bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ara‐C‐ and daunorubicin‐induced recruitment of Lyn in sphingomyelinase‐enriched membrane rafts
Authors: Grazide Solène , Maestre Nicolas , Veldman Robert Jan , Bezombes Christine , Maddens Stéphane , Levade Thierry , Laurent Guy , Jaffrézou Jean‐pierre 
Published: October 2002

ABSTRACTInduction of apoptosis by DNA‐damaging agents such as 1‐β‐D‐arabinofuranosylcytosine (Ara‐C) includes the activation of Lyn protein tyrosine kinase. We have previously established that Ara‐C‐induced activation of Lyn results in its binding to a neutral sphingomyelinase (SMase) and is requisite for its stimulation and the induction of apoptosis in U937 cells. However, the spacio‐temporal organization of these events is unclear. This study demonstrates that part of the total cellular SMase activity is sequestered in sphingomyelin‐enriched plasma membrane microdomains (rafts). Under Ara‐C and daunorubicin (DNR) treatment, Lyn is rapidly activated and translocated into rafts. The compartmentalization of Lyn (as well as neutral SMase activation and apoptosis) induced by these drugs was blocked by the tyrosine kinase inhibitor herbimycin A and raft disruption. In conclusion, this study establishes that DNA‐damaging agents such as Ara‐C and DNR rapidly induce Lyn activation and its translocation into membrane rafts. This in turn leads to neutral SMase activation and raft‐associated sphingomyelin hydrolysis with the concomitant generation of the proapoptotic lipid second messenger, ceramide. The apparent topological partitioning between DNA damage and apoptosis signaling (integrated into specialized plasma membrane domains) is discussed.
Crystal structures of leukotriene A4 hydrolase in complex with captopril and two competitive tight‐binding inhibitors
Authors: Thunnissen Marjolein M.G.M. , Andersson Björn , Samuelsson Bengt , Wong Chi‐huey , Haeggström Jesper Z. 
Published: October 2002

ABSTRACTLeukotriene (LT) A4 hydrolase/aminopeptidase is a bifunctional zinc enzyme that catalyzes the final step in the biosynthesis of LTB4, a potent chemoattractant and immune modulating lipid mediator. Here, we report a high‐resolution crystal structure of LTA4 hydrolase in complex with captopril, a classical inhibitor of the zinc peptidase angiotensin‐converting enzyme. Captopril makes few interactions with the protein, but its free thiol group is bound to the zinc, apparently accounting for most of its inhibitory action on LTA4 hydrolase. In addition, we have determined the structures of LTA4 hydrolase in complex with two selective tight‐binding inhibitors, a thioamine and a hydroxamic acid. Their common benzyloxyphenyl tail, designed to mimic the carbon backbone of LTA4, binds into a narrow hydrophobic cavity in the protein. The free hydroxyl group of the hydroxamic acid makes a suboptimal, monodentate complex with the zinc, and strategies for improved inhibitor design can be deduced from the structure. Taken together, the three crystal structures provide the molecular basis for the divergent pharmacological profiles of LTA4 hydrolase inhibitors. Moreover, they help define the binding pocket for the fatty acid‐derived epoxide LTA4 as well as the subsites for a tripeptide substrate, which in turn have important implications for the molecular mechanisms of enzyme catalyses.
Negative regulatory role of overexpression of PLCγ1in the expression of early growth response 1 gene in rat 3Y1 fibroblasts
Authors: Shin Soon Young , Ko Jesang , Chang Jong‐Soo , Min Do Sik , Choi Chan , Bae Sun Sik , Kim Myung Jong , Hyun Dae Sung , Kim Jung‐Hye , Han Mi Young , Kim Young‐Ho , Kim Yong Sik , Na Doe Sun , Suh Pann‐Ghill , Lee Young Han 
Published: October 2002

