Gene expression profiling in wild-type and metallothionein mutant fibroblast cell lines

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ÁNGELA D ARMENDÁRIZ

2006-01-01

Full Text Available The role of metallothioneins (MT in copper homeostasis is of great interest, as it appears to be partially responsible for the regulation of intracellular copper levels during adaptation to extracellular excess of the metal. To further investigate a possible role of MTs in copper metabolism, a genomics approach was utilized to evaluate the role of MT on gene expression. Microarray analysis was used to examine the effects of copper overload in fibroblast cells from normal and MT I and II double knock-out mice (MT-/-. As a first step, we compared genes that were significantly upregulated in wild-type and MT-/- cells exposed to copper. Even though wild-type and mutant cells are undistinguishable in terms of their morphological features and rates of growth, our results show that MT-/- cells do not respond with induction of typical markers of cellular stress under copper excess conditions, as observed in the wild-type cell line, suggesting that the transcription initiation rate or the mRNA stability of stress genes is affected when there is an alteration in the copper store capacity. The functional classification of other up-regulated genes in both cell lines indicates that a large proportion (>80% belong to two major categories: 1 metabolism; and 2 cellular physiological processes, suggesting that at the transcriptional level copper overload induces the expression of genes associated with diverse molecular functions. These results open the possibility to understand how copper homeostasis is being coordinated with other metabolic pathways.

Introgression from cultivated rice alters genetic structures of wild relative populations: implications for in situ conservation

Science.gov (United States)

Jin, Xin; Chen, Yu; Liu, Ping; Li, Chen; Cai, Xingxing; Rong, Jun

2018-01-01

Abstract Maintaining genetic integrity is essential for in situ and ex situ conservation of crop wild relative (CWR) species. However, introgression of crop alleles into CWR species/populations may change their genetic structure and diversity, resulting in more invasive weeds or, in contrast, the extinction of endangered populations. To determine crop-wild introgression and its consequences, we examined the genetic structure and diversity of six wild rice (Oryza rufipogon) populations under in situ conservation in China. Thirty-four simple sequence repeat (SSR) and 34 insertion/deletion markers were used to genotype the wild rice populations and two sets of rice cultivars (O. sativa), corresponding to the two types of molecular markers. Shared alleles and STRUCTURE analyses suggested a variable level of crop-wild introgression and admixture. Principal coordinates and cluster analyses indicated differentiation of wild rice populations, which was associated with the spatial distances to cultivated rice fields. The level of overall genetic diversity was comparable between wild rice populations and rice cultivars, but a great number of wild-specific alleles was detected in the wild populations. We conclude based on the results that crop-wild introgression can considerably alter the pattern of genetic structure and relationships of CWR populations. Appropriate measures should be taken for effective in situ conservation of CWR species under the scenario of crop-wild introgression. PMID:29308123

Evaluation of MIC Strip Isavuconazole test for susceptibility testing of wild-type and non-wild-type Aspergillus fumigatus isolates

DEFF Research Database (Denmark)

Arendrup, Maiken Cavling; Verweij, Paul; Nielsen, Henrik Vedel

2017-01-01

We evaluated the MIC Strip Isavuconazole test against EUCAST E.Def 9.3 by using 40 wild-type and 39 CYP51A mutant Aspergillus fumigatus strains. The strip full inhibition endpoint (FIE) and 80% growth inhibition endpoint were determined by two independent readers, reader 1 (R1) and R2. The essent......We evaluated the MIC Strip Isavuconazole test against EUCAST E.Def 9.3 by using 40 wild-type and 39 CYP51A mutant Aspergillus fumigatus strains. The strip full inhibition endpoint (FIE) and 80% growth inhibition endpoint were determined by two independent readers, reader 1 (R1) and R2...

UV-sensitivity and repair of UV-damage in Salmonella of wild type

International Nuclear Information System (INIS)

Kondratiev, Y.S.; Brukhansky, G.V.; Andreeva, I.V.; Skavronskaya, A.G.

1977-01-01

The UV-sensitivity of wild type Salmonella strains has been compared to that of wild type E.coli and its UV-sensitive mutants. Many wild type Salmonella strains are 4-5 times more sensitive than wild type E.coli and their inactivation curve is similar to that for E.coli with a mutation in the polA gene. Alkaline sucrose gradient centrifugation has shown a deficiency of these strains in normal excision repair of UV-damaged DNA. This deficiency is not a Salmonella genus feature because one strain as resistant as wild type E.coli was found. This resistant strain showed normal excision repair in alkaline sucrose gradient centrifugation experiments. The possible influence of plasmids and mutations in repair genes on the ability of Salmonella to repair UV-damaged DNA is discussed. (orig.) [de

UV-sensitivity and repair of UV-damage in Salmonella of wild type

Energy Technology Data Exchange (ETDEWEB)

Kondratiev, Y S; Brukhansky, G V; Andreeva, I V; Skavronskaya, A G [Akademiya Meditsinskikh Nauk SSSR, Moscow. Inst. Ehpidemiologii i Mikrobiologii

1977-12-01

The UV-sensitivity of wild type Salmonella strains has been compared to that of wild type E.coli and its UV-sensitive mutants. Many wild type Salmonella strains are 4-5 times more sensitive than wild type E.coli and their inactivation curve is similar to that for E.coli with a mutation in the polA gene. Alkaline sucrose gradient centrifugation has shown a deficiency of these strains in normal excision repair of UV-damaged DNA. This deficiency is not a Salmonella genus feature because one strain as resistant as wild type E.coli was found. This resistant strain showed normal excision repair in alkaline sucrose gradient centrifugation experiments. The possible influence of plasmids and mutations in repair genes on the ability of Salmonella to repair UV-damaged DNA is discussed.

Sabin and wild type polioviruses from children who presented with ...

African Journals Online (AJOL)

Conclusion: This study further confirms the presence of Sabin and wild-type poliovirus among children in Nigeria. The isolation of Sabin strain of poliovirus is advantageous to the polio eradication program as it is capable of inducing natural immunity in susceptible hosts. Transmission of wild-type poliovirus among children ...

Elucidating Functional Aspects of P-type ATPases

DEFF Research Database (Denmark)

Autzen, Henriette Elisabeth

2015-01-01

and helped enlighten how thapsigargin, a potent inhibitor of SERCA1a, depends on a water mediated hydrogen bond network when bound to SERCA1a. Furthermore, molecular dynamics (MD) simulations of the same P-type ATPase were used to assess a long-standing question whether cholesterol affects SERCA1a through...... similar to that of the wild type (WT) protein. The discrepancy between the newly determined crystal structure of LpCopA and the functional manifestations of the missense mutation in human CopA, could indicate that LpCopA is insufficient in structurally elucidating the effect of disease-causing mutations...... in the human CopA proteins. MD simulations, which combine coarse-grained (CG) and atomistic procedures, were set up in order to elucidate mechanistic implications exerted by the lipid bilayer on LpCopA. The MD simulations of LpCopA corroborated previous and new in vivo activity data and showed...

DNA binding properties of dioxin receptors in wild-type and mutant mouse hepatoma cells

International Nuclear Information System (INIS)

Cuthill, S.; Poellinger, L.

1988-01-01

The current model of action of 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin) entails stimulation of target gene transcription via the formation of dioxin-receptor complexes and subsequent accumulation of the complexes within the cell nucleus. Here, the authors have analyzed the DNA binding properties of the dioxin receptor in wild-type mouse hepatoma (Hepa 1c1c7) cells and a class of nonresponsive mutant cells which fail to accumulate dioxin-receptor complexes within the nucleus in vivo. In vitro, both the wild-type and mutant [ 3 H]dioxin-receptor complexes exhibited low affinity for DNA-cellulose (5-8% and around 4% retention, respectively) in the absence of prior biochemical manipulations. However, following chromatography on heparin-Sepharose, the wild-type but not the mutant dioxin receptor was transformed to a species with an increased affinity for DNA (40-50% retention on DNA-cellulose). The gross molecular structure of the mutant, non DNA binding dioxin receptor did not appear to be altered as compared to that of the wild-type receptor. These results imply that the primary deficiency in the mutant dioxin receptor form may reside at the DNA binding level and that, in analogy to steroid hormone receptors, DNA binding of the receptor may be an essential step in the regulation of target gene transcription by dioxin

O6-methylguanine-DNA methyltransferase in wild-type and ada mutants of Escherichia coli

International Nuclear Information System (INIS)

Mitra, S.; Pal, B.C.; Foote, R.S.

1982-01-01

O 6 -Methylguanine-DNA methyltransferase is induced in Escherichia coli during growth in low levels of N-methyl-N'-nitro-N-nitrosoguanidine. We have developed a sensitive assay for quantitating low levels of this activity with a synthetic DNA substrate containing 3 H-labeled O 6 -methylguanine as the only modified base. Although both wild-type and adaptation-deficient (ada) mutants of E. coli contained low but comparable numbers (from 13 to 60) of the enzyme molecules per cell, adaptation treatment caused a significant increase of the enzyme in the wild type but not in the ada mutants, suggesting that the ada mutation is in a regulatory locus and not in the structural gene for the methyltransferase

Left ventricular structure and function in black normotensive type 2 ...

African Journals Online (AJOL)

Keywords: Black normotensive patients, left ventricular function, type 2 DM. Résumé ... sickle cell disease and structural heart disease were excluded ... Pulmonary venous flow (PVF) velocity ... had abnormal ECG pattern compared with 30%.

The role of a topologically conserved isoleucine in glutathione transferase structure, stability and function

International Nuclear Information System (INIS)

Achilonu, Ikechukwu; Gildenhuys, Samantha; Fisher, Loren; Burke, Jonathan; Fanucchi, Sylvia; Sewell, B. Trevor; Fernandes, Manuel; Dirr, Heini W.

2010-01-01

The role of a topologically conserved isoleucine in the structure of glutathione transferase was investigated by replacing the Ile71 residue in human GSTA1-1 by alanine or valine. The common fold shared by members of the glutathione-transferase (GST) family has a topologically conserved isoleucine residue at the N-terminus of helix 3 which is involved in the packing of helix 3 against two β-strands in domain 1. The role of the isoleucine residue in the structure, function and stability of GST was investigated by replacing the Ile71 residue in human GSTA1-1 by alanine or valine. The X-ray structures of the I71A and I71V mutants resolved at 1.75 and 2.51 Å, respectively, revealed that the mutations do not alter the overall structure of the protein compared with the wild type. Urea-induced equilibrium unfolding studies using circular dichroism and tryptophan fluorescence suggest that the mutation of Ile71 to alanine or valine reduces the stability of the protein. A functional assay with 1-chloro-2,4-dinitrobenzene shows that the mutation does not significantly alter the function of the protein relative to the wild type. Overall, the results suggest that conservation of the topologically conserved Ile71 maintains the structural stability of the protein but does not play a significant role in catalysis and substrate binding

Insights into the molecular basis for substrate binding and specificity of the wild-type L-arginine/agmatine antiporter AdiC.

Science.gov (United States)

Ilgü, Hüseyin; Jeckelmann, Jean-Marc; Gapsys, Vytautas; Ucurum, Zöhre; de Groot, Bert L; Fotiadis, Dimitrios

2016-09-13

Pathogenic enterobacteria need to survive the extreme acidity of the stomach to successfully colonize the human gut. Enteric bacteria circumvent the gastric acid barrier by activating extreme acid-resistance responses, such as the arginine-dependent acid resistance system. In this response, l-arginine is decarboxylated to agmatine, thereby consuming one proton from the cytoplasm. In Escherichia coli, the l-arginine/agmatine antiporter AdiC facilitates the export of agmatine in exchange of l-arginine, thus providing substrates for further removal of protons from the cytoplasm and balancing the intracellular pH. We have solved the crystal structures of wild-type AdiC in the presence and absence of the substrate agmatine at 2.6-Å and 2.2-Å resolution, respectively. The high-resolution structures made possible the identification of crucial water molecules in the substrate-binding sites, unveiling their functional roles for agmatine release and structure stabilization, which was further corroborated by molecular dynamics simulations. Structural analysis combined with site-directed mutagenesis and the scintillation proximity radioligand binding assay improved our understanding of substrate binding and specificity of the wild-type l-arginine/agmatine antiporter AdiC. Finally, we present a potential mechanism for conformational changes of the AdiC transport cycle involved in the release of agmatine into the periplasmic space of E. coli.

Dynamics of the Peripheral Membrane Protein P2 from Human Myelin Measured by Neutron Scattering--A Comparison between Wild-Type Protein and a Hinge Mutant.

