bluuebunny/crossref_metadata_2025_split huggingface dataset:
Constructing ‘Farmer’ and ‘State’ Identities in Moral Discourses about Semi-subsistence Agriculture in North-east Brazil
Authors: PENNESI KAREN 
Published: August 2015

AbstractAnthropological analysis elucidates how discourses about agriculture in one North-east Brazilian community reflect relational roles of citizens and the state, the position of farmers in society, and the relationship of individuals to their work. In these discourses, farmers are positioned as moral, hard-working, autonomous citizens, justifying their participation in low-paying activities. The declining numbers of agricultural workers is explained as a result of individual laziness or government irresponsibility. In using these discourses to take stances publicly on agricultural issues, speakers assign responsibilities and moral status to agents. In constructing rural identities, such moral discourses emphasise the symbolic value of subsistence agriculture as its economic value declines.
‘Hands off my constitution’: Constitutional reform and the workings of democracy in Mali
Authors: Wing Susanna D. 
Published: August 2015

ABSTRACTConstitutions in Africa are often considered fragile. Mali is an example of both constitutional crisis and constitutional durability. While Mali was upheld as a democratic model, the 2012 military coup might lead one to argue that Mali's recent history reflects constitutional weakness. However, the swift reinstatement of the constitution, popular commitment to its preservation, and attempts to pursue the post-coup transition in a constitutional manner all illustrate the domestic and international legitimacy of Mali's 1992 constitution. This article analyses the process of the proposed Malian constitutional referendum that, though constitutional in itself, contributed to the March 2012 overthrow of President Touré. It argues that the history of participatory constitutionalism in Mali contributed to the movement against the referendum. Constitutional reform is a necessity for an enduring constitution and this article sheds light on constitutional reform in states with weak legislatures, and illustrates the process of reform and the political divide that surrounded it in Mali. It provides a unique analysis of the crisis in Mali while at the same time making a contribution to our understanding of constitutionalism and constitutional reform in Africa.
Streptomyces natalensis programmed cell death and morphological differentiation are dependent on oxidative stress
Authors: Beites Tiago , Oliveira Paulo , Rioseras Beatriz , Pires Sílvia D. S. , Oliveira Rute , Tamagnini Paula , Moradas-Ferreira Pedro , Manteca Ángel , Mendes Marta V. 
Published: August 2015

AbstractStreptomyces are aerobic Gram-positive bacteria characterized by a complex life cycle that includes hyphae differentiation and spore formation. Morphological differentiation is triggered by stressful conditions and takes place in a pro-oxidant environment, which sets the basis for an involvement of the oxidative stress response in this cellular process. Characterization of the phenotypic traits of Streptomyces natalensis ΔkatA1 (mono-functional catalase) and ΔcatR (Fur-like repressor of katA1 expression) strains in solid medium revealed that both mutants had an impaired morphological development process. The sub-lethal oxidative stress caused by the absence of KatA1 resulted in the formation of a highly proliferative and undifferentiated vegetative mycelium, whereas de-repression of CatR regulon, from which KatA1 is the only known representative, resulted in the formation of scarce aerial mycelium. Both mutant strains had the transcription of genes associated with aerial mycelium formation and biosynthesis of the hyphae hydrophobic layer down-regulated. The first round of the programmed cell death (PCD) was inhibited in both strains which caused the prevalence of the transient primary mycelium (MI) over secondary mycelium (MII). Our data shows that the first round of PCD and morphological differentiation in S. natalensis is dependent on oxidative stress in the right amount at the right time.
A new strategy to inhibit the excision reaction catalysed by HIV-1 reverse transcriptase: compounds that compete with the template–primer
Authors: Cruchaga Carlos , Anso Elena , Font María , Martino Virginia S. , Rouzaut Ana , Martinez-Irujo Juan J. 
Published: August 2015