ABSTRACTThe early growth response 1 (Egr‐1) gene product is a transcription factor that functions as an oikis factor. Loss of Egr‐1 expression is closely associated with tumor formation. Phospholipase Cγ1 (PLCγ1) is overexpressed in some tumors, and its overexpression causes anchorage‐independent growth. Here we report that overexpression of PLCγ1 and SH2‐SH3 domain of PLCγ1 decreased induction of Egr‐1 and the Egr‐1‐regulated genes TSP‐1 and PAI‐1. Results from the nuclear run‐on assay and transfection experiment with the proximal 455 base pair region of the Egr‐1 promoter (‐454 to +1) showed that Egr‐1 transcriptional activity was suppressed in PLCγ1–3Y1 cells whereas decay of Egr‐1 mRNA was similar in both cell lines. Serum response element‐and ternary complex factor Elk‐1‐mediated transcriptional activation of the reporter gene in response to EGF were also inhibited in PLCγ1–3Y1 cells. Pretreatment with the protein synthesis inhibitor cycloheximide (CHX) partially abrogated the serum‐induced suppression of Egr‐1 transcription in PLCγ1–3Y1 cells, suggesting that a CHX‐sensitive factor(s) is involved in the suppression of Egr‐1 transcription in PLCγ1–3Y1 cells. Our results demonstrated that overexpression of PLCγ1 functions as a negative modulator of the tumor suppressor Egr‐1 gene expression, possibly through inhibition of Elk‐1‐dependent transcriptional activity.—Shin, S. Y., Ko, J., Chang, J.‐S., Min, D. S., Choi, C., Bae, S. S., Kim, M. J., Hyun, D. S., Kim, J.‐H., Han, M. Y., Kim, Y.‐H., Kim, Y. S., Na, D. S., Suh, P.‐G., Lee, Y. H. Negative regulatory role of overexpression of PLCγ1 in the expression of early growth response 1 gene in rat 3Y1 fibroblasts. FASEB J. 16, 1504–1514 (2002)
Fatty acid augmentation of the cardiac slowly activating delayed rectifier current (IKs) is conferred by hminK
Authors: Doolan Gavin K. , Panchal Rekha G. , Fonnes Eva L. , Clarke Alison L. , Williams David A. , Petrou Steven 
Published: October 2002

ABSTRACTThe mechanism by which dietary fatty acids confer protection against cardiac arrhythmias and sudden cardiac death is not resolved. Here, we study the effects of several known cardioprotective and arrhythmogenic fatty acids on the slowly activating delayed rectifier potassium current (IKs), which is responsible for the repolarization phase of the cardiac action potential. cRNAs encoding either or both of the two subunits, KvLQT1 and hminK, that together produce IKs, were injected into Xenopus oocytes, and the effects of various fatty acids were determined. Docosahexaenoic acid (DHA) significantly augmented IKs as did the short‐chained fully saturated lauric acid, and to a lesser extent the cis‐unsaturated oleic acid. Eicosapentaenoic acid (EPA) was without significant effect on current magnitude, although it reduced the rate of activation. These results suggest that not all “antiarrhythmic” fatty acids target the same channel. To examine the role of hminK in this response, KvLQT1 was expressed alone. In this case, DHA, lauric acid, and oleic acid did not augment current, suggesting that hminK confers fatty acid sensitivity to IKs.
Nitric oxide promotes intracellular calcium release from mitochondria in striatal neurons
Authors: Horn Thomas F. W. , Wolf Gerald , Duffy Steven , Weiss Samuel , Keilhoff Gerburg , MacVicar Brian A. 
Published: October 2002

ABSTRACTOverproduction of nitric oxide by NMDA receptor stimulation is implicated in calcium deregulation and neurodegeneration of striatal neurons. We investigated the involvement of nitric oxide (NO) in inducing intracellular calcium release and in modifying calcium transients evoked by NMDA. NO application (4–10 fM) reversibly and repeatedly increased the intracellular calcium concentration [Ca2+]i in Fura‐2‐or fluo‐3‐loaded cultured mouse striatal neurons. NO‐induced [Ca2+]i responses persisted in the absence of extracellular calcium, indicating that Ca2+ was released from intracellular stores. The source of calcium was distinct from [Ca2+]i‐activated (ruthenium red and ryanodine sensitive) or IP3‐activated (thapsigargin‐sensitive) Ca2+ stores and was not dependent on cGMP production because a cell permeant analog, 8‐bromo‐cGMP, did not increase basal [Ca2+]i. Glucose removal potentiated the NO‐induced release of [Ca2+]i. In contrast, pretreatment with either the mitochondrial uncoupler carbonyl cyanide m‐chlorophenylhydrazone or cyclosporin A, a blocker of the mitochondrial permeability transition pore, prevented the [Ca2+]i increase after NO. The rise in [Ca2+]i during NO exposure was preceded by a decrease in mitochondrial membrane potential that was partly reversible during washout. Repeated applications of NMDA induced irreversible [Ca2+]i responses in a subpopulation of striatal cells that were greatly reduced by the NOS inhibitor NW‐nitro‐L‐arginine. Calcium transients were prolonged by conjoint application of NMDA and NO. We conclude that NMDA‐evoked [Ca2+]i transients are modulated by endogenous NO production, which leads to release of calcium from the mitochondrial pool. An NO‐activated mitochondrial permeability transition pore may lead to cell death after overstimulation of NMDA receptors.—Horn, T. F. W., Wolf, G., Duffy, S., Weiss, S., Keilhoff, G., MacVicar, B. A. Nitric oxide promotes intracellular calcium release from mitochondria in striatal neurons. FASEB J. 16, 1611–1622 (2002)
The PDZ‐scaffold protein INAD abolishes apparent store‐dependent regulation of the light‐activated cation channel TRP
Authors: Harteneck Christian , Kuchta Sabina Naranjo , Huber Armin , Paulsen Reinhard , Schultz Günter 
Published: October 2002