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Saara Laulumaa

Full Text Available Myelin protein P2 is a fatty acid-binding structural component of the myelin sheath in the peripheral nervous system, and its function is related to its membrane binding capacity. Here, the link between P2 protein dynamics and structure and function was studied using elastic incoherent neutron scattering (EINS. The P38G mutation, at the hinge between the β barrel and the α-helical lid, increased the lipid stacking capacity of human P2 in vitro, and the mutated protein was also functional in cultured cells. The P38G mutation did not change the overall structure of the protein. For a deeper insight into P2 structure-function relationships, information on protein dynamics in the 10 ps to 1 ns time scale was obtained using EINS. Values of mean square displacements mainly from protein H atoms were extracted for wild-type P2 and the P38G mutant and compared. Our results show that at physiological temperatures, the P38G mutant is more dynamic than the wild-type P2 protein, especially on a slow 1-ns time scale. Molecular dynamics simulations confirmed the enhanced dynamics of the mutant variant, especially within the portal region in the presence of bound fatty acid. The increased softness of the hinge mutant of human myelin P2 protein is likely related to an enhanced flexibility of the portal region of this fatty acid-binding protein, as well as to its interactions with the lipid bilayer surface requiring conformational adaptations.

Foot Type Biomechanics Part 2: are structure and anthropometrics related to function?

Science.gov (United States)

Mootanah, Rajshree; Song, Jinsup; Lenhoff, Mark W; Hafer, Jocelyn F; Backus, Sherry I; Gagnon, David; Deland, Jonathan T; Hillstrom, Howard J

2013-03-01

Many foot pathologies are associated with specific foot types. If foot structure and function are related, measurement of either could assist with differential diagnosis of pedal pathologies. Biomechanical measures of foot structure and function are related in asymptomatic healthy individuals. Sixty-one healthy subjects' left feet were stratified into cavus (n=12), rectus (n=27) and planus (n=22) foot types. Foot structure was assessed by malleolar valgus index, arch height index, and arch height flexibility. Anthropometrics (height and weight), age, and walking speed were measured. Foot function was assessed by center of pressure excursion index, peak plantar pressure, maximum force, and gait pattern parameters. Foot structure and anthropometric variables were entered into stepwise linear regression models to identify predictors of function. Measures of foot structure and anthropometrics explained 10-37% of the model variance (adjusted R(2)) for gait pattern parameters. When walking speed was included, the adjusted R(2) increased to 45-77% but foot structure was no longer a factor. Foot structure and anthropometrics predicted 7-47% of the model variance for plantar pressure and 16-64% for maximum force parameters. All multivariate models were significant (pFoot structure and function are related in asymptomatic healthy individuals. The structural parameters employed are basic measurements that do not require ionizing radiation and could be used in a clinical setting. Further research is needed to identify additional predictive parameters (plantar soft tissue characteristics, skeletal alignment, and neuromuscular control) and to include individuals with pathology. Copyright © 2012. Published by Elsevier B.V.

Wild-type and mutated IDH1/2 enzymes and therapy responses.

Science.gov (United States)

Molenaar, Remco J; Maciejewski, Jaroslaw P; Wilmink, Johanna W; van Noorden, Cornelis J F

2018-04-01

Isocitrate dehydrogenase 1 and 2 (IDH1/2) are key enzymes in cellular metabolism, epigenetic regulation, redox states, and DNA repair. IDH1/2 mutations are causal in the development and/or progression of various types of cancer due to supraphysiological production of D-2-hydroxyglutarate. In various tumor types, IDH1/2-mutated cancers predict for improved responses to treatment with irradiation or chemotherapy. The present review discusses the molecular basis of the sensitivity of IDH1/2-mutated cancers with respect to the function of mutated IDH1/2 in cellular processes and their interactions with novel IDH1/2-mutant inhibitors. Finally, lessons learned from IDH1/2 mutations for future clinical applications in IDH1/2 wild-type cancers are discussed.

Screening and Expression of a Silicon Transporter Gene (Lsi1) in Wild-Type Indica Rice Cultivars

Science.gov (United States)

Abiri, Rambod; Kalhori, Nahid; Atabaki, Narges

2017-01-01

Silicon (Si) is one of the most prevalent elements in the soil. It is beneficial for plant growth and development, and it contributes to plant defense against different stresses. The Lsi1 gene encodes a Si transporter that was identified in a mutant Japonica rice variety. This gene was not identified in fourteen Malaysian rice varieties during screening. Then, a mutant version of Lsi1 was substituted for the native version in the three most common Malaysian rice varieties, MR219, MR220, and MR276, to evaluate the function of the transgene. Real-time PCR was used to explore the differential expression of Lsi1 in the three transgenic rice varieties. Silicon concentrations in the roots and leaves of transgenic plants were significantly higher than in wild-type plants. Transgenic varieties showed significant increases in the activities of the enzymes SOD, POD, APX, and CAT; photosynthesis; and chlorophyll content; however, the highest chlorophyll A and B levels were observed in transgenic MR276. Transgenic varieties have shown a stronger root and leaf structure, as well as hairier roots, compared to the wild-type plants. This suggests that Lsi1 plays a key role in rice, increasing the absorption and accumulation of Si, then alters antioxidant activities, and improves morphological properties. PMID:28191468

Screening and Expression of a Silicon Transporter Gene (Lsi1 in Wild-Type Indica Rice Cultivars

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Mahbod Sahebi

2017-01-01

Full Text Available Silicon (Si is one of the most prevalent elements in the soil. It is beneficial for plant growth and development, and it contributes to plant defense against different stresses. The Lsi1 gene encodes a Si transporter that was identified in a mutant Japonica rice variety. This gene was not identified in fourteen Malaysian rice varieties during screening. Then, a mutant version of Lsi1 was substituted for the native version in the three most common Malaysian rice varieties, MR219, MR220, and MR276, to evaluate the function of the transgene. Real-time PCR was used to explore the differential expression of Lsi1 in the three transgenic rice varieties. Silicon concentrations in the roots and leaves of transgenic plants were significantly higher than in wild-type plants. Transgenic varieties showed significant increases in the activities of the enzymes SOD, POD, APX, and CAT; photosynthesis; and chlorophyll content; however, the highest chlorophyll A and B levels were observed in transgenic MR276. Transgenic varieties have shown a stronger root and leaf structure, as well as hairier roots, compared to the wild-type plants. This suggests that Lsi1 plays a key role in rice, increasing the absorption and accumulation of Si, then alters antioxidant activities, and improves morphological properties.

Screening and Expression of a Silicon Transporter Gene (Lsi1) in Wild-Type Indica Rice Cultivars.

Science.gov (United States)

Sahebi, Mahbod; Hanafi, Mohamed M; Rafii, M Y; Azizi, Parisa; Abiri, Rambod; Kalhori, Nahid; Atabaki, Narges

2017-01-01

Silicon (Si) is one of the most prevalent elements in the soil. It is beneficial for plant growth and development, and it contributes to plant defense against different stresses. The Lsi1 gene encodes a Si transporter that was identified in a mutant Japonica rice variety. This gene was not identified in fourteen Malaysian rice varieties during screening. Then, a mutant version of Lsi1 was substituted for the native version in the three most common Malaysian rice varieties, MR219, MR220, and MR276, to evaluate the function of the transgene. Real-time PCR was used to explore the differential expression of Lsi1 in the three transgenic rice varieties. Silicon concentrations in the roots and leaves of transgenic plants were significantly higher than in wild-type plants. Transgenic varieties showed significant increases in the activities of the enzymes SOD, POD, APX, and CAT; photosynthesis; and chlorophyll content; however, the highest chlorophyll A and B levels were observed in transgenic MR276. Transgenic varieties have shown a stronger root and leaf structure, as well as hairier roots, compared to the wild-type plants. This suggests that Lsi1 plays a key role in rice, increasing the absorption and accumulation of Si, then alters antioxidant activities, and improves morphological properties.

Mathematical model of the SOS response regulation in wild-type Escherichia coli

International Nuclear Information System (INIS)

Aksenov, S.V.

1997-01-01

Regulation of the SOS response in Escherichia coli, which is a set of inducible cellular reactions introduced after DNA damage, is due to specific interaction of LexA and RecA proteins. LexA protein is a common repressor of the genes of the SOS system, and RecA protein, once transiently activated by the so-called SOS-inducing signal, promotes LexA protein destruction. We have described the SOS regulation by means of differential equations with regard to LexA and RecA concentrations elsewhere. The 'input' function for model equations is the level of the SOS-inducing signal against time. Here we present a means for calculating the concentration of single-stranded DNA (SOS-inducing signal) as a function of time in wild-type cells after ultraviolet irradiation. With model equations one can simulate kinetic curves of SOS regulatory proteins after DNA damage to survey the SOS response kinetics. Simulation of LexA protein kinetics agrees with experimental data. We compare simulated LexA kinetic curves in wild-type and uνr - mutant bacteria, which is useful in investigating the way uνrABC-dependent excision repair modulates the SOS response kinetics. Possible applications of the model to investigating various aspects of the SOS induction are discussed

Comparative effects of chlorpyrifos in wild type and cannabinoid Cb1 receptor knockout mice

Energy Technology Data Exchange (ETDEWEB)

Baireddy, Praveena; Liu, Jing; Hinsdale, Myron; Pope, Carey, E-mail: carey.pope@okstate.edu

2011-11-15

Endocannabinoids (eCBs) modulate neurotransmission by inhibiting the release of a variety of neurotransmitters. The cannabinoid receptor agonist WIN 55.212-2 (WIN) can modulate organophosphorus (OP) anticholinesterase toxicity in rats, presumably by inhibiting acetylcholine (ACh) release. Some OP anticholinesterases also inhibit eCB-degrading enzymes. We studied the effects of the OP insecticide chlorpyrifos (CPF) on cholinergic signs of toxicity, cholinesterase activity and ACh release in tissues from wild type (+/+) and cannabinoid CB1 receptor knockout (-/-) mice. Mice of both genotypes (n = 5-6/treatment group) were challenged with CPF (300 mg/kg, 2 ml/kg in peanut oil, sc) and evaluated for functional and neurochemical changes. Both genotypes exhibited similar cholinergic signs and cholinesterase inhibition (82-95% at 48 h after dosing) in cortex, cerebellum and heart. WIN reduced depolarization-induced ACh release in vitro in hippocampal slices from wild type mice, but had no effect in hippocampal slices from knockouts or in striatal slices from either genotype. Chlorpyrifos oxon (CPO, 100 {mu}M) reduced release in hippocampal slices from both genotypes in vitro, but with a greater reduction in tissues from wild types (21% vs 12%). CPO had no significant in vitro effect on ACh release in striatum. CPF reduced ACh release in hippocampus from both genotypes ex vivo, but reduction was again significantly greater in tissues from wild types (52% vs 36%). In striatum, CPF led to a similar reduction (20-23%) in tissues from both genotypes. Thus, while CB1 deletion in mice had little influence on the expression of acute toxicity following CPF, CPF- or CPO-induced changes in ACh release appeared sensitive to modulation by CB1-mediated eCB signaling in a brain-regional manner. -- Highlights: Black-Right-Pointing-Pointer C57Bl/6 mice showed dose-related cholinergic toxicity following subcutaneous chlorpyrifos exposure. Black-Right-Pointing-Pointer Wild type and

Comparative effects of chlorpyrifos in wild type and cannabinoid Cb1 receptor knockout mice

International Nuclear Information System (INIS)

Baireddy, Praveena; Liu, Jing; Hinsdale, Myron; Pope, Carey

2011-01-01

Endocannabinoids (eCBs) modulate neurotransmission by inhibiting the release of a variety of neurotransmitters. The cannabinoid receptor agonist WIN 55.212-2 (WIN) can modulate organophosphorus (OP) anticholinesterase toxicity in rats, presumably by inhibiting acetylcholine (ACh) release. Some OP anticholinesterases also inhibit eCB-degrading enzymes. We studied the effects of the OP insecticide chlorpyrifos (CPF) on cholinergic signs of toxicity, cholinesterase activity and ACh release in tissues from wild type (+/+) and cannabinoid CB1 receptor knockout (−/−) mice. Mice of both genotypes (n = 5–6/treatment group) were challenged with CPF (300 mg/kg, 2 ml/kg in peanut oil, sc) and evaluated for functional and neurochemical changes. Both genotypes exhibited similar cholinergic signs and cholinesterase inhibition (82–95% at 48 h after dosing) in cortex, cerebellum and heart. WIN reduced depolarization-induced ACh release in vitro in hippocampal slices from wild type mice, but had no effect in hippocampal slices from knockouts or in striatal slices from either genotype. Chlorpyrifos oxon (CPO, 100 μM) reduced release in hippocampal slices from both genotypes in vitro, but with a greater reduction in tissues from wild types (21% vs 12%). CPO had no significant in vitro effect on ACh release in striatum. CPF reduced ACh release in hippocampus from both genotypes ex vivo, but reduction was again significantly greater in tissues from wild types (52% vs 36%). In striatum, CPF led to a similar reduction (20–23%) in tissues from both genotypes. Thus, while CB1 deletion in mice had little influence on the expression of acute toxicity following CPF, CPF- or CPO-induced changes in ACh release appeared sensitive to modulation by CB1-mediated eCB signaling in a brain-regional manner. -- Highlights: ► C57Bl/6 mice showed dose-related cholinergic toxicity following subcutaneous chlorpyrifos exposure. ► Wild type and cannabinoid CB1 receptor knockout littermates

Modelling biological control with wild-type and genetically modified baculoviruses in the Helicoverpa armigera-cotton system

NARCIS (Netherlands)

Sun, X.; Werf, van der W.; Bianchi, F.J.J.A.; Hu, Z.; Vlak, J.M.