Inhibitors of the excision reaction catalysed by HIV-1 RT (reverse transcriptase) represent a promising approach in the fight against HIV, because these molecules would interfere with the main mechanism of resistance of this enzyme towards chain-terminating nucleotides. Only a limited number of compounds have been demonstrated to inhibit this reaction to date, including NNRTIs (non-nucleoside RT inhibitors) and certain pyrophosphate analogues. We have found previously that 2GP (2-O-galloylpunicalin), an antiviral compound extracted from the leaves of Terminalia triflora, was able to inhibit both the RT and the RNase H activities of HIV-1 RT without affecting cell proliferation or viability. In the present study, we show that 2GP also inhibited the ATP- and PPi-dependent phosphorolysis catalysed by wild-type and AZT (3′-azido-3′-deoxythymidine)-resistant enzymes at sub-micromolar concentrations. Kinetic and direct-binding analysis showed that 2GP was a non-competitive inhibitor against the nucleotide substrate, whereas it competed with the binding of RT to the template–primer (Kd=85 nM). As expected from its mechanism of action, 2GP was active against mutations conferring resistance to NNRTIs and AZT. The combination of AZT with 2GP was highly synergistic when tested in the presence of pyrophosphate, indicating that the inhibition of RT-catalysed phosphorolysis was responsible for the synergy found. Although other RT inhibitors that compete with the template–primer have been described, this is the first demonstration that these compounds can be used to block the excision of chain terminating nucleotides, providing a rationale for their combination with nucleoside analogues.
β-1,4-Glucanase-like protein from the cyanobacterium Synechocystis PCC6803 is a β-1,3-1,4-glucanase and functions in salt stress tolerance
Authors: Tamoi Masahiro , Kurotaki Hideki , Fukamizo Tamo 
Published: August 2015

In the present study, we characterized the gene (Cyanobase accession number slr0897) designated Ssglc encoding a β-1,4-glucanase-like protein (SsGlc) from Synechocystis PCC6803. The deduced amino acid sequence for Ssglc showed a high degree of similarity to sequences of GH (glycoside hydrolase) family 9 β-1,4-glucanases (cellulases) from various sources. Surprisingly, the recombinant protein obtained from the Escherichia coli expression system was able to hydrolyse barley β-glucan and lichenan (β-1,3-1,4-glucan), but not cellulose (β-1,4-glucan), curdlan (β-1,3-glucan), or laminarin (β-1,3-1,6-glucan). A 1H-NMR analysis of the enzymatic products revealed that the enzyme hydrolyses the β-1,4-glycosidic linkage of barley β-glucan through an inverting mechanism. The data indicated that SsGlc was a novel type of GH9 glucanase which could specifically hydrolyse the β-1,3-1,4-linkage of glucan. The growth of mutant Synechocystis cells in which the Ssglc gene was disrupted by a kanamycin-resistance cartridge gene was almost the same as that of the wild-type cells under continuous light (40 μmol of photons/m2 per s), a 12 h light (40 μmol of photons/m2 per s)/12 h dark cycle, cold stress (4 °C), and high light stress (200 μmol of photons/m2 per s). However, under salt stress (300–450 mM NaCl), growth of the Ssglc-disrupted mutant cells was significantly inhibited as compared with that of the wild-type cells. The Ssglc-disrupted mutant cells showed a decreased rate of O2 consumption and NaHCO3-dependent O2 evolution as compared with the wild-type cells under salt stress. Under osmotic stress (100–400 mM sorbitol), there was no difference in growth between the wild-type and the Ssglc-disrupted mutant cells. These results suggest that SsGlc functions in salt stress tolerance in Synechocystis PCC6803.
Distribution of mRNA for human epiregulin, a differentially expressed member of the epidermal growth factor family
Authors: TOYODA Hitoshi , KOMURASAKI Toshi , UCHIDA Daisuke , MORIMOTO Sigeo 
Published: August 2015

We have recently identified epiregulin as a new growth regulator and a member of the epidermal growth factor (EGF) family. Epiregulin has certain characteristics that are different from those of the classical members of the EGF family, EGF and transforming growth factor α, including mitogenic responses on several normal cells and binding to EGF receptors on epidermoid carcinoma A431 cells. In the present study we cloned and identified the expression of human epiregulin transcript. The human epiregulin gene encoded a 163-residue putative transmembrane precursor containing an EGF-like domain in the internal segment, and the structural organization was similar to that of other members of the EGF family that bind to EGF receptors. Northern blot analysis showed the expression of human epiregulin to be mainly on peripheral blood macrophages and the placenta in normal tissues, and was highest on epithelial tumour cell lines in various types of tumour cell lines. The expression profile was quite different from that of other members of the EGF family in normal and tumour cells. Recombinant expression in mammalian cells also showed that human epiregulin was secreted as a soluble form of approx. 5 kDa that is biologically active on the basis of the stimulation of DNA synthesis. Our findings suggest that epiregulin is involved in certain physiological processes such as maintenance or development of normal cell growth, and the progression of carcinomas.
Identification of human complement factor H as a chemotactic protein for monocytes
Authors: NABIL Karim , RIHN Bertrand , JAURAND Marie-Claude , VIGNAUD Jean-Michel , RIPOCHE Jean , MARTINET Yves , MARTINET Nadine 
Published: August 2015