ABSTRACTIn fly photoreceptor cells, light initiates a G protein‐coupled phospholipase Cβ‐dependent signaling cascade that results in the depolarization of the cell membrane, which is mediated by the cation channels TRP and TRPL. Together with phospholipase Cβ and an eye‐specific protein kinase C, TRP is tethered to the scaffolding protein INAD, which forms a multimolecular signaling complex. Divergent data from expressed TRP and studies from photoreceptor cells have brought up a controversy whether or not a capacitative calcium entry (CCE) mechanism is involved in the Drosophila phototransduction pathway. Our initial characterization of TRP from photoreceptors of Calliphora vicina supported the hypothesis of a CCE mechanism, as heterologously expressed TRP was stimulated after application of thapsigargin. The situation changed when the PDZ domain protein INAD was coexpressed with TRP. In cells coexpressing TRP and INAD, no calcium entry was detectable on application of store depletion protocols. Suppression of CCE by INAD was not observed when the described interaction was disrupted by mutations in TRP and INAD. Our data show that apparent activation of TRP by CCE is abolished by INAD. Within the complex, the proteins necessary for phototransduction mutually influence their activities. The results support the hypothesis of a store‐independent activation of TRP.
Severely reduced neutrophil adhesion and impaired host defense against fecal and commensal bacteria in CD18–/–P‐selectin–/–double null mice
Authors: Forlow Stephen Bradley , Foley Patricia L. , Ley Klaus 
Published: October 2002

ABSTRACTLeukocyte recruitment to sites of inflammation requires the functions of selectins and integrins. P‐selectin null (CD62P–/–) mice show a mild and CD18 null (CD18–/–) mice a more severe neutrophil recruitment defect in some inflammatory models. To investigate the possible cooperative interactions between CD18 integrins and P‐selectin in mediating neutrophil recruitment, we generated CD18–/–CD62P–/–double null mice. CD18–/–CD62P–/–mice were apparently normal at weaning and fertile but later failed to gain weight, showed increased susceptibility to infection by fecal and commensal bacteria, and survived only 5–6 months. Some CD18–/–CD62P–/–mice showed severe spontaneous skin lesions; most showed neutrophil infiltration in the lungs and liver, and positive bacterial cultures from internal organs. The number and velocity of rolling leukocytes in tumor necrosis factor a treated venules of CD18–/–CD62P–/–mice was similar to those in wild‐type mice, but neutrophil adhesion was severely reduced. Only 25% of adhered leukocytes were neutrophils in CD18–/–CD62P‐/mice vs. >90% in wild‐type, CD62P–/–, and CD18–/–single mutants. Our data show that removing both P‐selectin and CD18 integrins from mice leads to severe neutrophil recruitment defects and spontaneous pathology.—Forlow, S. B., Foley, P. L., Ley, K. Severely reduced neutrophil adhesion and impaired host defense against fecal and commensal bacteria in CD18–/–P‐selectin–/–double null mice. FASEB J. 16, 1488–1496 (2002)
Augmentation of cardiac contractility with no change in L‐type Ca2+ current in transgenic mice with a cardiac‐directed expression of the human adenylyl cyclase type 8 (AC8)
Authors: Georget Marie , Mateo Philippe , Vandecasteele Grégoire , Jurevičius Jonas , Lipskaia Larissa , Defer Nicole , Hanoune Jacques , Hoerter Jacqueline , Fischmeister Rodolphe 
Published: October 2002