2006-01-01

A comprehensive model was developed to simulate virus epizootics in a stage structured insect population and analyse scenarios for the biological control of cotton bollworm (CBW), Helicoverpa armigera, in cotton, using wild-type or genetically modified baculoviruses. In simulations on dosage and

Functional flexibility in wild bonobo vocal behaviour

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Zanna Clay

2015-08-01

Full Text Available A shared principle in the evolution of language and the development of speech is the emergence of functional flexibility, the capacity of vocal signals to express a range of emotional states independently of context and biological function. Functional flexibility has recently been demonstrated in the vocalisations of pre-linguistic human infants, which has been contrasted to the functionally fixed vocal behaviour of non-human primates. Here, we revisited the presumed chasm in functional flexibility between human and non-human primate vocal behaviour, with a study on our closest living primate relatives, the bonobo (Pan paniscus. We found that wild bonobos use a specific call type (the “peep” across a range of contexts that cover the full valence range (positive-neutral-negative in much of their daily activities, including feeding, travel, rest, aggression, alarm, nesting and grooming. Peeps were produced in functionally flexible ways in some contexts, but not others. Crucially, calls did not vary acoustically between neutral and positive contexts, suggesting that recipients take pragmatic information into account to make inferences about call meaning. In comparison, peeps during negative contexts were acoustically distinct. Our data suggest that the capacity for functional flexibility has evolutionary roots that predate the evolution of human speech. We interpret this evidence as an example of an evolutionary early transition away from fixed vocal signalling towards functional flexibility.

Activation Of Wild-Type Hras Suppresses The Earliest Stages Of Pancreatic Cancer

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Jamie Weyandt

2015-08-01

Conclusions: Loss of wild-type Hras promotes the earliest stages of pancreatic tumorigenesis, and moreover results in more rapid progression of the disease. As such, mechanisms leading to activation of wild-type Ras proteins, including but not limited to redox-dependent reactions, may influence the development of pancreatic cancer.

Recovery of deficient homologous recombination in Brca2-depleted mouse cells by wild-type Rad51 expression.

Science.gov (United States)

Lee, Shauna A; Roques, Céline; Magwood, Alissa C; Masson, Jean-Yves; Baker, Mark D

2009-02-01

The BRCA2 tumor suppressor is important in maintaining genomic stability. BRCA2 is proposed to control the availability, cellular localization and DNA binding activity of the central homologous recombination protein, RAD51, with loss of BRCA2 resulting in defective homologous recombination. Nevertheless, the roles of BRCA2 in regulating RAD51 and how other proteins implicated in RAD51 regulation, such as RAD52 and RAD54 function relative to BRCA2 is not known. In this study, we tested whether defective homologous recombination in Brca2-depleted mouse hybridoma cells could be rectified by expression of mouse Rad51 or the Rad51-interacting mouse proteins, Rad52 and Rad54. In the Brca2-depleted cells, defective homologous recombination can be restored by over-expression of wild-type mouse Rad51, but not mouse Rad52 or Rad54. Correction of the homologous recombination defect requires Rad51 ATPase activity. A sizeable fraction ( approximately 50%) of over-expressed wild-type Rad51 is nuclear localized. The restoration of homologous recombination in the presence of a low (i.e., non-functional) level of Brca2 by wild-type Rad51 over-expression is unexpected. We suggest that Rad51 may access the nuclear compartment in a Brca2-independent manner and when Rad51 is over-expressed, the normal requirement for Brca2 control over Rad51 function in homologous recombination is dispensable. Our studies support loss of Rad51 function as a critical underlying factor in the homologous recombination defect in the Brca2-depleted cells.

Molecular Modeling and Structural Stability of Wild-Type and Mutant CYP51 from Leishmania major: In Vitro and In Silico Analysis of a Laboratory Strain

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Masoud Keighobadi

2018-03-01

Full Text Available Cutaneous leishmaniasis is a neglected tropical disease and a major public health in the most countries. Leishmania major is the most common cause of cutaneous leishmaniasis. In the Leishmania parasites, sterol 14α-demethylase (CYP51, which is involved in the biosynthesis of sterols, has been identified as an attractive target for development of new therapeutic agents. In this study, the sequence and structure of CYP51 in a laboratory strain (MRHO/IR/75/ER of L. major were determined and compared to the wild-type strain. The results showed 19 mutations including seven non-synonymous and 12 synonymous ones in the CYP51 sequence of strain MRHO/IR/75/ER. Importantly, an arginine to lysine substitution at position of 474 resulted in destabilization of CYP51 (ΔΔG = 1.17 kcal/mol in the laboratory strain; however, when the overall effects of all substitutions were evaluated by 100 ns molecular dynamics simulation, the final structure did not show any significant changes (p-value < 0.05 in stability parameter of the strain MRHO/IR/75/ER compared to the wild-type protein. The energy level for the CYP51 of wild-type and MRHO/IR/75/ER strain were −40,027.1 and −39,706.48 Kcal/mol respectively. The overall Root-mean-square deviation (RMSD deviation between two proteins was less than 1 Å throughout the simulation and Root-mean-square fluctuation (RMSF plot also showed no substantial differences between amino acids fluctuation of the both protein. The results also showed that, these mutations were located on the protein periphery that neither interferes with protein folding nor with substrate/inhibitor binding. Therefore, L. major strain MRHO/IR/75/ER is suggested as a suitable laboratory model for studying biological role of CYP51 and inhibitory effects of sterol 14α-demethylase inhibitors.

Interactions between the arbuscular mycorrhizal (AM) fungus Glomus intraradices and nontransformed tomato roots of either wild-type or AM-defective phenotypes in monoxenic cultures.

Science.gov (United States)

Bago, Alberto; Cano, Custodia; Toussaint, Jean-Patrick; Smith, Sally; Dickson, Sandy

2006-09-01

Monoxenic symbioses between the arbuscular mycorrhizal (AM) fungus Glomus intraradices and two nontransformed tomato root organ cultures (ROCs) were established. Wild-type tomato ROC from cultivar "RioGrande 76R" was employed as a control for mycorrhizal colonization and compared with its mutant line (rmc), which exhibits a highly reduced mycorrhizal colonization (rmc) phenotype. Structural features of the two root lines were similar when grown either in soil or under in vitro conditions, indicating that neither monoxenic culturing nor the rmc mutation affected root development or behavior. Colonization by G. intraradices in monoxenic culture of the wild-type line was low (<10%) but supported extensive development of extraradical mycelium, branched absorbing structures, and spores. The reduced colonization of rmc under monoxenic conditions (0.6%) was similar to that observed previously in soil. Extraradical development of runner hyphae was low and proportional to internal colonization. Few spores were produced. These results might suggest that carbon transfer may be modified in the rmc mutant. Our results support the usefulness of monoxenically obtained mycorrhizas for investigation of AM colonization and intraradical symbiotic functioning.

Phenylbutyrate Sensitizes Human Glioblastoma Cells Lacking Wild-Type P53 Function to Ionizing Radiation

International Nuclear Information System (INIS)

Lopez, Carlos A.; Feng, Felix Y.; Herman, Joseph M.; Nyati, Mukesh K.; Lawrence, Theodore S.; Ljungman, Mats

2007-01-01

Purpose: Histone deacetylase (HDAC) inhibitors induce growth arrest, differentiation, and apoptosis in cancer cells. Phenylbutyrate (PB) is a HDAC inhibitor used clinically for treatment of urea cycle disorders. Because of its low cytotoxicity, cerebrospinal fluid penetration, and high oral bioavailability, we investigated PB as a potential radiation sensitizer in human glioblastoma cell lines. Methods and Materials: Four glioblastoma cell lines were selected for this study. Phenylbutyrate was used at a concentration of 2 mM, which is achievable in humans. Western blots were used to assess levels of acetylated histone H3 in tumor cells after treatment with PB. Flow cytometry was used for cell cycle analysis. Clonogenic assays were performed to assess the effect of PB on radiation sensitivity. We used shRNA against p53 to study the role of p53 in radiosensitization. Results: Treatment with PB alone resulted in hyperacetylation of histones, confirmed by Western blot analysis. The PB alone resulted in cytostatic effects in three cell lines. There was no evidence of G 1 arrest, increase in sub-G 1 fraction or p21 protein induction. Clonogenic assays showed radiosensitization in two lines harboring p53 mutations, with enhancement ratios (± SE) of 1.5 (± 0.2) and 1.3 (± 0.1), respectively. There was no radiopotentiating effect in two cell lines with wild-type p53, but knockdown of wild-type p53 resulted in radiosensitization by PB. Conclusions: Phenylbutyrate can produce p21-independent cytostasis, and enhances radiation sensitivity in p53 mutant human glioblastoma cells in vitro. This suggests the potential application of combined PB and radiotherapy in glioblastoma harboring mutant p53

Biochemical characterization and structural modeling of human cathepsin E variant 2 in comparison to the wild-type protein

Science.gov (United States)

Puizdar, Vida; Zajc, Tajana; Žerovnik, Eva; Renko, Miha; Pieper, Ursula; Eswar, Narayanan; Šali, Andrej; Dolenc, Iztok; Turk, Vito

2014-01-01

Cathepsin E splice variant 2 appears in a number of gastric carcinoma. Here, we report detecting this variant in HeLa cells using polyclonal antibodies and biotinylated inhibitor pepstatin A. An overexpression of GFP fusion proteins of cathepsin E and its splice variant within HEK-293T cells was performed to show their localization. Their distribution under a fluorescence microscope showed that they are colocalized. We also expressed variant 1 and variant 2 of cathepsins E, with propeptide and without it, in Echerichia coli. After refolding from the inclusion bodies, the enzymatic activity and circular dichroism spectra of the splice variant 2 were compared to those of the wild-type mature active cathepsins E. While full-length cathepsin E variant1 is activated at acid pH, the splice variant remains inactive. In contrast to the active cathepsin E, the splice variant 2 predominantly assumes β-sheet structure, prone to oligomerization, at least under in vitro conditions, as shown by Atomic Force Microscopy as shallow disk-like particles. A comparative structure model of splice variant 2 was computed based on its alignment to the known structure of cathepsin E intermediate (Protein Data Bank code 1TZS), and used to rationalize its conformational properties and loss of activity. PMID:22718633

Comparative behaviour of lab.-cultured and wild-type Dacus oleae flies in the field

International Nuclear Information System (INIS)

Prokopy, R.J.; Haniotakis, G.E.; Economopoulos, A.P.

