CP, has been refined at 2.6 Å resolution to a crystallographic R factor of 20.2%. These structures provide important new insights into substrate recognition and ligand-induced conformational changes. Comparison of these structures with the structures of OTCase complexed with the bisubstrate analogue PALO or CP and L-norvaline reveals that binding of the first substrate, CP, induces a global conformational change involving relative domain movement, whereas the binding of the second substrate brings the flexible SMG loop, which is equivalent to the 240s loop in aspartate transcarbamylase, into the active site. The model reveals structural features that define the substrate specificity of the enzyme and that regulate the order of binding and release of products.
Atomic coordinates and structural factors (codes 1EP9 and 1FVO respectively) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ, U.S.A.
Characterization of a novel phosphatidylinositol 3-phosphate-binding protein containing two FYVE fingers in tandem that is targeted to the Golgi
Authors: CHEUNG Peter C. F. , TRINKLE-MULCAHY Laura , COHEN Philip , LUCOCQ John M. 
Published: August 2015

We have identified a novel protein of predicted molecular mass 40kDa that contains two FYVE domains in tandem and has therefore been named TAFF1 (TAndem FYVE Fingers-1). The protein is expressed predominantly in heart and binds to PtdIns3P specifically, even though the FYVE domains in TAFF1 lacks the first Arg of the consensus sequence R(K/R)HHCR, critical for the PtdIns3P binding of other FYVE domains identified so far. The first Arg is replaced by a Thr and Ser in the N-terminal and C-terminal FYVE domains of TAFF1 respectively. Mutational analysis indicates that both FYVE domains are required for high affinity binding to PtdIns3P. Cell localization studies using a green fluorescent protein fusion show that TAFF1 is localized to the Golgi, and that the Golgi targeting sequence is located within the N-terminal 187 residues and not in either FYVE domain.
The sequence of TAFF1has been deposited with the GenBank®, EMBL, DDBJ and GSDB Nucleotide Sequence Databases under accession number AF311602.
Differences in tissue-specific and embryonic expression of mouse Ceacam1 and Ceacam2 genes
Authors: HAN Eric , PHAN Dillon , LO Piao , POY Matthew N. , BEHRINGER Richard , NAJJAR Sonia M. , LIN Sue-Hwa 
Published: August 2015

The intercellular adhesion molecule CEACAM1, also known as C-CAM1 (where CAM is cell-adhesion molecule), can function as a tumour suppressor in several carcinomas, including those of the prostate, breast, bladder and colon. This suggests that CEACAM1 may play an important role in the regulation of normal cell growth and differentiation. However, there is no direct evidence to support this putative function of CEACAM1. To elucidate its physiological function by targeted gene deletion, we isolated the Ceacam genes from a mouse 129 Sv/Ev library. Although there is only one Ceacam1 gene in humans and one in rats, two homologous genes (Ceacam1 and Ceacam2) have been identified in the mouse. Our sequence analysis revealed that the genes encoded nine exons and spanned approx. 16-17 kb (Ceacam1) and 25kb (Ceacam2). The genes were highly similar (79.6%). The major differences in the protein-coding regions were located in exons 2, 5 and 6 (76.9%, 87.0% and 78.5% similarity respectively). In addition, introns 2, 5 and 7 were also significantly different, being 29.7%, 59.8% and 64.5% similar respectively. While most of these differences were due to nucleotide substitutions, two insertions of 418 and 5849bp occurred in intron 2 of Ceacam2, and another two insertions of 1384 and 197bp occurred in introns 5 and 7 respectively. To determine whether functional redundancy exists between Ceacam1 and Ceacam2, we examined their expression in 16 mouse tissues by using semi-quantitative reverse transcription-PCR. As in human and rat, in the mouse Ceacam1 mRNA was highly abundant in the liver, small intestine, prostate and spleen. In contrast, Ceacam2 mRNA was only detected in kidney, testis and, to a lesser extent, spleen. Reverse transcription-PCR using testis RNA indicated that Ceacam2 in the testis is an alternatively spliced form containing only exons 1, 2, 5, 6, 8 and 9. In the mouse embryo, Ceacam1 mRNA was detected at day 8.5, disappeared between days 9.5 and 12.5, and re-appeared at day 19. On the other hand, no Ceacam2 mRNA was detected throughout embryonic development. The different tissue expression patterns and regulation during embryonic development suggest that the CEACAM1 and CEACAM2 proteins, although highly similar, may have different functions both during mouse development and in adulthood.
The Ceacam1 and Ceacam2 sequences have been deposited in the GenBank®/EMBL/DDBJ/GSDB Nucleotide Sequence Databases with accession numbers AF287911 and AF287912 respectively.
Reading and spelling skills inGerman third graders: Examining the role of student and context characteristics
Authors: von Suchodoletz Antje , Larsen Ross A. A. , Gunzenhauser Catherine , Fäsche Anika 
Published: August 2015