ABSTRACTThe β‐adrenergic cascade is severely impaired in heart failure (HF), in part because of a reduction in the activity of the two dominant cardiac adenylyl cyclase (AC) isoforms, AC5 and AC6. Hence, cardiac‐directed AC overexpression is a conceivable therapeutic strategy in HF. In this study, we explored the consequences at the cellular and organ level of a cardiac‐directed expression of the human AC8 in the transgenic mouse line AC8TG. Unlike AC5 and AC6, which are inhibited by intracellular Ca2+, AC8 is stimulated by Ca2+‐calmodulin. Langendorff perfused hearts from AC8TG mice had a twofold higher left ventricular systolic pressure, a 40% faster heart rate, a 37% faster relaxation, and a 30% higher sensitivity to external Ca2+ than nontransgenic control mice (NTG). Cell shortening measured in isolated ventricular myocytes developed 22% faster and relaxed 43% faster in AC8TG than in NTG mice. Likewise, Ca2+ transients measured in fluo‐3 AM‐loaded myocytes were 30% higher and relaxed 24% faster in AC8TG compared with NTG mice. In spite of the large increase in Ca2+ transients and contraction, expression of AC8 had no effect on the whole‐cell L‐type Ca2+ current (ICa,L) amplitude. Moreover, ICa,L was unchanged even when AC8 was activated by raising intracellular Ca2+. Thus, cardiac expression of AC8 leads to an increase in cAMP that activates specifically Ca2+ uptake into the sarcoplasmic reticulum but not Ca2+ influx at the sarcolemma, suggesting a strong compartmentation of the cAMP signal.
RETRACTED: NCX‐4016, a nitric oxide‐releasing aspirin, protects endothelial cells against apoptosis by modulating mitochondrial function
Authors: Fiorucci Stefano , Mencarelli Andrea , Mannucci Roberta , Distrutti Eleonora , Morelli Antonio , del Soldato Piero , Moncada Salvador 
Published: October 2002

ABSTRACTWe investigated the effect of a nitric oxide (NO)‐releasing derivative of aspirin (NCX‐4016) on a mitochondria‐dependent model of apoptosis in human umbilical endothelial cells (HUVEC). Exposure of HUVEC to staurosporine caused a progressive fall in mitochondrial membrane potential (∆ψm) and apoptosis. Exposure to an NO donor, (z)‐1‐[2‐(2‐aminoethyl)‐N‐(2ammonioethyl)amino]diazen‐1‐ium‐1,2‐diolate (DETA‐NO), caused an early (1–3h) hyperpolarization of ∆ψm and reduction of apoptosis that was followed (at 4–8 h) by an accelerated collapse of ∆ψm and cell death. In contrast, treatment with NCX‐4016, but not with aspirin or a non‐NO‐releasing analogue of NCX‐4016, protected HUVEC against the apoptotic actions of staurosporine for up to 8 h. Confocal microscopy demonstrated that although NCX4016 released NO in subcellular compartments, DETA‐NO caused a generalized increase in cytosolic fluorescence. Exposure to DETA‐NO resulted in a rapid and profound inhibition of cell respiration (78.3 ± 6.4%), whereas NCX‐4016 caused a less pronounced reduction in oxygen consumption (43.5 ± 5.3%). Staurosporine caused a time‐dependent activation of proapoptotic caspases. NCX‐4016 prevented this activation, whereas DETA‐NO failed to inhibit caspase activity. In contrast to DETA‐NO, NCX‐4016 did not increase mitochondrial oxidative stress. These data demonstrated that NCX‐4016 conveys NO directly inside endothelial cells and modulates mitochondrial function.
Angiogenesis stimulated by PDGF‐CC, a novel member in the PDGF family, involves activation of PDGFR‐aa and ‐ap receptors
Authors: Cao Renhai , Bråkenhielm Ebba , Li Xuri , Pietras Kristian , Widenfalk Johan , Östman Arne , Eriksson Ulf , Cao Yihai 
Published: October 2002

ABSTRACTA newly discovered PDGF isoform, PDGF‐CC, is expressed in actively angiogenic tissues such as placenta, some embryonic tissues, and tumors. We test the possibility that PDGF‐CC promotes angiogenesis in vivo. The core domain (mature form) of human PDGF‐CC is sufficiently potent to stimulate neovascularization in the mouse cornea. The corneal angiogenic response induced by PDGF‐CC is robust although the area of neovascularization is smaller than those of FGF‐2‐and VEGF‐stimulated angiogenesis. Similarly, PDGF‐BB and PDGF‐AB induce angiogenic responses virtually indistinguishable from PDGF‐CCstimulated vessels. In contrast, PDGF‐AA displays only a weak angiogenic response in the mouse cornea. Although there was no significant difference in incorporation of mural cells to the newly formed blood vessels induced by PDGF‐BB and ‐CC, the percentage of mural cell positive vessels induced by PDGF‐AA was greater than those induced by FGF‐2, PDGF‐BB, and PDGF‐CC. In the developing chick embryo, PDGF‐CC induced branch sprouts from established blood vessels. In PDGF receptor‐transfected endothelial cells, PDGF‐CC activated the PDGF receptor alpha subunit (PDGFR‐a). PDGF‐CC, but not PDGF‐AA, was able to activate PDGFR‐p receptor in endothelial cells that coexpress both α and β forms of receptors. Thus, the PDGF‐CC‐mediated angiogenic response is most likely transduced by PDGF‐aa and ‐ap receptors. These data demonstrate that the PDGF family is a complex and important group of proangiogenic factors.—Cao, R., Bråkenhielm, E., Li, X., Pietras, K., Widenfalk, J, Östman, A., Eriksson, U., Cao, Y. Angiogenesis stimulated by PDGF‐CC, a novel member in the PDGF family, involves activation of PDGFR‐aa and ‐ap receptors. FASEB J. 16, 1575–1583 (2002)
Truncation Analysis of TatA and TatB Defines the Minimal Functional Units Required for Protein Translocation
Authors: Lee Philip A. , Buchanan Grant , Stanley Nicola R. , Berks Ben C. , Palmer Tracy 
Published: October 2002