1975-01-01

Under field conditions, the authors compared the responses of lab.-type (ca. 85 generations under artificial conditions) and wild-type Dacus oleae flies to host plant colour and odour, host fruit colour and shape, small rectangles of different colours and shades, and McPhail-type traps of different colours baited with different odours. Except for the lab.-type flies being relatively more attracted toward red fruit models and small red rectangles and relatively less attracted toward yellow fruit models and small yellow rectangles than the wild type, the qualitative nature of the responses of the two fly types toward the various experimental treatments was essentially the same. Quantitatively, however, consistently smaller percentages of the released lab.-type than the released wild-type flies were recaptured, suggesting that the mobility, flight pattern, or vigour of the two types of flies may be different. (author)

Biophysical investigation of type A PutAs reveals a conserved core oligomeric structure

Energy Technology Data Exchange (ETDEWEB)

Korasick, David A. [Department of Biochemistry, University of Missouri, Columbia MO USA; Singh, Harkewal [Department of Chemistry, University of Missouri, Columbia MO USA; Pemberton, Travis A. [Department of Chemistry, University of Missouri, Columbia MO USA; Luo, Min [Department of Chemistry, University of Missouri, Columbia MO USA; Dhatwalia, Richa [Department of Chemistry, University of Missouri, Columbia MO USA; Tanner, John J. [Department of Biochemistry, University of Missouri, Columbia MO USA; Department of Chemistry, University of Missouri, Columbia MO USA

2017-08-01

Many enzymes form homooligomers, yet the functional significance of self-association is seldom obvious. Herein, we examine the connection between oligomerization and catalytic function for proline utilization A (PutA) enzymes. PutAs are bifunctional enzymes that catalyze both reactions of proline catabolism. Type A PutAs are the smallest members of the family, possessing a minimal domain architecture consisting of N-terminal proline dehydrogenase and C-terminal l-glutamate-γ-semialdehyde dehydrogenase modules. Type A PutAs form domain-swapped dimers, and in one case (Bradyrhizobium japonicum PutA), two of the dimers assemble into a ring-shaped tetramer. Whereas the dimer has a clear role in substrate channeling, the functional significance of the tetramer is unknown. To address this question, we performed structural studies of four-type A PutAs from two clades of the PutA tree. The crystal structure of Bdellovibrio bacteriovorus PutA covalently inactivated by N-propargylglycine revealed a fold and substrate-channeling tunnel similar to other PutAs. Small-angle X-ray scattering (SAXS) and analytical ultracentrifugation indicated that Bdellovibrio PutA is dimeric in solution, in contrast to the prediction from crystal packing of a stable tetrameric assembly. SAXS studies of two other type A PutAs from separate clades also suggested that the dimer predominates in solution. To assess whether the tetramer of B. japonicum PutA is necessary for catalytic function, a hot spot disruption mutant that cleanly produces dimeric protein was generated. The dimeric variant exhibited kinetic parameters similar to the wild-type enzyme. These results implicate the domain-swapped dimer as the core structural and functional unit of type A PutAs.

Impact of type 2 diabetes and duration of type 2 diabetes on cardiac structure and function

DEFF Research Database (Denmark)

Jørgensen, Peter G; Jensen, Magnus T; Mogelvang, Rasmus

2016-01-01

BACKGROUND: Contemporary treatment of type 2 diabetes (T2D) has improved patient outcome and may also have affected myocardial structure and function. We aimed to describe the effect of T2D and T2D duration on cardiac structure and function in a large outpatient population. METHODS: We performed...... comprehensive echocardiography on a representative sample of 1004 persons including a representative sample of 770 patients with T2D without known heart disease and 234 age- and sex-matched controls. RESULTS: T2D was associated with increased left ventricular (LV) wall thicknesses and decreased LV internal...... dysfunction persisted after multivariable adjustment (P=0.013). CONCLUSIONS: In patients with T2D, LV structural and functional alterations persist and are accentuated with increasing diabetes duration despite reductions in overall risk of cardiovascular disease in this patient population....

Structural and functional properties of alkali-treated high-amylose rice starch.

Science.gov (United States)

Cai, Jinwen; Yang, Yang; Man, Jianmin; Huang, Jun; Wang, Zhifeng; Zhang, Changquan; Gu, Minghong; Liu, Qiaoquan; Wei, Cunxu

2014-02-15

Native starches were isolated from mature grains of high-amylose transgenic rice TRS and its wild-type rice TQ and treated with 0.1% and 0.4% NaOH for 7 and 14 days at 35 °C. Alkali-treated starches were characterised for structural and functional properties using various physical methods. The 0.1% NaOH treatment had no significant effect on structural and functional properties of starches except that it markedly increased the hydrolysis of starch by amylolytic enzymes. The 0.4% NaOH treatment resulted in some changes in structural and functional properties of starches. The alkali treatment affected granule morphology and decreased the electron density between crystalline and amorphous lamellae of starch. The effect of alkali on the crystalline structure including long- and short-range ordered structure was not pronounced. Compared with control starch, alkali-treated TRS starches had lower amylose content, higher onset and peak gelatinisation temperatures, and faster hydrolysis of starch by HCl and amylolytic enzymes. Copyright © 2013 Elsevier Ltd. All rights reserved.

Comparison of the proteomes of three yeast wild type strains: CEN.PK2, FY1679 and W303

DEFF Research Database (Denmark)

Rogowska-Wrzesinska, A.; Mose Larsen, P.; Blomberg, A.

2001-01-01

Yeast deletion strains created during gene function analysis projects very often show drastic phenotypic differences depending on the genetic background used. These results indicate the existence of important molecular differences between the CEN.PK2, FY1679 and W303 wild type strains...

A Ten-Week Biochemistry Lab Project Studying Wild-Type and Mutant Bacterial Alkaline Phosphatase

Science.gov (United States)

Witherow, D. Scott

2016-01-01

This work describes a 10-week laboratory project studying wild-type and mutant bacterial alkaline phosphatase, in which students purify, quantitate, and perform kinetic assays on wild-type and selected mutants of the enzyme. Students also perform plasmid DNA purification, digestion, and gel analysis. In addition to simply learning important…

Structural and functional analysis of APOA5 mutations identified in patients with severe hypertriglyceridemia[S

Science.gov (United States)

Mendoza-Barberá, Elena; Julve, Josep; Nilsson, Stefan K.; Lookene, Aivar; Martín-Campos, Jesús M.; Roig, Rosa; Lechuga-Sancho, Alfonso M.; Sloan, John H.; Fuentes-Prior, Pablo; Blanco-Vaca, Francisco

2013-01-01

During the diagnosis of three unrelated patients with severe hypertriglyceridemia, three APOA5 mutations [p.(Ser232_Leu235)del, p.Leu253Pro, and p.Asp332ValfsX4] were found without evidence of concomitant LPL, APOC2, or GPIHBP1 mutations. The molecular mechanisms by which APOA5 mutations result in severe hypertriglyceridemia remain poorly understood, and the functional impairment/s induced by these specific mutations was not obvious. Therefore, we performed a thorough structural and functional analysis that included follow-up of patients and their closest relatives, measurement of apoA-V serum concentrations, and sequencing of the APOA5 gene in 200 nonhyperlipidemic controls. Further, we cloned, overexpressed, and purified both wild-type and mutant apoA-V variants and characterized their capacity to activate LPL. The interactions of recombinant wild-type and mutated apoA-V variants with liposomes of different composition, heparin, LRP1, sortilin, and SorLA/LR11 were also analyzed. Finally, to explore the possible structural consequences of these mutations, we developed a three-dimensional model of full-length, lipid-free human apoA-V. A complex, wide array of impairments was found in each of the three mutants, suggesting that the specific residues affected are critical structural determinants for apoA-V function in lipoprotein metabolism and, therefore, that these APOA5 mutations are a direct cause of hypertriglyceridemia. PMID:23307945

Differential induction of Toll-like receptors & type 1 interferons by Sabin attenuated & wild type 1 polioviruses in human neuronal cells.

Science.gov (United States)

Mohanty, Madhu C; Deshpande, Jagadish M

2013-01-01

Polioviruses are the causative agent of paralytic poliomyelitis. Attenuated polioviruses (Sabin oral poliovirus vaccine strains) do not replicate efficiently in neurons as compared to the wild type polioviruses and therefore do not cause disease. This study was aimed to investigate the differential host immune response to wild type 1 poliovirus (wild PV) and Sabin attenuated type 1 poliovirus (Sabin PV) in cultured human neuronal cells. By using flow cytometry and real time PCR methods we examined host innate immune responses and compared the role of toll like receptors (TLRs) and cytoplasmic RNA helicases in cultured human neuronal cells (SK-N-SH) infected with Sabin PV and wild PV. Human neuronal cells expressed very low levels of TLRs constitutively. Sabin PV infection induced significantly higher expression of TLR3, TLR7 and melanoma differentiation-associated protein-5 (MDA-5) m-RNA in neuronal cells at the beginning of infection (up to 4 h) as compared to wild PV. Further, Sabin PV also induced the expression of interferon α/β at early time point of infection. The induced expression of IFN α/β gene by Sabin PV in neuronal cells could be suppressed by inhibiting TLR7. Neuronal cell innate immune response to Sabin and wild polioviruses differ significantly for TLR3, TLR7, MDA5 and type 1 interferons. Effects of TLR7 activation and interferon production and Sabin virus replication in neuronal cells need to be actively investigated in future studies.

Structural consistency analysis of recombinant and wild-type human serum albumin

Science.gov (United States)

Cao, Hui-Ling; Sun, Li-Hua; Liu, Li; Li, Jian; Tang, Lin; Guo, Yun-Zhu; Mei, Qi-Bing; He, Jian-Hua; Yin, Da-Chuan

2017-01-01

Recombinant human serum albumin (rHSA) is potential alternatives for human serum albumin (HSA) which may ease severe shortage of HSA worldwide. In theory, rHSA and HSA are the same. Structure decides function. Therefore, the 3D structural consistency analysis of rHSA and HSA is outmost importance, which is the base of their function consistency. In this paper, the crystal structures of rHSA at resolution limit of 2.22 Å and HSA at 2.30 Å were determined by X-ray diffraction (XRD), which were deposited in the Protein Data Bank (PDB) with accession codes 4G03 (rHSA) and 4G04 (HSA). The differences between rHSA and HSA were systematically analyzed from the crystallization behavior, diffraction data and three-dimensional (3D) structure. The superimposed contrasted analysis indicated that rHSA and HSA achieved a structural similarity of 99% with an r.m.s. deviation of 0.397 Å for the corresponding overall Cα atoms. In addition, the number of α-helices in the rHSA or HSA molecule was verified to be 30. As a result, rHSA can potentially replace HSA. The study provides a theoretical and experimental basis for the clinical and additional applications of rHSA. Meanwhile, it is also a good example for applications of genetic engineering.

Effect of wild-type Shigella species and attenuated Shigella vaccine candidates on small intestinal barrier function, antigen trafficking, and cytokine release.

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Maria Fiorentino

Full Text Available Bacterial dysentery due to Shigella species is a major cause of morbidity and mortality worldwide. The pathogenesis of Shigella is based on the bacteria's ability to invade and replicate within the colonic epithelium, resulting in severe intestinal inflammatory response and epithelial destruction. Although the mechanisms of pathogenesis of Shigella in the colon have been extensively studied, little is known on the effect of wild-type Shigella on the small intestine and the role of the host response in the development of the disease. Moreover, to the best of our knowledge no studies have described the effects of apically administered Shigella flexneri 2a and S. dysenteriae 1 vaccine strains on human small intestinal enterocytes. The aim of this study was to assess the coordinated functional and immunological human epithelial responses evoked by strains of Shigella and candidate vaccines on small intestinal enterocytes. To model the interactions of Shigella with the intestinal mucosa, we apically exposed monolayers of human intestinal Caco2 cells to increasing bacterial inocula. We monitored changes in paracellular permeability, examined the organization of tight-junctions and the pro-inflammatory response of epithelial cells. Shigella infection of Caco2 monolayers caused severe mucosal damage, apparent as a drastic increase in paracellular permeability and disruption of tight junctions at the cell-cell boundary. Secretion of pro-inflammatory IL-8 was independent of epithelial barrier dysfunction. Shigella vaccine strains elicited a pro-inflammatory response without affecting the intestinal barrier integrity. Our data show that wild-type Shigella infection causes a severe alteration of the barrier function of a small intestinal cell monolayer (a proxy for mucosa and might contribute (along with enterotoxins to the induction of watery diarrhea. Diarrhea may be a mechanism by which the host attempts to eliminate harmful bacteria and transport them

Structures of a bi-functional Kunitz-type STI family inhibitor of serine and aspartic proteases: Could the aspartic protease inhibition have evolved from a canonical serine protease-binding loop?

Science.gov (United States)

Guerra, Yasel; Valiente, Pedro A; Pons, Tirso; Berry, Colin; Rudiño-Piñera, Enrique

2016-08-01

Bi-functional inhibitors from the Kunitz-type soybean trypsin inhibitor (STI) family are glycosylated proteins able to inhibit serine and aspartic proteases. Here we report six crystal structures of the wild-type and a non-glycosylated mutant of the bifunctional inhibitor E3Ad obtained at different pH values and space groups. The crystal structures show that E3Ad adopts the typical β-trefoil fold of the STI family exhibiting some conformational changes due to pH variations and crystal packing. Despite the high sequence identity with a recently reported potato cathepsin D inhibitor (PDI), three-dimensional structures obtained in this work show a significant conformational change in the protease-binding loop proposed for aspartic protease inhibition. The E3Ad binding loop for serine protease inhibition is also proposed, based on structural similarity with a novel non-canonical conformation described for the double-headed inhibitor API-A from the Kunitz-type STI family. In addition, structural and sequence analyses suggest that bifunctional inhibitors of serine and aspartic proteases from the Kunitz-type STI family are more similar to double-headed inhibitor API-A than other inhibitors with a canonical protease-binding loop. Copyright © 2016. Published by Elsevier Inc.

