bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of cis-regulatory elements of the vascular endothelial growth inhibitor gene promoter
Authors: XIAO Qingli , HSU Chung Y. , CHEN Hong , MA Xiucui , XU Jan , LEE Jin-Moo 
Published: June 2005

VEGI (vascular endothelial growth inhibitor), a member of the tumour necrosis factor superfamily, has been reported to inhibit endothelial cell proliferation, angiogenesis and tumour growth. We identified and cloned approx. 2.2 kb of the VEGI promoter from mouse cerebral endothelial cells. The promoter contained an atypical TATA-box-binding protein sequence TAAAAAA residing at −32/−26 relative to the transcription initiation site (+1), 83 bp upstream from the ATG start codon. To investigate critical sequences in the VEGI promoter, a series of deleted and truncated segments were constructed from a 2300 bp promoter construct (−2201/+96) linked to a luciferase reporter gene. Transient transfection of cerebral microvascular cells (bEND.3) and rat C6 glioma cells demonstrated that a 1700 bp deletion from the −2201 to −501 did not significantly affect promoter activity; however, a truncated construct (−501/+96) lacking the region between −312 and −57 resulted in nearly 90% loss of promoter activity. A consensus NF-κB (nuclear factor κB) and several SP1 (specificity protein-1)-binding sequences were identified within the deleted segment. Supershift analysis revealed that NF-κB subunits, p50 and p65, interacted with the VEGI promoter. Exposure of cerebral endothermic cells to the pro-inflammatory cytokine, tumour necrosis factor-α, increased VEGI mRNA levels and DNA-binding activities, whereas an NF-κB inhibitor attenuated this increase. In addition, p65 overexpression enhanced, whereas p50 overexpression decreased, the luciferase activity. Furthermore, mutation of the NF-κB DNA binding site blocked this p65- and tumour necrosis factor-α-induced luciferase activity. These findings suggest that the transcription factor NF-κB plays an important role in the regulation of VEGI expression.
Stabilization of nucleic acids by unusual polyamines produced by an extreme thermophile, Thermus thermophilus
Authors: TERUI Yusuke , OHNUMA Mio , HIRAGA Kaori , KAWASHIMA Etsuko , OSHIMA Tairo 
Published: June 2005

Extreme thermophiles produce two types of unusual polyamine: long linear polyamines such as caldopentamine and caldohexamine, and branched polyamines such as quaternary ammonium compounds [e.g. tetrakis(3-aminopropyl)ammonium]. To clarify the physiological roles of long linear and branched polyamines in thermophiles, we synthesized them chemically and tested their effects on the stability of ds (double-stranded) and ss (single-stranded) DNAs and tRNA in response to thermal denaturation, as measured by differential scanning calorimetry. Linear polyamines stabilized dsDNA in proportion to the number of amino nitrogen atoms within their molecular structure. We used the empirical results to derive formulae that estimate the melting temperature of dsDNA in the presence of polyamines of a particular molecular composition. ssDNA and tRNA were stabilized more effectively by tetrakis(3-aminopropyl)ammonium than any of the other polyamines tested. We propose that long linear polyamines are effective to stabilize DNA, and tetrakis(3-aminopropyl)ammonium plays important roles in stabilizing RNAs in thermophile cells.
Molecular cloning and functional characterization of a high-affinity zinc importer (DrZIP1) from zebrafish (Danio rerio)
Authors: QIU Andong , SHAYEGHI Majid , HOGSTRAND Christer 
Published: June 2005

Zinc is a vital micronutrient to all organisms and a potential toxicant to aquatic animals. It is therefore of importance to understand the mechanism of zinc regulation. In the present study, we molecularly cloned and functionally characterized a zinc transporter of the SLC39A family [commonly referred to as the ZIP (Zrt- and Irt-related protein) family] from the gill of zebrafish (Danio rerio) (DrZIP1). DrZIP1 protein was found to localize at the plasma membrane and to function as a zinc uptake transporter when being expressed in either chinook salmon (Oncorhynchus tshawytscha) embryonic 214 cells or Xenopus laevis oocytes. In comparison with pufferfish transporter proteins (FrZIP2 and FrECaC) that are known to facilitate cellular zinc uptake, DrZIP1 appears to have high affinity to bind and transport zinc, suggesting that it may be a high-affinity zinc uptake transporter (Km&lt;0.5 μM) in fish. Orthologues of DrZIP1 were also identified in both freshwater and seawater pufferfish (Tetraodon nigroviridis and Takifugu rubripes), indicating that these proteins may be functionally conserved among different fish species. DrZIP1 mRNA is expressed in all the tissues examined in the present study and thus DrZIP1 may be a constitutive zinc uptake transporter in many cell types of zebrafish.
Activation of protein synthesis in cardiomyocytes by the hypertrophic agent phenylephrine requires the activation of ERK and involves phosphorylation of tuberous sclerosis complex 2 (TSC2)
Authors: ROLFE Mark , McLEOD Laura E. , PRATT Phillip F. , PROUD Christopher G. 
Published: June 2005