We used chromatographic separation to purify to homogeneity a monomeric monocyte chemotactic protein of 150 kDa contained in mesothelioma pleural effusions. It was identified by N-terminal amino acid sequencing and immunoblotting as complement factor H, an inhibitor of the alternative complement pathway. Specific antibodies against factor H inhibited the monocyte chemotactic activity of the purified protein, which was most active at 10 nM. Factor H is a restrictive factor of alternative complement pathway activation. The new chemotactic function assigned to factor H in recruiting monocytes to the mesothelioma site might contribute to malignant cell phagocytosis via the iC3b/complement receptor type 3 pathway. These functions link the humoral and cellular immune systems.
Exposure of endothelial cells to cyclic strain induces elevations of cytosolic Ca2+ concentration through mobilization of intracellular and extracellular pools
Authors: ROSALES Oscar R. , ISALES Carlos M. , BARRETT Paula Q. , BROPHY Colleen , SUMPIO Bauer E. 
Published: August 2015

We have previously reported that exposure of endothelial cells to cyclic strain elicited a rapid but transient generation of inositol 1,4,5-trisphosphate (IP3), which reached a peak 10 s after the initiation of cyclic deformation. To address the effect of cyclic strain on intracellular Ca2+ concentration ([Ca2+]i) and its temporal relationship to IP3 generation, confluent bovine aortic endothelial cells were grown on flexible membranes, loaded with aequorin and the membranes placed in a custom-designed flow-through chamber. The chamber was housed inside a photomultiplier tube, and vacuum was utilized to deform the membranes. Our results indicate that the initiation of 10% average strain induced a rapid increase in [Ca2+]i which contained two distinct components: a large initial peak 12 s after the initiation of stretch which closely followed the IP3 peak, and a subsequent lower but sustained phase. Pretreatment with 5 μMGdCl3 for 10 min or nominally Ca2+-free medium (CFM) for 3 min reduced the magnitude of the initial rise and abolished the sustained phase. Repetitive 10% average strain at a frequency of 60 cycles/min also elicited a single IP3 peak at 10 s. However, there was also a large initial [Ca2+]i peak followed by multiple smaller transient [Ca2+]i elevations. Preincubation with 5 μMGdCl3 or CFM diminished the initial [Ca2+]i transient and markedly inhibited the late-phase component. Preincubation with 25 μM 2,5-di-(t-butyl)-1,4-benzohydroquinone (BHQ) attenuated the initial [Ca2+]i transient. Cyclic-strain-mediated IP3 formation in confluent endothelial cells at 10 s, however, was not modified by pretreatment with 25 μM BHQ, 500 μM NiCl2, 10 nM charybdotoxin, 5 μM GdCl3 or CFM. We conclude that in endothelial cells exposed to cyclic strain, Ca2+ enters the cytosol from intracellular and extracellular pools but IP3 formation is not dependent on Ca2+ entry via the plasma membrane.
Identification of three internalization sequences in the cytoplasmic tail of the 46 kDa mannose 6-phosphate receptor
Authors: DENZER Kristin , WEBER Birgit , HILLE-REHFELD Annette , VON FIGURA Kurt , POHLMANN Regina 
Published: August 2015

The cytoplasmic tail of the human 46 kDa mannose 6-phosphate receptor (MPR 46) is necessary for rapid internalization of the receptor and sufficient to mediate internalization of a resident plasma membrane protein. To localize the internalization sequences within the 67 amino acids of the cytoplasmic tail, the tail was progressively shortened from its C-terminus, internal deletions of between four and eight amino acids were introduced into the tail, and individual residues were substituted by alanine, glycine or serine. Three sequences were identified that contribute to the internalization of MPR 46. The first is located within the 23 juxtamembrane cytoplasmic residues of the tail. It contains four essential residues within a heptapeptide and does not resemble known internalization signals. The second sequence contains as a critical residue Tyr-45. The third region is located within the C-terminal seven residues and contains a di-leucine pair as essential residues. The first and third sequences were shown to function as autonomous internalization sequences. Substitution of critically important residues within a single internalization sequence was tolerated, with no or only a moderate decrease in the internalization rate. When essential residues from two or all three internalization sequences were substituted, however, the internalization rate was decreased by more than 60% and 90% respectively. This indicates that the autonomous internalization signals in the cytoplasmic tail of MPR 46 function in an additive manner, but are partly redundant.
Regulation of rat AMP deaminase 3 (isoform C) by development and skeletal muscle fibre type
Authors: MAHNKE-ZIZELMAN Donna K. , D'CUNHA Jonathan , WOJNAR Jean M. , BROGLEY Michele A. , SABINA Richard L. 
Published: August 2015