Differential induction of Toll-like receptors & type 1 interferons by Sabin attenuated & wild type 1 polioviruses in human neuronal cells

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Madhu C Mohanty

2013-01-01

Full Text Available Background & objectives: Polioviruses are the causative agent of paralytic poliomyelitis. Attenuated polioviruses (Sabin oral poliovirus vaccine strains do not replicate efficiently in neurons as compared to the wild type polioviruses and therefore do not cause disease. This study was aimed to investigate the differential host immune response to wild type 1 poliovirus (wild PV and Sabin attenuated type 1 poliovirus (Sabin PV in cultured human neuronal cells. Methods: By using flow cytometry and real time PCR methods we examined host innate immune responses and compared the role of toll like receptors (TLRs and cytoplasmic RNA helicases in cultured human neuronal cells (SK-N-SH infected with Sabin PV and wild PV. Results: Human neuronal cells expressed very low levels of TLRs constitutively. Sabin PV infection induced significantly higher expression of TLR3, TLR7 and melanoma differentiation-associated protein-5 (MDA-5 m-RNA in neuronal cells at the beginning of infection (up to 4 h as compared to wild PV. Further, Sabin PV also induced the expression of interferon α/β at early time point of infection. The induced expression of IFN α/β gene by Sabin PV in neuronal cells could be suppressed by inhibiting TLR7. Interpretation & conclusions: Neuronal cell innate immune response to Sabin and wild polioviruses differ significantly for TLR3, TLR7, MDA5 and type 1 interferons. Effects of TLR7 activation and interferon production and Sabin virus replication in neuronal cells need to be actively investigated in future studies.

lambda. -prophage induction in repair-deficient and wild type E. coli strains by. gamma. -rays and heavy ions

Energy Technology Data Exchange (ETDEWEB)

Bonev, M.N.; Kozubek, S.; Krasavin, E.A.; Amirtajev, K.G. (Joint Inst. for Nuclear Research, Dubna (USSR))

1990-05-01

{lambda}-prophage induction in repair-deficient and wild-type E. coli strains by heavy ions and {gamma}-rays was investigated. The dose dependence of the fraction of induced cells has been measured and its initial slope ({lambda}-induction potency) determined. Induction by {gamma}-rays was found to be more efficient in a polA-repair-deficient strain; the value of {lambda}-induction potency is zero in lexA{sup -} and recA{sup -} strains. The {lambda}-induction potency potency increased with LET for wild-type cells but remained constant in polA{sup -} mutant cells. It is suggested that DNA damage triggering the {lambda}-prophage induction in the case of ionizing radiation could be a type of DNA single-strand break with complex structures which cannot be repaired by fast repair processes, and requires a substantial level of energy deposition for induction in a DNA molecule. (author).

Age structure changes and extraordinary lifespan in wild medfly populations.

Science.gov (United States)

Carey, James R; Papadopoulos, Nikos T; Müller, Hans-Georg; Katsoyannos, Byron I; Kouloussis, Nikos A; Wang, Jane-Ling; Wachter, Kenneth; Yu, Wei; Liedo, Pablo

2008-06-01

The main purpose of this study was to test the hypotheses that major changes in age structure occur in wild populations of the Mediterranean fruit fly (medfly) and that a substantial fraction of individuals survive to middle age and beyond (> 3-4 weeks). We thus brought reference life tables and deconvolution models to bear on medfly mortality data gathered from a 3-year study of field-captured individuals that were monitored in the laboratory. The average time-to-death of captured females differed between sampling dates by 23.9, 22.7, and 37.0 days in the 2003, 2004, and 2005 field seasons, respectively. These shifts in average times-to-death provided evidence of changes in population age structure. Estimates indicated that middle-aged medflies (> 30 days) were common in the population. A surprise in the study was the extraordinary longevity observed in field-captured medflies. For example, 19 captured females but no reference females survived in the laboratory for 140 days or more, and 6 captured but no reference males survived in the laboratory for 170 days or more. This paper advances the study of aging in the wild by introducing a new method for estimating age structure in insect populations, demonstrating that major changes in age structure occur in field populations of insects, showing that middle-aged individuals are common in the wild, and revealing the extraordinary lifespans of wild-caught individuals due to their early life experience in the field.

Pharmacologic Treatment Assigned for Niemann Pick Type C1 Disease Partly Changes Behavioral Traits in Wild-Type Mice.

Science.gov (United States)

Schlegel, Victoria; Thieme, Markus; Holzmann, Carsten; Witt, Martin; Grittner, Ulrike; Rolfs, Arndt; Wree, Andreas

2016-11-09

Niemann-Pick Type C1 (NPC1) is an autosomal recessive inherited disorder characterized by accumulation of cholesterol and glycosphingolipids. Previously, we demonstrated that BALB/c-npc1 nih Npc1 -/- mice treated with miglustat, cyclodextrin and allopregnanolone generally performed better than untreated Npc1 -/- animals. Unexpectedly, they also seemed to accomplish motor tests better than their sham-treated wild-type littermates. However, combination-treated mutant mice displayed worse cognition performance compared to sham-treated ones. To evaluate effects of these drugs in healthy BALB/c mice, we here analyzed pharmacologic effects on motor and cognitive behavior of wild-type mice. For combination treatment mice were injected with allopregnanolone/cyclodextrin weekly, starting at P7. Miglustat injections were performed daily from P10 till P23. Starting at P23, miglustat was embedded in the chow. Other mice were treated with miglustat only, or sham-treated. The battery of behavioral tests consisted of accelerod, Morris water maze, elevated plus maze, open field and hot-plate tests. Motor capabilities and spontaneous motor behavior were unaltered in both drug-treated groups. Miglustat-treated wild-type mice displayed impaired spatial learning compared to sham- and combination-treated mice. Both combination- and miglustat-treated mice showed enhanced anxiety in the elevated plus maze compared to sham-treated mice. Additionally, combination treatment as well as miglustat alone significantly reduced brain weight, whereas only combination treatment reduced body weight significantly. Our results suggest that allopregnanolone/cyclodextrin ameliorate most side effects of miglustat in wild-type mice.

Pharmacologic Treatment Assigned for Niemann Pick Type C1 Disease Partly Changes Behavioral Traits in Wild-Type Mice

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Victoria Schlegel

2016-11-01

Full Text Available Niemann-Pick Type C1 (NPC1 is an autosomal recessive inherited disorder characterized by accumulation of cholesterol and glycosphingolipids. Previously, we demonstrated that BALB/c-npc1nihNpc1−/− mice treated with miglustat, cyclodextrin and allopregnanolone generally performed better than untreated Npc1−/− animals. Unexpectedly, they also seemed to accomplish motor tests better than their sham-treated wild-type littermates. However, combination-treated mutant mice displayed worse cognition performance compared to sham-treated ones. To evaluate effects of these drugs in healthy BALB/c mice, we here analyzed pharmacologic effects on motor and cognitive behavior of wild-type mice. For combination treatment mice were injected with allopregnanolone/cyclodextrin weekly, starting at P7. Miglustat injections were performed daily from P10 till P23. Starting at P23, miglustat was embedded in the chow. Other mice were treated with miglustat only, or sham-treated. The battery of behavioral tests consisted of accelerod, Morris water maze, elevated plus maze, open field and hot-plate tests. Motor capabilities and spontaneous motor behavior were unaltered in both drug-treated groups. Miglustat-treated wild-type mice displayed impaired spatial learning compared to sham- and combination-treated mice. Both combination- and miglustat-treated mice showed enhanced anxiety in the elevated plus maze compared to sham-treated mice. Additionally, combination treatment as well as miglustat alone significantly reduced brain weight, whereas only combination treatment reduced body weight significantly. Our results suggest that allopregnanolone/cyclodextrin ameliorate most side effects of miglustat in wild-type mice.

Functional analysis in mouse embryonic stem cells reveals wild-type activity for three MSH6 variants found in suspected Lynch syndrome patients.

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Eva A L Wielders

Full Text Available Lynch syndrome confers an increased risk to various types of cancer, in particular early onset colorectal and endometrial cancer. Mutations in mismatch repair (MMR genes underlie Lynch syndrome, with the majority of mutations found in MLH1 and MSH2. Mutations in MSH6 have also been found but these do not always cause a clear cancer predisposition phenotype and MSH6-defective tumors often do not show the standard characteristics of MMR deficiency, such as microsatellite instability. In particular, the consequences of MSH6 missense mutations are challenging to predict, which further complicates genetic counseling. We have previously developed a method for functional characterization of MSH2 missense mutations of unknown significance. This method is based on endogenous gene modification in mouse embryonic stem cells using oligonucleotide-directed gene targeting, followed by a series of functional assays addressing the MMR functions. Here we have adapted this method for the characterization of MSH6 missense mutations. We recreated three MSH6 variants found in suspected Lynch syndrome families, MSH6-P1087R, MSH6-R1095H and MSH6-L1354Q, and found all three to behave like wild type MSH6. Thus, despite suspicion for pathogenicity from clinical observations, our approach indicates these variants are not disease causing. This has important implications for counseling of mutation carriers.

Structure/Function Analysis of Protein-Protein Interactions and Role of Dynamic Motions in Mercuric Ion Reductase

Energy Technology Data Exchange (ETDEWEB)

Miller, Susan M.

2005-05-18

This report summarizes the activities and findings of our structure/function studies of the bacterial detoxification enzyme mercuric ion reductase. The objectives of the work were to obtain crystal structure information for the catalytic core of this enzyme, use the information to investigate the importance of specific parts of the enzyme to its function, and investigate the role of one domain of the enzyme in its function within cells. We describe the accomplishments towards these goals including many structures of the wild type and mutant forms of the enzyme that highlight its interactions with its Hg(II) substrate, elucidation of the role of the N-terminal domain in vitro and in vivo, and elucidation of the roles of at two conserved residues in the core in the mechanism of catalysis.

Pulmonary hypertension in wild type mice and animals with genetic deficit in KCa2.3 and KCa3.1 channels.

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Christine Wandall-Frostholm

Full Text Available In vascular biology, endothelial KCa2.3 and KCa3.1 channels contribute to arterial blood pressure regulation by producing membrane hyperpolarization and smooth muscle relaxation. The role of KCa2.3 and KCa3.1 channels in the pulmonary circulation is not fully established. Using mice with genetically encoded deficit of KCa2.3 and KCa3.1 channels, this study investigated the effect of loss of the channels in hypoxia-induced pulmonary hypertension.Male wild type and KCa3.1-/-/KCa2.3T/T(+DOX mice were exposed to chronic hypoxia for four weeks to induce pulmonary hypertension. The degree of pulmonary hypertension was evaluated by right ventricular pressure and assessment of right ventricular hypertrophy. Segments of pulmonary arteries were mounted in a wire myograph for functional studies and morphometric studies were performed on lung sections. Chronic hypoxia induced pulmonary hypertension, right ventricular hypertrophy, increased lung weight, and increased hematocrit levels in either genotype. The KCa3.1-/-/KCa2.3T/T(+DOX mice developed structural alterations in the heart with increased right ventricular wall thickness as well as in pulmonary vessels with increased lumen size in partially- and fully-muscularized vessels and decreased wall area, not seen in wild type mice. Exposure to chronic hypoxia up-regulated the gene expression of the KCa2.3 channel by twofold in wild type mice and increased by 2.5-fold the relaxation evoked by the KCa2.3 and KCa3.1 channel activator NS309, whereas the acetylcholine-induced relaxation - sensitive to the combination of KCa2.3 and KCa3.1 channel blockers, apamin and charybdotoxin - was reduced by 2.5-fold in chronic hypoxic mice of either genotype.Despite the deficits of the KCa2.3 and KCa3.1 channels failed to change hypoxia-induced pulmonary hypertension, the up-regulation of KCa2.3-gene expression and increased NS309-induced relaxation in wild-type mice point to a novel mechanism to counteract pulmonary

Wild type measles virus attenuation independent of type I IFN

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Horvat Branka

2008-02-01

Full Text Available Abstract Background Measles virus attenuation has been historically performed by adaptation to cell culture. The current dogma is that attenuated virus strains induce more type I IFN and are more resistant to IFN-induced protection than wild type (wt. Results The adaptation of a measles virus isolate (G954-PBL by 13 passages in Vero cells induced a strong attenuation of this strain in vivo. The adapted virus (G954-V13 differs from its parental strain by only 5 amino acids (4 in P/V/C and 1 in the M gene. While a vaccine strain, Edmonston Zagreb, could replicate equally well in various primate cells, both G954 strains exhibited restriction to the specific cell type used initially for their propagation. Surprisingly, we observed that both G954 strains induced type I IFN, the wt strain inducing even more than the attenuated ones, particularly in human plasmacytoid Dendritic Cells. Type I IFN-induced protection from the infection of both G954 strains depended on the cell type analyzed, being less efficient in the cells used to grow the viral strain. Conclusion Thus, mutations in M and P/V/C proteins can critically affect MV pathogenicity, cellular tropism and lead to virus attenuation without interfering with the α/β IFN system.