The hypertrophic Gq-protein-coupled receptor agonist PE (phenylephrine) activates protein synthesis. We showed previously that activation of protein synthesis by PE requires MEK [MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase] and mTOR (mammalian target of rapamycin). However, it remained unclear whether ERK activation was required and which downstream components were involved in activating mTOR and protein synthesis. Using an adenovirus encoding the MKP3 (MAPK phosphatase 3) to inhibit ERK activity, we demonstrate that ERK is essential for the activation of protein synthesis by PE. Activation and phosphorylation of S6K1 (ribosomal protein S6 kinase 1) and phosphorylation of eIF4E (eukaryotic initiation factor 4E)-binding protein (both are mTOR targets) were also inhibited by MKP3, suggesting that ERK is also required for the activation of mTOR signalling. PE stimulation of cardiomyocytes induced the phosphorylation of TSC2 (tuberous sclerosis complex 2), a negative regulator of mTOR activity. TSC2 was phosphorylated only weakly at Thr1462, but phosphorylated at additional sites within the sequence RXRXX(S/T). This differs from the phosphorylation induced by insulin, indicating that MEK/ERK signalling targets distinct sites in TSC2. This phosphorylation may be mediated by p90RSK (90 kDa ribosomal protein S6K), which is activated by ERK, and appears to involve phosphorylation at Ser1798. Activation of protein synthesis by PE is partially insensitive to the mTOR inhibitor rapamycin. Inhibition of the MAPK-interacting kinases by CGP57380 decreases the phosphorylation of eIF4E and PE-induced protein synthesis. Moreover, CGP57380+rapamycin inhibited protein synthesis to the same extent as blocking ERK activation, suggesting that MAPK-interacting kinases and regulation of mTOR each contribute to the activation of protein synthesis by PE in cardiomyocytes.
FRAP analysis of nucleocytoplasmic dynamics of the vitamin D receptor splice variant VDRB1: preferential targeting to nuclear speckles
Authors: SUNN Kathryn L. , EISMAN John A. , GARDINER Edith M. , JANS David A. 
Published: June 2005

Although the key components of the cellular nuclear transport machinery have largely been characterized through extensive efforts in recent years, in vivo measurements of the kinetics of nuclear protein import/export are patently few. The present study applies the approach of FRAP (fluorescence recovery after photobleaching) to examine the nucleocytoplasmic flux of a novel human VDRB1 (vitamin D receptor B1) isoform in living cells. Through an N-terminal extension containing a consensus nuclear targeting sequence, VDRB1 is capable of localizing in nuclear speckles adjacent to SC-35 (35 kDa splicing component)-containing speckles as well as in the nucleoplasm, dependent on ligand. Investigation of VDRB1 nucleocytoplasmic transport using FRAP indicates for the first time that the VDRB1 has a serum-modulated, active nuclear import mechanism. There is no evidence of an efficient, active export mechanism for VDRB1, probably as a result of nuclear retention. VDRB1 nuclear import in the absence of serum occurred more rapidly and to a greater extent to nuclear speckles compared with import to other nuclear sites. This preferential transport from the cytoplasm to and accumulation within nuclear speckles is consistent with the idea that the latter represent dynamic centres of VDRB1 interaction with other nuclear proteins. The results are consistent with the existence of specialized pathways to target proteins to nuclear subdomains.
Candidacidal effects of two antimicrobial peptides: histatin 5 causes small membrane defects, but LL-37 causes massive disruption of the cell membrane
Authors: den HERTOG Alice L. , van MARLE Jan , van VEEN Henk A. , van't HOF Wim , BOLSCHER Jan G. M. , VEERMAN Enno C. I. , NIEUW AMERONGEN Arie V. 
Published: June 2005