AMP deaminase (AMPD) is characterized by a multigene family in rodents and man. Highly conserved rat and human AMPD1 and AMPD2 genes produce protein products that exhibit cross-species immunoreactivities (AMPD1, rat isoform A and human isoform M; AMPD2, rat isoform B and human isoform L). A third gene, AMPD3, has been described in humans, but antisera raised against its purified protein product (isoform E) reportedly does not cross-react with a third activity purified from rat tissues (isoform C). This study was designed to address this latter issue by cloning, sequencing and expressing rat AMPD3 cDNA species. Similarly to the human AMPD3 gene, the rat AMPD3 gene produces multiple transcripts that differ at or near their 5′ ends. The boundary at which these alternative sequences diverge is precisely conserved in both species. Across the region that is common to all rat and human AMPD3 cDNA species, nucleotide and predicted amino acid sequences are 89% and 93% identical respectively, although the rat open reading frame is lacking two separate in-frame codons in the 5′ end. Extreme 5′ regions between the two species are entirely divergent, and one alternative rat sequence is predicted to confer at least 36 additional N-terminal residues to its encoded AMPD3 polypeptide. A comparison of 3′ untranslated regions indicates that the rat sequence is 250 bp longer and contains multiple consensus polyadenylation signals. Examination of relative rat AMPD3 gene expression shows (1) variable patterns of alternative mRNA abundance across adult tissues, (2) developmental regulation in skeletal muscle and liver, and (3) greater mRNA abundance in adult red (soleus) than in mixed (plantaris) and white (outer gastrocnemius) skeletal muscle. Finally, baculoviral expression of rat and human AMPD3 proteins produces enzymes that are chromatographically and kinetically similar. Moreover, both recombinant activities immunoreact with anti-C and anti-E serum. These combined results demonstrate that rat isoform C and human isoform E are homologous cross-species AMPD3 proteins.
Regulatory role of prostaglandin E2 in induction of cyclo-oxygenase-2 by a thromboxane A2 analogue (U46619) and basic fibroblast growth factor in porcine aortic smooth-muscle cells
Authors: KARIM Souad , BERROU Eliane , LÉVY-TOLEDANO Sylviane , BRYCKAERT Marijke , MACLOUF Jacques 
Published: August 2015

U46619, a thromboxane A2 analogue, and basic fibroblast growth factor (FGF-2) both induced the expression of the inducible cyclo-oxygenase (Cox)-2 in porcine aortic smooth-muscle cells. This induction was dose-dependent (submaximal at 300 nM for U46619 and 1 ng/ml for FGF-2) and time-dependent, with similar intensity and maximal expression at 2 h. Under these conditions, both inducers stimulated rapid activation of extracellular signal-regulated kinase (ERK2) at 5–10 min, a transient and lower intensity being induced by U46619 whereas that induced by FGF-2 was sustained (&gt; 1 h). PD98059, an inhibitor of the ERK pathway, inhibited the expression of Cox-2. In contrast, activation of Jun-N-terminal kinase (JNK1) was sustained with U46619 but poorly induced by FGF-2. Cox-2 expression induced by U46619 or FGF-2 was similarly reduced by prostaglandin (PGE2), forskolin or dibutyryl-cAMP, suggesting a regulatory effect of adenylate cyclase on Cox-2 expression. However, activation of ERK2 by FGF-2 was not affected by PGE2 whereas that of JNK1 by U46619 was inhibited, suggesting that inhibition of COX-2 expression by cAMP may be downstream of ERK2. The effects of PGE2 and forskolin on Cox-2 and phosphorylation of JNK1 were reversed with the protein kinase A inhibitor H89. In addition, endogenous PGE2 down-regulated the expression of Cox-2 by the two inducers, as stimulation of the cells in the presence of different Cox inhibitors increased the expression of the protein. Overall, these results suggest that exogenous and endogenous PGE2 exert negative inhibitory effects on Cox-2 expression mediated by stimulation of protein kinase A.
Nuclear factor κB is required for the transcriptional control of type II NO synthase in regenerating liver
Authors: DÍAZ-GUERRA Mara J. M. , VELASCO Marta , MARTÍN-SANZ Paloma , BOSCÁ Lisardo 
Published: August 2015