Chronic intermittent fasting improves cognitive functions and brain structures in mice.

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Liaoliao Li

Full Text Available Obesity is a major health issue. Obesity started from teenagers has become a major health concern in recent years. Intermittent fasting increases the life span. However, it is not known whether obesity and intermittent fasting affect brain functions and structures before brain aging. Here, we subjected 7-week old CD-1 wild type male mice to intermittent (alternate-day fasting or high fat diet (45% caloric supplied by fat for 11 months. Mice on intermittent fasting had better learning and memory assessed by the Barnes maze and fear conditioning, thicker CA1 pyramidal cell layer, higher expression of drebrin, a dendritic protein, and lower oxidative stress than mice that had free access to regular diet (control mice. Mice fed with high fat diet was obese and with hyperlipidemia. They also had poorer exercise tolerance. However, these obese mice did not present significant learning and memory impairment or changes in brain structures or oxidative stress compared with control mice. These results suggest that intermittent fasting improves brain functions and structures and that high fat diet feeding started early in life does not cause significant changes in brain functions and structures in obese middle-aged animals.

Chronic intermittent fasting improves cognitive functions and brain structures in mice.

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Li, Liaoliao; Wang, Zhi; Zuo, Zhiyi

2013-01-01

Obesity is a major health issue. Obesity started from teenagers has become a major health concern in recent years. Intermittent fasting increases the life span. However, it is not known whether obesity and intermittent fasting affect brain functions and structures before brain aging. Here, we subjected 7-week old CD-1 wild type male mice to intermittent (alternate-day) fasting or high fat diet (45% caloric supplied by fat) for 11 months. Mice on intermittent fasting had better learning and memory assessed by the Barnes maze and fear conditioning, thicker CA1 pyramidal cell layer, higher expression of drebrin, a dendritic protein, and lower oxidative stress than mice that had free access to regular diet (control mice). Mice fed with high fat diet was obese and with hyperlipidemia. They also had poorer exercise tolerance. However, these obese mice did not present significant learning and memory impairment or changes in brain structures or oxidative stress compared with control mice. These results suggest that intermittent fasting improves brain functions and structures and that high fat diet feeding started early in life does not cause significant changes in brain functions and structures in obese middle-aged animals.

Identifying the Integrated Neural Networks Involved in Capsaicin-Induced Pain Using fMRI in Awake TRPV1 Knockout and Wild-Type Rats

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Jason Richard Yee

2015-02-01

Full Text Available In the present study, we used functional MRI in awake rats to investigate the pain response that accompanies intradermal injection of capsaicin into the hindpaw. To this end, we used BOLD imaging together with a 3D segmented, annotated rat atlas and computational analysis to identify the integrated neural circuits involved in capsaicin-induced pain. The specificity of the pain response to capsaicin was tested in a transgenic model that contains a biallelic deletion of the gene encoding for the transient receptor potential cation channel subfamily V member 1 (TRPV1. Capsaicin is an exogenous ligand for the TRPV1 receptor, and in wild-type rats, activated the putative pain neural circuit. In addition, capsaicin-treated wild-type rats exhibited activation in brain regions comprising the Papez circuit and habenular system, systems that play important roles in the integration of emotional information, and learning and memory of aversive information, respectively. As expected, capsaicin administration to TRPV1-KO rats failed to elicit the robust BOLD activation pattern observed in wild-type controls. However, the intradermal injection of formalin elicited a significant activation of the putative pain pathway as represented by such areas as the anterior cingulate, somatosensory cortex, parabrachial nucleus, and periaqueductal gray. Notably, comparison of neural responses to capsaicin in wild-type versus knock-out rats uncovered evidence that capsaicin may function in an antinociceptive capacity independent of TRPV1 signaling. Our data suggest that neuroimaging of pain in awake, conscious animals has the potential to inform the neurobiological basis of full and integrated perceptions of pain.

Genetic relationships and epidemiological links between wild type 1 poliovirus isolates in Pakistan and Afghanistan.

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Angez, Mehar; Shaukat, Shahzad; Alam, Muhammad M; Sharif, Salmaan; Khurshid, Adnan; Zaidi, Syed Sohail Zahoor

2012-02-22

Efforts have been made to eliminate wild poliovirus transmission since 1988 when the World Health Organization began its global eradication campaign. Since then, the incidence of polio has decreased significantly. However, serotype 1 and serotype 3 still circulate endemically in Pakistan and Afghanistan. Both countries constitute a single epidemiologic block representing one of the three remaining major global reservoirs of poliovirus transmission. In this study we used genetic sequence data to investigate transmission links among viruses from diverse locations during 2005-2007. In order to find the origins and routes of wild type 1 poliovirus circulation, polioviruses were isolated from faecal samples of Acute Flaccid Paralysis (AFP) patients. We used viral cultures, two intratypic differentiation methods PCR, ELISA to characterize as vaccine or wild type 1 and nucleic acid sequencing of entire VP1 region of poliovirus genome to determine the genetic relatedness. One hundred eleven wild type 1 poliovirus isolates were subjected to nucleotide sequencing for genetic variation study. Considering the 15% divergence of the sequences from Sabin 1, Phylogenetic analysis by MEGA software revealed that active inter and intra country transmission of many genetically distinct strains of wild poliovirus type 1 belonged to genotype SOAS which is indigenous in this region. By grouping wild type 1 polioviruses according to nucleotide sequence homology, three distinct clusters A, B and C were obtained with multiple chains of transmission together with some silent circulations represented by orphan lineages. Our results emphasize that there was a persistent transmission of wild type 1 polioviruses in Pakistan and Afghanistan during 2005-2007. The epidemiologic information provided by the sequence data can contribute to the formulation of better strategies for poliomyelitis control to those critical areas, associated with high risk population groups which include migrants

Genetic relationships and epidemiological links between wild type 1 poliovirus isolates in Pakistan and Afghanistan

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Angez Mehar

2012-02-01

Full Text Available Abstract Background/Aim Efforts have been made to eliminate wild poliovirus transmission since 1988 when the World Health Organization began its global eradication campaign. Since then, the incidence of polio has decreased significantly. However, serotype 1 and serotype 3 still circulate endemically in Pakistan and Afghanistan. Both countries constitute a single epidemiologic block representing one of the three remaining major global reservoirs of poliovirus transmission. In this study we used genetic sequence data to investigate transmission links among viruses from diverse locations during 2005-2007. Methods In order to find the origins and routes of wild type 1 poliovirus circulation, polioviruses were isolated from faecal samples of Acute Flaccid Paralysis (AFP patients. We used viral cultures, two intratypic differentiation methods PCR, ELISA to characterize as vaccine or wild type 1 and nucleic acid sequencing of entire VP1 region of poliovirus genome to determine the genetic relatedness. Results One hundred eleven wild type 1 poliovirus isolates were subjected to nucleotide sequencing for genetic variation study. Considering the 15% divergence of the sequences from Sabin 1, Phylogenetic analysis by MEGA software revealed that active inter and intra country transmission of many genetically distinct strains of wild poliovirus type 1 belonged to genotype SOAS which is indigenous in this region. By grouping wild type 1 polioviruses according to nucleotide sequence homology, three distinct clusters A, B and C were obtained with multiple chains of transmission together with some silent circulations represented by orphan lineages. Conclusion Our results emphasize that there was a persistent transmission of wild type1 polioviruses in Pakistan and Afghanistan during 2005-2007. The epidemiologic information provided by the sequence data can contribute to the formulation of better strategies for poliomyelitis control to those critical areas

Reconstitution of wild type viral DNA in simian cells transfected with early and late SV40 defective genomes.

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O'Neill, F J; Gao, Y; Xu, X

1993-11-01

The DNAs of polyomaviruses ordinarily exist as a single circular molecule of approximately 5000 base pairs. Variants of SV40, BKV and JCV have been described which contain two complementing defective DNA molecules. These defectives, which form a bipartite genome structure, contain either the viral early region or the late region. The defectives have the unique property of being able to tolerate variable sized reiterations of regulatory and terminus region sequences, and portions of the coding region. They can also exchange coding region sequences with other polyomaviruses. It has been suggested that the bipartite genome structure might be a stage in the evolution of polyomaviruses which can uniquely sustain genome and sequence diversity. However, it is not known if the regulatory and terminus region sequences are highly mutable. Also, it is not known if the bipartite genome structure is reversible and what the conditions might be which would favor restoration of the monomolecular genome structure. We addressed the first question by sequencing the reiterated regulatory and terminus regions of E- and L-SV40 DNAs. This revealed a large number of mutations in the regulatory regions of the defective genomes, including deletions, insertions, rearrangements and base substitutions. We also detected insertions and base substitutions in the T-antigen gene. We addressed the second question by introducing into permissive simian cells, E- and L-SV40 genomes which had been engineered to contain only a single regulatory region. Analysis of viral DNA from transfected cells demonstrated recombined genomes containing a wild type monomolecular DNA structure. However, the complete defectives, containing reiterated regulatory regions, could often compete away the wild type genomes. The recombinant monomolecular genomes were isolated, cloned and found to be infectious. All of the DNA alterations identified in one of the regulatory regions of E-SV40 DNA were present in the recombinant

Supramolecular structure of enterobacterial wild-type lipopolysaccharides (LPS), fractions thereof, and their neutralization by Pep19-2.5.

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Brandenburg, Klaus; Heinbockel, Lena; Correa, Wilmar; Fukuoka, Satoshi; Gutsmann, Thomas; Zähringer, Ulrich; Koch, Michel H J

2016-04-01

Lipopolysaccharides (LPS) belong to the strongest immune-modulating compounds known in nature, and are often described as pathogen-associated molecular patterns (PAMPs). In particular, at higher concentrations they are responsible for sepsis and the septic shock syndrome associated with high lethality. Since most data are indicative that LPS aggregates are the bioactive units, their supramolecular structures are considered to be of outmost relevance for deciphering the molecular mechanisms of its bioactivity. So far, however, most of the data available addressing this issue, were published only for the lipid part (lipid A) and the core-oligosaccharide containing rough LPS, representing the bioactive unit. By contrast, it is well known that most of the LPS specimen identified in natural habitats contain the smooth-form (S-form) LPS, which carry additionally a high-molecular polysaccharide (O-chain). To fill this lacuna and going into a more natural system, here various wild-type (smooth form) LPS including also some LPS fractions were investigated by small-angle X-ray scattering with synchrotron radiation to analyze their aggregate structure. Furthermore, the influence of a recently designed synthetic anti-LPS peptide (SALP) Pep19-2.5 on the aggregate structure, on the binding thermodynamics, and on the cytokine-inducing activity of LPS were characterized, showing defined aggregate changes, high affinity binding and inhibition of cytokine secretion. The data obtained are suitable to refine our view on the preferences of LPS for non-lamellar structures, representing the highest bioactive forms which can be significantly influenced by the binding with neutralizing peptides such as Pep19-2.5. Copyright © 2016 Elsevier Inc. All rights reserved.

Investigating the structure and dynamics of the PIK3CA wild-type and H1047R oncogenic mutant.

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Paraskevi Gkeka

2014-10-01

Full Text Available The PIK3CA gene is one of the most frequently mutated oncogenes in human cancers. It encodes p110α, the catalytic subunit of phosphatidylinositol 3-kinase alpha (PI3Kα, which activates signaling cascades leading to cell proliferation, survival, and cell growth. The most frequent mutation in PIK3CA is H1047R, which results in enzymatic overactivation. Understanding how the H1047R mutation causes the enhanced activity of the protein in atomic detail is central to developing mutant-specific therapeutics for cancer. To this end, Surface Plasmon Resonance (SPR experiments and Molecular Dynamics (MD simulations were carried out for both wild-type (WT and H1047R mutant proteins. An expanded positive charge distribution on the membrane binding regions of the mutant with respect to the WT protein is observed through MD simulations, which justifies the increased ability of the mutated protein variant to bind to membranes rich in anionic lipids in our SPR experiments. Our results further support an auto-inhibitory role of the C-terminal tail in the WT protein, which is abolished in the mutant protein due to loss of crucial intermolecular interactions. Moreover, Functional Mode Analysis reveals that the H1047R mutation alters the twisting motion of the N-lobe of the kinase domain with respect to the C-lobe and shifts the position of the conserved P-loop residues in the vicinity of the active site. These findings demonstrate the dynamical and structural differences of the two proteins in atomic detail and propose a mechanism of overactivation for the mutant protein. The results may be further utilized for the design of mutant-specific PI3Kα inhibitors that exploit the altered mutant conformation.