The effects of antimicrobial peptides on artificial membranes have been well-documented; however, reports on the ultrastructural effects on the membranes of micro-organisms are relatively scarce. We compared the effects of histatin 5 and LL-37, two antimicrobial peptides present in human saliva, on the functional and morphological properties of the Candida albicans cell membrane. Fluorescence microscopy and immunogold transmission electron microscopy revealed that LL-37 remained associated with the cell wall and cell membrane, whereas histatin 5 transmigrated over the membrane and accumulated intracellularly. Freeze-fracture electron microscopy revealed that LL-37 severely affected the membrane morphology, resulting in the disintegration of the membrane bilayer into discrete vesicles, and an instantaneous efflux of small molecules such as ATP as well as larger molecules such as proteins with molecular masses up to 40 kDa. The effects of histatin 5 on the membrane morphology were less pronounced, but still resulted in the efflux of nucleotides. As the morphological defects induced by histatin 5 are much smaller than those induced by LL-37, but the efflux of nucleotides is similar at comparable candidacidal concentrations, we suggest that the loss of nucleotides plays an important role in the killing process.
Rgt1, a glucose sensing transcription factor, is required for transcriptional repression of the HXK2 gene in Saccharomyces cerevisiae
Authors: PALOMINO Aaron , HERRERO Pilar , MORENO Fernando 
Published: June 2005

Expression of HXK2, a gene encoding a Saccharomyces cerevisiae bifunctional protein with catalytic and regulatory functions, is controlled by glucose availability, being activated in the presence of glucose and inhibited when the levels of the sugar are low. In the present study, we identified Rgt1 as a transcription factor that, together with the Med8 protein, is essential for repression of the HXK2 gene in the absence of glucose. Rgt1 represses HXK2 expression by binding specifically to the motif (CGGAAAA) located at −395 bp relative to the ATG translation start codon in the HXK2 promoter. Disruption of the RGT1 gene causes an 18-fold increase in the level of HXK2 transcript in the absence of glucose. Rgt1 binds to the RGT1 element of HXK2 promoter in a glucose-dependent manner, and the repression of target gene depends on binding of Rgt1 to DNA. The physiological significance of the connection between two glucose-signalling pathways, the Snf3/Rgt2 that causes glucose induction and the Mig1/Hxk2 that causes glucose repression, was also analysed.
Guanine-specific DNA damage induced by γ-irradiated histone
Authors: FURUKAWA Ayako , HIRAKU Yusuke , OIKAWA Shinji , LUXFORD Catherine , DAVIES Michael J. , KAWANISHI Shosuke 
Published: June 2005

In γ-irradiation, •OH is directly generated from water and causes DNA damage leading to carcinogenesis. Exposure of proteins to γ-irradiation, in the presence of oxygen, gives high yields of hydroperoxides. To clarify whether these hydroperoxides, particularly those formed on DNA-binding histone proteins, participate in γ-irradiation-induced carcinogenesis, experiments using 32P-labelled DNA fragments obtained from human cancer-related genes were undertaken. Histone protein-hydroperoxides induced significant DNA damage in the presence of Cu(I). Histone H1- and H3-hydroperoxides showed stronger DNA damage compared with histone H2A- and H4-hydroperoxides at 0.7 μM. Histone H1-hydroperoxides caused Cu(I)-dependent DNA damage predominantly at guanine residues, especially at 5′-GGC-3′, 5′-GGA-3′, 5′-GGT-3′ and single G bases. In contrast, histone H3-hydroperoxides/Cu(I) induced DNA damage at 5′-G in GG sequences; this sequence specificity is identical with that generated by 2,2′-azobis (2-amidinopropane) dihydrochloride, which is known to produce peroxyl radicals (RO2•). The difference in site specificity of DNA damage induced by histone H1- and H3-hydroperoxides may arise from their amino acid composition or their mode of binding to DNA. The histone H1-hydroperoxides/Cu(I) system also induced 8-oxo-7,8-dihydro-2′-deoxyguanosine formation in calf thymus DNA. It is concluded that histone protein-hydroperoxides can induce guanine-specific DNA damage, which may contribute to γ-irradiation-induced carcinogenesis.
Molecular cloning and characterization of a novel β-1,3-xylanase possessing two putative carbohydrate-binding modules from a marine bacterium Vibrio sp. strain AX-4
Authors: KIYOHARA Masashi , SAKAGUCHI Keishi , YAMAGUCHI Kuniko , ARAKI Toshiyoshi , NAKAMURA Takashi , ITO Makoto 
Published: June 2005