A concerted activation of transcription factors involved in the transactivation of type II NO synthase (iNOS) gene occurred after partial hepatectomy (PH), resulting in the transient expression of iNOS. The corresponding mRNA and protein levels of iNOS reached a maximum at 4 h and 8 h post-PH respectively. This induction was preceded by an early and transient activation of nuclear factor κB (NF-κB). Analysis of the κB inhibitory (I) proteins showed an important role for IκBα in the process of NF-κB activation, whereas the contribution of IκBβ was less evident. Interferon regulatory factor 1, which has been described as an important activator of iNOS expression, was up-regulated after PH but failed to bind to the corresponding DNA binding sequences of the iNOS promoter. The transcriptional control of iNOS after PH, was compared with the events associated with the hepatic expression of this enzyme in animals challenged with lipopolysaccharide, showing a differential pattern of transcription-factor activation and IκB degradation between both models. Transfection of hepatoma cell lines with iNOS promoter constructs, followed by stimulation with post-PH sera, revealed the requirement of NF-κB activation for iNOS expression. These data suggest that there is an important role for the restricted NF-κB activation in the temporal pattern of iNOS expression in regenerating liver.
Sequence of human carboxypeptidase D reveals it to be a member of the regulatory carboxypeptidase family with three tandem active site domains
Authors: TAN Fulong , REHLI Michael , KRAUSE Stefan W. , SKIDGEL Randal A. 
Published: August 2015

We have cloned the cDNA for human carboxypeptidase D (CPD), a new B-type metallocarboxypeptidase that is membrane bound and has an acidic pH optimum. The 5.8 kb of cDNA sequenced contains an open reading frame of 4131 bp encoding 1377 amino acid residues. The sequence is similar (75% identity) to duck gp180, a protein that was isolated, cloned and sequenced as a hepatitis B virus-binding protein but not characterized as a carboxypeptidase. Hydropathic analysis revealed a hydrophobic region at the N-terminus, representing the signal peptide, and one near the C-terminus that probably represents the transmembrane anchor. The most striking feature is the presence of three tandem carboxypeptidase homology domains that have sequence similarity to the regulatory B-type carboxypeptidase family, typified by carboxypeptidases M, E and N. Because of the three repeats, CPD is about three times larger (175–180 kDa) than other members of this family (approx. 50–62 kDa). Domain 2 is most closely related to carboxypeptidases M, E and N (45–48% identity), followed by domain 1 (37–38%) and domain 3 (20–27%). There is much higher sequence identity in regions containing putative active site residues, and all catalytically important residues are strictly conserved in domains 1 and 2. In domain 3, however, only 1 of 8 active site residues is conserved, indicating that this portion might not be catalytically active. Northern blotting of mRNA from human tissues and cells showed high levels of CPD mRNA in placenta, pancreas and Hep G2 hepatoma cells, and smaller amounts in skeletal muscle, heart and HT-29 colon carcinoma and melanoma cell lines.
Expression, purification and characterization of recombinant caprine N-acetylglucosamine-6-sulphatase
Authors: LITJENS Tom , BIELICKI Julie , ANSON Donald S. , FRIDERICI Karen , JONES Margaret Z. , HOPWOOD John J. 
Published: August 2015

Mucopolysaccharidosis type IIID or Sanfilippo D syndrome is a lysosomal storage disorder caused by the deficiency of N-acetylglucosamine-6-sulphatase (Glc6S). In addition to human patients, a Nubian goat with this disorder has been described and the caprine Glc6S (cGlc6S) cDNA cloned. In this study, the full-length cGlc6S cDNA was inserted into the expression vector, pEFNeo, which placed the cGlc6S cDNA under the transcriptional control of the human polypeptide chain elongation factor promoter. The pEFNeo expression vector also contains the human growth hormone polyadenylation signal and the genes encoding resistance to ampicillin and G418. The cGlc6S expression construct was electroporated into Chinese hamster ovary (CHO-K1) cells, and stably transfected clones were isolated. One clone, CHOrcGlc6S.17, which secreted the highest Glc6S activity into the culture medium, was selected and cultured in cell factories. The secreted recombinant cGlc6S (rcGlc6S) precursor was purified to homogeneity from conditioned medium by a two-column procedure which consisted of a Cu2+-chelating Sepharose column followed by TSK G3000SW gel filtration. The native molecular mass of rcGlc6S was estimated to be 102 kDa and the subunit size was 94 kDa. The kinetic properties of cGlc6S were similar to those of human Glc6S isolated from liver. rcGlc6S was endocytosed by fibroblasts from patients with mucopolysaccharidosis type IIID via the mannose 6-phosphate receptor-mediated pathway resulting in correction of the storage phenotype of these cells.