Rearing in seawater mesocosms improves the spawning performance of growth hormone transgenic and wild-type coho salmon.

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Rosalind A Leggatt

Full Text Available Growth hormone (GH transgenes can significantly accelerate growth rates in fish and cause associated alterations to their physiology and behaviour. Concern exists regarding potential environmental risks of GH transgenic fish, should they enter natural ecosystems. In particular, whether they can reproduce and generate viable offspring under natural conditions is poorly understood. In previous studies, GH transgenic salmon grown under contained culture conditions had lower spawning behaviour and reproductive success relative to wild-type fish reared in nature. However, wild-type salmon cultured in equal conditions also had limited reproductive success. As such, whether decreased reproductive success of GH transgenic salmon is due to the action of the transgene or to secondary effects of culture (or a combination has not been fully ascertained. Hence, salmon were reared in large (350,000 L, semi-natural, seawater tanks (termed mesocosms designed to minimize effects of standard laboratory culture conditions, and the reproductive success of wild-type and GH transgenic coho salmon from mesocosms were compared with that of wild-type fish from nature. Mesocosm rearing partially restored spawning behaviour and success of wild-type fish relative to culture rearing, but remained lower overall than those reared in nature. GH transgenic salmon reared in the mesocosm had similar spawning behaviour and success as wild-type fish reared in the mesocosm when in full competition and without competition, but had lower success in male-only competition experiments. There was evidence of genotype×environmental interactions on spawning success, so that spawning success of transgenic fish, should they escape to natural systems in early life, cannot be predicted with low uncertainty. Under the present conditions, we found no evidence to support enhanced mating capabilities of GH transgenic coho salmon compared to wild-type salmon. However, it is clear that GH transgenic

Dietary Supplementation of Hericium erinaceus Increases Mossy Fiber-CA3 Hippocampal Neurotransmission and Recognition Memory in Wild-Type Mice

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Federico Brandalise

2017-01-01

Full Text Available Hericium erinaceus (Bull. Pers. is a medicinal mushroom capable of inducing a large number of modulatory effects on human physiology ranging from the strengthening of the immune system to the improvement of cognitive functions. In mice, dietary supplementation with H. erinaceus prevents the impairment of spatial short-term and visual recognition memory in an Alzheimer model. Intriguingly other neurobiological effects have recently been reported like the effect on neurite outgrowth and differentiation in PC12 cells. Until now no investigations have been conducted to assess the impact of this dietary supplementation on brain function in healthy subjects. Therefore, we have faced the problem by considering the effect on cognitive skills and on hippocampal neurotransmission in wild-type mice. In wild-type mice the oral supplementation with H. erinaceus induces, in behaviour test, a significant improvement in the recognition memory and, in hippocampal slices, an increase in spontaneous and evoked excitatory synaptic current in mossy fiber-CA3 synapse. In conclusion, we have produced a series of findings in support of the concept that H. erinaceus induces a boost effect onto neuronal functions also in nonpathological conditions.

Dietary Supplementation of Hericium erinaceus Increases Mossy Fiber-CA3 Hippocampal Neurotransmission and Recognition Memory in Wild-Type Mice.

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Brandalise, Federico; Cesaroni, Valentina; Gregori, Andrej; Repetti, Margherita; Romano, Chiara; Orrù, Germano; Botta, Laura; Girometta, Carolina; Guglielminetti, Maria Lidia; Savino, Elena; Rossi, Paola

2017-01-01

Hericium erinaceus (Bull.) Pers. is a medicinal mushroom capable of inducing a large number of modulatory effects on human physiology ranging from the strengthening of the immune system to the improvement of cognitive functions. In mice, dietary supplementation with H. erinaceus prevents the impairment of spatial short-term and visual recognition memory in an Alzheimer model. Intriguingly other neurobiological effects have recently been reported like the effect on neurite outgrowth and differentiation in PC12 cells. Until now no investigations have been conducted to assess the impact of this dietary supplementation on brain function in healthy subjects. Therefore, we have faced the problem by considering the effect on cognitive skills and on hippocampal neurotransmission in wild-type mice. In wild-type mice the oral supplementation with H. erinaceus induces, in behaviour test, a significant improvement in the recognition memory and, in hippocampal slices, an increase in spontaneous and evoked excitatory synaptic current in mossy fiber-CA3 synapse. In conclusion, we have produced a series of findings in support of the concept that H. erinaceus induces a boost effect onto neuronal functions also in nonpathological conditions.

The wild type as concept and in experimental practice: A history of its role in classical genetics and evolutionary theory.

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Holmes, Tarquin

2017-06-01

Wild types in genetics are specialised strains of laboratory experimental organism which principally serve as standards against which variation is measured. As selectively inbred lineages highly isolated from ancestral wild populations, there appears to be little wild or typical about them. I will nonetheless argue that they have historically been successfully used as stand-ins for nature, allowing knowledge produced in the laboratory to be extrapolated to the natural world. In this paper, I will explore the 19th century origins of the wild type concept, the theoretical and experimental innovations which allowed concepts and organisms to move from wild nature to laboratory domestication c. 1900 (resulting in the production of standardised lab strains), and the conflict among early geneticists between interactionist and atomist accounts of wild type, which would eventually lead to the conceptual disintegration of wild types and the triumph of genocentrism and population genetics. I conclude by discussing how the strategy of using wild type strains to represent nature in the lab has nonetheless survived the downfall of the wild type concept and continues to provide, significant limitations acknowledged, an epistemically productive means of investigating heredity and evolutionary variation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Fusarium spp. Associated with Field-Grown Grain of Near-Isogenic Low Lignin and Wild-Type Sorghum

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Fusarium spp. associated with field-grown grain of near-isogenic low lignin and wild-type sorghum. Deanna Funnell-Harris and Jeff Pedersen, USDA-ARS, Lincoln, NE Previous studies indicated that low lignin brown midrib (bmr) sorghum may be more resistant to Fusarium spp. than wild-type and that phen...

Transplacental and oral transmission of wild-type bluetongue virus serotype 8 in cattle after experimental infection

NARCIS (Netherlands)

Backx, A.; Heutink, C.G.; Rooij, van E.M.A.; Rijn, van P.A.

2009-01-01

Potential vertical transmission of wild-type bluetongue virus serotype 8 (BTV-8) in cattle was explored in this experiment. We demonstrated transplacental transmission of wild-type BTV-8 in one calf and oral infection with BTV-8 in another calf. Following the experimental BTV-8 infection of seven

Structural and functional studies of conserved nucleotide-binding protein LptB in lipopolysaccharide transport

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Wang, Zhongshan [Biomedical Research Centre, Norwich Medical School, University of East Anglia, Norwich Research Park, NR4 7TJ (United Kingdom); College of Life Sciences, Sichuan University, Chengdu 610065 (China); Biomedical Sciences Research Complex, School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST (United Kingdom); Xiang, Quanju [College of Life Sciences, Sichuan University, Chengdu 610065 (China); Biomedical Sciences Research Complex, School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST (United Kingdom); Department of Microbiology, College of Resource and Environment Science, Sichuan Agriculture University, Yaan 625000 (China); Zhu, Xiaofeng [College of Life Sciences, Sichuan University, Chengdu 610065 (China); Dong, Haohao [Biomedical Sciences Research Complex, School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST (United Kingdom); He, Chuan [School of Electronics and Information, Wuhan Technical College of Communications, No. 6 Huangjiahu West Road, Hongshan District, Wuhan, Hubei 430065 (China); Wang, Haiyan; Zhang, Yizheng [College of Life Sciences, Sichuan University, Chengdu 610065 (China); Wang, Wenjian, E-mail: Wenjian166@gmail.com [Laboratory of Department of Surgery, The First Affiliated Hospital, Sun Yat-sen University, 58 Zhongshan Road II, Guangzhou, Guangdong 510080 (China); Dong, Changjiang, E-mail: C.Dong@uea.ac.uk [Biomedical Research Centre, Norwich Medical School, University of East Anglia, Norwich Research Park, NR4 7TJ (United Kingdom)

2014-09-26

Highlights: • Determination of the structure of the wild-type LptB in complex with ATP and Mg{sup 2+}. • Demonstrated that ATP binding residues are essential for LptB’s ATPase activity and LPS transport. • Dimerization is required for the LptB’s function and LPS transport. • Revealed relationship between activity of the LptB and the vitality of E. coli cells. - Abstract: Lipopolysaccharide (LPS) is the main component of the outer membrane of Gram-negative bacteria, which plays an essential role in protecting the bacteria from harsh conditions and antibiotics. LPS molecules are transported from the inner membrane to the outer membrane by seven LPS transport proteins. LptB is vital in hydrolyzing ATP to provide energy for LPS transport, however this mechanism is not very clear. Here we report wild-type LptB crystal structure in complex with ATP and Mg{sup 2+}, which reveals that its structure is conserved with other nucleotide-binding proteins (NBD). Structural, functional and electron microscopic studies demonstrated that the ATP binding residues, including K42 and T43, are crucial for LptB’s ATPase activity, LPS transport and the vitality of Escherichia coli cells with the exceptions of H195A and Q85A; the H195A mutation does not lower its ATPase activity but impairs LPS transport, and Q85A does not alter ATPase activity but causes cell death. Our data also suggest that two protomers of LptB have to work together for ATP hydrolysis and LPS transport. These results have significant impacts in understanding the LPS transport mechanism and developing new antibiotics.

Structural and functional studies of conserved nucleotide-binding protein LptB in lipopolysaccharide transport

International Nuclear Information System (INIS)

Wang, Zhongshan; Xiang, Quanju; Zhu, Xiaofeng; Dong, Haohao; He, Chuan; Wang, Haiyan; Zhang, Yizheng; Wang, Wenjian; Dong, Changjiang

2014-01-01

Highlights: • Determination of the structure of the wild-type LptB in complex with ATP and Mg 2+ . • Demonstrated that ATP binding residues are essential for LptB’s ATPase activity and LPS transport. • Dimerization is required for the LptB’s function and LPS transport. • Revealed relationship between activity of the LptB and the vitality of E. coli cells. - Abstract: Lipopolysaccharide (LPS) is the main component of the outer membrane of Gram-negative bacteria, which plays an essential role in protecting the bacteria from harsh conditions and antibiotics. LPS molecules are transported from the inner membrane to the outer membrane by seven LPS transport proteins. LptB is vital in hydrolyzing ATP to provide energy for LPS transport, however this mechanism is not very clear. Here we report wild-type LptB crystal structure in complex with ATP and Mg 2+ , which reveals that its structure is conserved with other nucleotide-binding proteins (NBD). Structural, functional and electron microscopic studies demonstrated that the ATP binding residues, including K42 and T43, are crucial for LptB’s ATPase activity, LPS transport and the vitality of Escherichia coli cells with the exceptions of H195A and Q85A; the H195A mutation does not lower its ATPase activity but impairs LPS transport, and Q85A does not alter ATPase activity but causes cell death. Our data also suggest that two protomers of LptB have to work together for ATP hydrolysis and LPS transport. These results have significant impacts in understanding the LPS transport mechanism and developing new antibiotics

Wild-type and A315T mutant TDP-43 exert differential neurotoxicity in a Drosophila model of ALS

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Estes, Patricia S.; Boehringer, Ashley; Zwick, Rebecca; Tang, Jonathan E.; Grigsby, Brianna; Zarnescu, Daniela C.

2011-01-01

The RNA-binding protein TDP-43 has been linked to amyotrophic lateral sclerosis (ALS) both as a causative locus and as a marker of pathology. With several missense mutations being identified within TDP-43, efforts have been directed towards generating animal models of ALS in mouse, zebrafish, Drosophila and worms. Previous loss of function and overexpression studies have shown that alterations in TDP-43 dosage recapitulate hallmark features of ALS pathology, including neuronal loss and locomotor dysfunction. Here we report a direct in vivo comparison between wild-type and A315T mutant TDP-43 overexpression in Drosophila neurons. We found that when expressed at comparable levels, wild-type TDP-43 exerts more severe effects on neuromuscular junction architecture, viability and motor neuron loss compared with the A315T allele. A subset of these differences can be compensated by higher levels of A315T expression, indicating a direct correlation between dosage and neurotoxic phenotypes. Interestingly, larval locomotion is the sole parameter that is more affected by the A315T allele than wild-type TDP-43. RNA interference and genetic interaction experiments indicate that TDP-43 overexpression mimics a loss-of-function phenotype and suggest a dominant-negative effect. Furthermore, we show that neuronal apoptosis does not require the cytoplasmic localization of TDP-43 and that its neurotoxicity is modulated by the proteasome, the HSP70 chaperone and the apoptosis pathway. Taken together, our findings provide novel insights into the phenotypic consequences of the A315T TDP-43 missense mutation and suggest that studies of individual mutations are critical for elucidating the molecular mechanisms of ALS and related neurodegenerative disorders. PMID:21441568

The effect of UVB on flavonoid biosynthesis in wild type and mutant petunia and arabidopsis

International Nuclear Information System (INIS)

Ryan, K.G.; Swinny, E.E.; Markham, K.R.; Winefield, C.