We cloned a novel β-1,3-xylanase gene, consisting of a 1728-bp open reading frame encoding 576 amino acid residues, from a marine bacterium, Vibrio sp. strain AX-4. Sequence analysis revealed that the β-1,3-xylanase is a modular enzyme composed of a putative catalytic module belonging to glycoside hydrolase family 26 and two putative carbohydrate-binding modules belonging to family 31. The recombinant enzyme hydrolysed β-1,3-xylan to yield xylo-oligosaccharides with different numbers of xylose units, mainly xylobiose, xylotriose and xylotetraose. However, the enzyme did not hydrolyse β-1,4-xylan, β-1,4-mannan, β-1,4-glucan, β-1,3-xylobiose or p-nitrophenyl-β-xyloside. When β-1,3-xylo-oligosaccharides were used as the substrate, the kcat value of the enzyme for xylopentaose was found to be 40 times higher than that for xylotetraose, and xylotriose was extremely resistant to hydrolysis by the enzyme. A PSI-BLAST search revealed two possible catalytic Glu residues (Glu-138 as an acid/base catalyst and Glu-234 as a nucleophile), both of which are generally conserved in glycoside hydrolase superfamily A. Replacement of these two conserved Glu residues with Asp and Gln resulted in a significant decrease and complete loss of enzyme activity respectively, without a change in their CD spectra, suggesting that these Glu residues are the catalytic residues of β-1,3-xylanase. The present study also clearly shows that the non-catalytic putative carbohydrate-binding modules play an important role in the hydrolysis of insoluble β-1,3-xylan, but not that of soluble glycol-β-1,3-xylan. Furthermore, repeating a putative carbohydrate-binding module strongly enhanced the hydrolysis of the insoluble substrate.
Simultaneous Dual Fast and Slow Pathway Conduction upon Induction of Typical Atrioventricular Nodal Reentrant Tachycardia: Electrophysiologic Characteristics in a Series of Patients
Authors: TOMASI CORRADO , DE PONTI ROBERTO , TRITTO MASSIMO , BARILLI ANGELA LUCIANA , BOTTONI NICOLA , ZARDINI MARCO , MENOZZI CARLO , SPADACINI GIAMMARIO , SALERNO‐URIARTE JORGE ANTONIO 
Published: June 2005

Introduction: Simultaneous dual atrioventricular nodal conduction (SDNC) through slow (SP) and fast pathway (FP) is a rare phenomenon observed upon the induction of atrioventricular nodal reciprocating tachycardia (AVNRT). The aim of this study is to report the electrophysiological features of patients showing typical AVNRT induced through SDNC. Methods and Results: Among 461 consecutive patients with typical AVNRT submitted to radiofrequency catheter ablation (RFCA), seven patients (1.5%) with SDNC at tachycardia onset (group I: 6 female; age 60–72 years, mean 65.2 ± 3.8 years) and 118 age‐matched controls (group II: 60 female; age 60–88 years, mean 68.4 ± 6.8 years) were considered. Controls were further subdivided into two subgroups according to age: subgroup A (94 patients, age 60–75 years) and subgroup B (24 patients, age >75 years). The value of the following parameters was significantly higher in group I than in group II and in subgroup A: A–H interval [113 ± 26 vs. 89 ± 27 (P < 0.01) vs. 84 ± 19 (P < 0.001)], ventriculoatrial conduction effective refractory period [355 ± 85 vs. 293 ± 87 (P < 0.05) vs. 281 ± 82 (P < 0.05)], SP conduction time upon AVNRT induction [444 ± 104 vs. 350 ± 72 (P < 0.01); vs. 345 ± 67 (P < 0.001)], AVNRT cycle length [484 ± 103 vs. 396 ± 71 ms (P < 0.05); vs. 384 ± 69 (P < 0.05)], and rate of AVNRT induction from ventricle [71% vs. 10% (P = 0.001); vs. 6% (P = 0.001)]. Differences were mostly not significant between group I and subgroup B. SP location and RFCA success rate were similar in all groups. Conclusion: In a population of AVNRT patients, SDNC at AVNRT induction is infrequent and it prevails beyond the fifth decade of life and in females. SDNC is associated with peculiar AVN conduction features, which resemble the age‐related modifications of AVN conduction.
Ventricular Arrhythmias Following Exposure of Failing Hearts to Oxidative Stress in Vitro
Authors: BRIGADEAU FRANÇOIS , GELÉ PATRICK , MARQUIÉ CHRISTELLE , SOUDAN BENOÎT , LACROIX DOMINIQUE 
Published: June 2005