BackgroundEducational processes and outcomes are influenced by a multitude of factors, including individual and contextual characteristics. Recently, studies have demonstrated that student and context characteristics may produce unique and cumulative effects on educational outcomes.AimsThe study aimed to investigate (1) the relative contribution of student, classroom, and school characteristics to reading fluency and orthographic spelling, (2) the relative contribution of specific predictors to reading fluency and orthographic spelling within the sets of student, classroom, and school characteristics, and (3) whether the contribution of student, classroom, and school characteristics differs for reading fluency and orthographic spelling.SampleParticipants were 789German third‐grade students from 56 classrooms in 34 schools.MethodStudents completed an intelligence test and a questionnaire assessing self‐control. Reading fluency and orthographic spelling performance were assessed using standardized achievement tests. Multilevel structural equation modelling was used to control for the hierarchical structure of educational data.Results and conclusionVariances in students' reading and spelling skills were in large part explained by student characteristics (>90%). Classroom and school characteristics yielded little variance. Student‐level intelligence and self‐control were significantly related to reading fluency. For orthographic spelling, student‐level intelligence and self‐control, class‐average intelligence, and, at the school level, the socio‐economic status of the school's neighbourhood were significant predictors. Future research needs to investigate relevant classroom and school factors that may directly and indirectly relate to academic outcomes.
Continuous Matching via Vector Field Flow
Authors: Corman Etienne , Ovsjanikov Maks , Chambolle Antonin 
Published: August 2015

AbstractWe present a new method for non‐rigid shape matching designed to enforce continuity of the resulting correspondence. Our method is based on the recently proposed functional map representation, which allows efficient manipulation and inference but often fails to provide a continuous point‐to‐point mapping. We address this problem by exploiting the connection between the operator representation of mappings and flows of vector fields. In particular, starting from an arbitrary continuous map between two surfaces we find an optimal flow that makes the final correspondence operator as close as possible to the initial functional map. Our method also helps to address the symmetric ambiguity problem inherent in many intrinsic correspondence methods when matching symmetric shapes. We provide practical and theoretical results showing that our method can be used to obtain an orientation preserving or reversing map starting from a functional map that represents the mixture of the two. We also show how this method can be used to improve the quality of maps produced by existing shape matching methods, and compare the resulting map's continuity with results obtained by other operator‐based techniques.
Use of endobronchial valve insertion to treat relapsing pneumothorax: a case report and literature review
Authors: Qi Fei , Tian Qing , Chen Liang'an , Li Chunyan , Zhang Shu , Liu Xingchen , Xiao Binbin 
Published: August 2015

AbstractBackgorund and Aims: Unidirectional endobronchial valves have recently been shown to be beneficial as treatment for persistent air leaks. This report presents a first case of endobronchial valve implantation to treat relapsing pneumothorax in a Chinese patient, and also presents a review of the literature on the use of one‐way valve insertion for the treatment of persistent air leaks.Methods: The patient did undergo a recent but failed chest tube intervention. By bronchoscopy and using Chartis® system measurements, the upper left lobe (including the left apical bronchus) was closed using a catheter.Results: After the expected decrease in airflow following bronchial occlusion, increased air pressure and decreased spilled air were noted; it was concluded that the pneumothorax was located in the left upper lobe. A Zephyr® endobronchial valve was placed in the left upper apical bronchus. The health benefits of the procedure were noticed in the following days.Conclusion: Our review suggests that the use of endobronchial valves could be used as an effective, minimally invasive, low‐risk intervention for patients with pneumothorax that cannot be treated surgically.