ABSTRACT

The TatA and TatB proteins are essential components of the twin arginine protein translocation pathway in
Escherichia coli
. C-terminal truncation analysis of the TatA protein revealed that a plasmid-expressed TatA protein shortened by 40 amino acids is still fully competent to support protein translocation. Similar truncation analysis of TatB indicated that the final 30 residues of TatB are dispensable for function. Further deletion experiments with TatB indicated that removal of even 70 residues from its C terminus still allowed significant transport. These results imply that the transmembrane and amphipathic helical regions of TatA and TatB are critical for their function but that the C-terminal domains are not essential for Tat transport activity. A chimeric protein comprising the N-terminal region of TatA fused to the amphipathic and C-terminal domains of TatB supports a low level of Tat activity in a strain in which the wild-type copy of either
tatA
or
tatB
(but not both) is deleted.
Evidence for a Type III Secretion System in
Aeromonas salmonicida
subsp.
salmonicida
Authors: Burr Sarah E. , Stuber Katja , Wahli Thomas , Frey Joachim 
Published: October 2002

ABSTRACT

Aeromonas salmonicida
subsp.
salmonicida
, the etiological agent of furunculosis, is an important fish pathogen. We have screened this bacterium with a broad-host-range probe directed against
yscV
, the gene that encodes the archetype of a highly conserved family of inner membrane proteins found in every known type III secretion system. This has led to the identification of seven open reading frames that encode homologues to proteins functioning within the type III secretion systems of
Yersinia
species. Six of these proteins are encoded by genes comprising a
virA
operon. The
A. salmonicida
subsp.
salmonicida yscV
homologue,
ascV
, was inactivated by marker replacement mutagenesis and used to generate an isogenic
ascV
mutant. Comparison of the extracellular protein profiles from the
ascV
mutant and the wild-type strain indicates that
A. salmonicida
subsp.
salmonicida
secretes proteins via a type III secretion system. The recently identified ADP-ribosylating toxin AexT was identified as one such protein. Finally, we have compared the toxicities of the wild-type
A. salmonicida
subsp.
salmonicida
strain and the
ascV
mutant against RTG-2 rainbow trout gonad cells. While infection with the wild-type strain results in significant morphological changes, including cell rounding, infection with the
ascV
mutant has no toxic effect, indicating that the type III secretion system we have identified plays an important role in the virulence of this pathogen.
A Polysaccharide Deacetylase Gene (
pdaA
) Is Required for Germination and for Production of Muramic δ-Lactam Residues in the Spore Cortex of
Bacillus subtilis
Authors: Fukushima Tatsuya , Yamamoto Hiroki , Atrih Abdelmadjid , Foster Simon J. , Sekiguchi Junichi 
Published: October 2002

ABSTRACT

The predicted amino acid sequence of
Bacillus subtilis yfjS
(renamed
pdaA
) exhibits high similarity to those of several polysaccharide deacetylases. β-Galactosidase fusion experiments and results of Northern hybridization with sporulation sigma mutants indicated that the
pdaA
gene is transcribed by Eσ
G
RNA polymerase.
pdaA
-deficient spores were bright by phase-contrast microscopy, and the spores were induced to germination on the addition of
l
-alanine. Germination-associated spore darkening, a slow and partial decrease in absorbance, and slightly lower dipicolinic acid release compared with that by the wild-type strain were observed. In particular, the release of hexosamine-containing materials was lacking in the
pdaA
mutant. Muropeptide analysis indicated that the
pdaA
-deficient spores completely lacked muramic δ-lactam. A
pdaA-gfp
fusion protein constructed in strain 168 and
pdaA
-deficient strains indicated that the protein is localized in
B. subtilis
spores. The biosynthetic pathway of muramic δ-lactam is discussed.