Introduction: There is experimental evidence that heart failure (HF) is an oxidative stress and that HF myocytes may be damaged by oxygen‐derived free radicals. However, the arrhythmogenicity of these radicals has not been studied in HF. Methods and Results: Isolated perfused hearts were obtained from sham‐operated (SHAM, n = 6), and fast pacing (250 ms, 2 weeks)‐induced heart failure porcines (HF, n = 8). Epicardial conduction was mapped in the longitudinal and transverse directions and ventricular arrhythmias were closely monitored after perfusion of 100, 300, and 1000 μmol/L H2O2. Left ventricular epicardium was sampled for action potentials recordings in the same conditions. Myocardial levels of thiobarbituric acid reactive substances and antioxidant enzymatic capacity were also assessed. Epicardial conduction velocities were unaffected by H2O2 in both groups. Isolated ventricular premature beats and runs of slow ventricular rhythm with H2O2 more frequently occurred in HF compared to SHAM despite an increased antioxidant capacity including Cu/Zn and Mn superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase. Sustained arrhythmias were not observed. Higher thiobarbituric acid reactive substances levels were found in HF confirming endogenous oxidative stress. Action potential duration at plateau level was increased following H2O2 in SHAM but not in HF epicardial fibers where a toxic effect developed at 1000 μmol/L. Conclusion: Oxidative stress with concomitant increase in antioxidant capacity develops in this HF model. There is a greater proclivity to oxidative stress‐mediated arrhythmias in HF. These arrhythmias are mainly extrasystoles or slow ventricular rhythms and not dependent on abnormal myocardial conduction.
Changes of Pulmonary Vein Orifice Size and Location throughout the Cardiac Cycle: Dynamic Analysis Using Magnetic Resonance Cine Imaging
Authors: LICKFETT LARS , DICKFELD TIMM , KATO RITSUSHI , TANDRI HARIKRISHNA , VASAMREDDY CHANDRASEKHAR R. , BERGER RONALD , BLUEMKE DAVID , LÜDERITZ BERNDT , HALPERIN HENRY , CALKINS HUGH 
Published: June 2005

Introduction: Anatomically guided left atrial ablation is used increasingly for treatment of atrial fibrillation (AF). Three‐dimensional mapping systems used for pulmonary veins (PV) encircling ablation procedures anticipate a stable size and position of the PV orifice. The aim of the current study was therefore to analyze changes of PV orifice size and location throughout the cardiac cycle using cine magnetic resonance imaging (MRI). Methods and Results: Twenty‐five healthy volunteers were studied using a 1.5 Tesla MRI system. MR angiograms were acquired with a breath‐hold three‐dimensional fast‐spoiled gradient‐echo imaging (3D FSPGR) sequence in the coronal plane before and after gadolinium injection. Maximum intensity projections and multiplanar reformations were performed to reconstruct images of the PV. Bright blood cine imaging in the axial view was acquired by a steady‐state free precession pulse sequence. Twenty bright blood images were obtained per cardiac cycle. The axial (anterior‐posterior) PV orifice diameter was measured in all 20 images. For analysis of PV movement the location of the orifice posterior edge was plotted on scale paper.PV orifice size depends on the stage of the cardiac cycle with the largest diameter in late atrial diastole and a mean decrease of 32.5% during atrial systole. Location changes of the PV orifice are in the range of up to 7.2 mm and larger in the coronal (lateral‐medial) than in the sagittal (anterior‐posterior) direction. Conclusion: PV orifice size and location is not as stable as anticipated by three‐dimensional mapping systems used for PV encircling left atrial ablation procedures. RF application close to the presumed orifice location should therefore be avoided to minimize the risk of PV stenosis.
The degradation of lignin and quantitative aspects of ruminant digestion
Authors: Porter P. , Singleton A. G. 
Published: June 2005

1. The technique for the quantitative estimation of lignin by digestion with 72% H2S04has been investigated. Two methods of pretreatment designed to eliminate interference products of carbohydrates and proteins were compared, namely (a) acid and pepsin and (b) acid detergent. The latter technique gave low lignin yields owing to the colloidal dispersion of lignin in the acid detergent treatment. Erroneous results arise from an inadequate digestion in 72% H2SO4. It was found preferable to filter the product lignin without prior dilution of the acid since white particles of material, assumed to be cellulose, flocculated from solution on dilution. Also there is some loss due to colloidal dispersion of lignin during refluxing in dilute acid.2. Analysis of soda lignin and sulphuric acid lignin preparations from hay, duodenal contents and faeces of sheep with exteriorized flow through a re-entrant duodenal fistula provided evidence for the degradation of lignin by digestive processes. Demethoxylation of lignin occurred mainly in the stomach, suggesting that this was the main site of breakdown.3. A new displacement method of measuring duodenal flow is described. Quantitative results obtained also indicate that lignin is degraded mainly in the stomach to the extent of about 10%.4. These results suggest that faecal lignin is an adequate marker for the estimation of duodenal flow, and hence of the extent of changes in the stomach and intestines respectively.