2000-01-01

Full text: Flavonoids may protect plants against damage by UVB radiation. Flavonoid composition and mRNA expression were determined following growth of plants under natural light, and under natural light with low UVB and with enhanced UVB. In wild-type Arabidopsis and Petunia, UVB induced an increase in total levels of flavonols and this was due to an up-regulation of, several genes coding for key enzymes in the phenylpropanoid pathway. In addition, UVB induced a higher rate of production of the di-hydroxylated si flavonol, quercetin glycoside than of the mono-hydroxylated equivalent, of kaempferol glycoside. Thus the ratio of quercetin to kaempferol increased with UVB treatment in wild type plants, and this suggests that the flavonoid r 3'hydroxylase (F3'H) enzyme, which converts dihydrokaempferol to dihydroquercetin, may play a key role in plant protection from UVB. Mutant plants of both species lacking this F3'H gene were grown under similar UV conditions. Leaves of the mutant Arabidopsis plant (tt7) did not contain quercetin, even under the enhanced UVB treatment. Under the low UVB treatment the total amount of flavonol was similar to the wild-type (Ler), but with increasing UVB, total flavonol (i.e. kaempferol) levels were significantly higher than in similarly treated wild type plants. In the Petunia F3'H mutant, low levels of quercetin were found even in the low UVB treatment, which indicates this variety may be producing some quercetin via an alternative pathway. Under UVB radiation, total flavonoids increased to levels significantly higher than in similarly treated wild type plants, and most of this material was kaempferol. These observations suggest that quercetin is the preferred protective flavonol in wild type plants, due perhaps to enhanced antioxidant or free radical scavenging activity. In mutant plants lacking the F3'H enzyme, the response is to produce a larger amount of a less effective photoprotectant

Explaining the resurgent popularity of the wild: motivations for wild plant gathering in the Biosphere Reserve Grosses Walsertal, Austria.

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Schunko, Christoph; Grasser, Susanne; Vogl, Christian R

2015-06-30

Wild plant gathering becomes again a popular and fashionable activity in Europe after gathering practices have been increasingly abandoned over the last decades. Recent ethnobotanical research documented a diversity of gathering practices from people of diverse socio-economic and cultural backgrounds who gather in urban and rural areas. Few efforts were though made to study the motivations for gathering wild plants and to understand the resurgent popularity of wild plant gathering. This paper addresses the following research questions: (1) which motivations activate wild plant gatherers? (2) which motivation-types of gatherers exist in the Grosses Walsertal? (3) how do the motivations for gathering relate to the socio-demographic background of gatherers? Field research was conducted in the Grosses Walsertal, Austria in the years 2008 and 2009 in two field research periods. Thirty-six local farmers were first interviewed with semi-structured interviews. The motivations identified in these interviews were then included in a structured questionnaire, which was used to interview 353 residents of the valley. Pupils of local schools participated in the data collection as interviewers. Principal Component Analysis was used to categorize the motivations and to identify motivation-types of wild plant gatherers. Generalized Linear Models were calculated to identify relations between motivations and the socio-demographic background of gatherers. The respondents listed 13 different motivations for gathering wild plants and four motivations for not gathering. These 17 motivations were grouped in five motivation-types of wild plant gatherers, which are in decreasing importance: product quality, fun, tradition, not-gathering, income. Women, older respondents and homegardeners gather wild plants more often for fun; older respondents gather more often for maintaining traditions; non-homegardeners more frequently mention motivations for not gathering. The resurgent popularity of

Effects of regionally applied heating on the respiration of wild type ...

African Journals Online (AJOL)

Nocturnal dark respiration (Rn) in wild type and transgenic soybean plants ... Illinois, USA under ambient and elevated CO2 conditions was examined in this study. ... Experimental plants were transferred to a controlled growth chamber at V4 ...

Somatic loss of function mutations in neurofibromin 1 and MYC associated factor X genes identified by exome-wide sequencing in a wild-type GIST case

International Nuclear Information System (INIS)

Belinsky, Martin G.; Rink, Lori; Cai, Kathy Q.; Capuzzi, Stephen J.; Hoang, Yen; Chien, Jeremy; Godwin, Andrew K.; Mehren, Margaret von

2015-01-01

Approximately 10–15 % of gastrointestinal stromal tumors (GISTs) lack gain of function mutations in the KIT and platelet-derived growth factor receptor alpha (PDGFRA) genes. An alternate mechanism of oncogenesis through loss of function of the succinate-dehydrogenase (SDH) enzyme complex has been identified for a subset of these “wild type” GISTs. Paired tumor and normal DNA from an SDH-intact wild-type GIST case was subjected to whole exome sequencing to identify the pathogenic mechanism(s) in this tumor. Selected findings were further investigated in panels of GIST tumors through Sanger DNA sequencing, quantitative real-time PCR, and immunohistochemical approaches. A hemizygous frameshift mutation (p.His2261Leufs*4), in the neurofibromin 1 (NF1) gene was identified in the patient’s GIST; however, no germline NF1 mutation was found. A somatic frameshift mutation (p.Lys54Argfs*31) in the MYC associated factor X (MAX) gene was also identified. Immunohistochemical analysis for MAX on a large panel of GISTs identified loss of MAX expression in the MAX-mutated GIST and in a subset of mainly KIT-mutated tumors. This study suggests that inactivating NF1 mutations outside the context of neurofibromatosis may be the oncogenic mechanism for a subset of sporadic GIST. In addition, loss of function mutation of the MAX gene was identified for the first time in GIST, and a broader role for MAX in GIST progression was suggested. The online version of this article (doi:10.1186/s12885-015-1872-y) contains supplementary material, which is available to authorized users

Comparative analysis of the protein compositions between wild type and body color mutant of helicoverpa armigera adult

International Nuclear Information System (INIS)

He Lihua; Chen Jin'e; Liu Yan; Wang Yongqiang; Liu Peigang; Meng Zhiqi

2012-01-01

To gain an in-depth understanding of the fineness and regulation mechanism of body color mutant of Helicoverpa armigera Hbner, the protein composition differences between adult of dominant mutant, recessive mutant and wild type were studied using the SDS-PAGE combined with MALDI-TOF-TOF/MS and bioinformatics analysis. The results indicated that the protein composition of the dominant mutant and wild type had little difference. However, there were obvious differences between the recessive mutant and wild-type. Three specific stripe were chosen for mass spectrometry and bioinformatics analysis, and two types of proteins related to energy metabolism and cytoskeleton were identified. These findings suggested that the two types of proteins may be associated with occurrence and regulation of body color mutant traits of H. armigera. (authors)

Wild-type MIC distributions for aminoglycoside and cyclic polypeptide antibiotics used for treatment of Mycobacterium tuberculosis infections.

Science.gov (United States)

Juréen, P; Angeby, K; Sturegård, E; Chryssanthou, E; Giske, C G; Werngren, J; Nordvall, M; Johansson, A; Kahlmeter, G; Hoffner, S; Schön, T

2010-05-01

The aminoglycosides and cyclic polypeptides are essential drugs in the treatment of multidrug-resistant tuberculosis, underscoring the need for accurate and reproducible drug susceptibility testing (DST). The epidemiological cutoff value (ECOFF) separating wild-type susceptible strains from non-wild-type strains is an important but rarely used tool for indicating susceptibility breakpoints against Mycobacterium tuberculosis. In this study, we established wild-type MIC distributions on Middlebrook 7H10 medium for amikacin, kanamycin, streptomycin, capreomycin, and viomycin using 90 consecutive clinical isolates and 21 resistant strains. Overall, the MIC variation between and within runs did not exceed +/-1 MIC dilution step, and validation of MIC values in Bactec 960 MGIT demonstrated good agreement. Tentative ECOFFs defining the wild type were established for all investigated drugs, including amikacin and viomycin, which currently lack susceptibility breakpoints for 7H10. Five out of seven amikacin- and kanamycin-resistant isolates were classified as susceptible to capreomycin according to the current critical concentration (10 mg/liter) but were non-wild type according to the ECOFF (4 mg/liter), suggesting that the critical concentration may be too high. All amikacin- and kanamycin-resistant isolates were clearly below the ECOFF for viomycin, and two of them were below the ECOFF for streptomycin, indicating that these two drugs may be considered for treatment of amikacin-resistant strains. Pharmacodynamic indices (peak serum concentration [Cmax]/MIC) were more favorable for amikacin and viomycin compared to kanamycin and capreomycin. In conclusion, our data emphasize the importance of establishing wild-type MIC distributions for improving the quality of drug susceptibility testing against Mycobacterium tuberculosis.

Sabin and wild type polioviruses from children who presented with acute flaccid paralysis in Nigeria.

Science.gov (United States)

Adedeji, A O; Okonko, I O; Adu, F D

2012-09-01

Sensitive poliovirus surveillance to detect vaccine-derived-polioviruses will continue to increase in importance. Isolating and identifying poliovirus strains from children of pediatrics age in Nigeria. A total of 120 fecal samples were randomly collected from children under the age of five who presented with acute flaccid paralysis. Samples were tested by tissue culture technique and further characterized by intratypic differentiation testing using ELISA and PCR methods. The study confirmed the presence of 22(18.3%) enteroviral isolates comprising 19(86.4%) polioviruses and 3(13.6%) non-polio enteroviruses. These 19 polioviruses include: Sabin-type poliovirus-1 (15.8%), poliovirus-2 (10.5%), poliovirus-3 (10.5%) and wild-type poliovirus-1 (63.2%) isolates. It showed that poliovirus infection was higher in children ages 6-11 months (18.9%), females (18.4%), northern states (91.0%) with no vaccination record (75.0%). Wild-type poliovirus-1 was isolated from the stool samples of 12(54.6%) children from northern states and in all age groups except 18-23 months. No significant differences (P >0.05) between poliovirus infection and age (18.9% vs. 17.7%; 81.9% vs. 18.2%) and sex (18.3% vs. 18.4%). There was significant differences (Pvaccination (75.0% vs. 0.0%). No wild-type poliovirus was found in those with complete vaccination. This study further confirms the presence of Sabin and wild-type poliovirus among children in Nigeria. The isolation of Sabin strain of poliovirus is advantageous to the polio eradication program as it is capable of inducing natural immunity in susceptible hosts. Transmission of wild-type poliovirus among children with incomplete vaccination poses a serious threat to polio eradication program in Nigeria. Environmental and serological surveillance with larger sample size are important for monitoring poliovirus circulation in Nigeria.

Repair of gamma radiation damage in wild type and a radiation sensitive mutant of Deinococcus radiodurans

International Nuclear Information System (INIS)

Mizuma, Nagayo

1989-01-01

In an effort to examine production and repair of radiation-induced single and double strand breaks in the DNA, a repair-deficient wild type and a repair-deficient mutant, UV17, of Deinococcus radiodurans were subjected to Co-60 gamma irradiation at a dose rate of 6.3 kGy/hr for wild type and 3.9 kGy/hr for UV17 mutant. The shoulder of the curve of UV17 mutant was narrow but existed with the intercept of 0.7 kGy and the corresponding value of the wild type was 4.2 kGy. Mutant cells exhibited about 6 fold increases in sensitivity for the shoulder relative to the wild type. The D 37 doses in the wild type and the mutant were 0.57 kGy and 0.25 kGy, respectively. From the survival curves, difference in the sensitivity between two strains was mainly due to difference of repair capacity than the number of radiation sensitive target. Sedimentation rate of the main component in the irradiated cells of UV17 mutant increased almost to the level of unirradiated control by the postincubation at 30deg C for 3 hrs. The results indicated that this sensitive mutant also exhibited an ability to restore single strand breaks after exposure to a sublethal dose of 0.6 kGy. When restitution of double strand breaks was analyzed by sedimentation in a neutral sucrose gradient, the wild type showed restitution to DNA-membrane complex from large part of the breaks. For UV17 mutant, the apparent increase in DNA-membrane complex formation was seen after 3 hours incubation. Large part of the decrease in the activities of peak 2 was recovered in the peak 1 for the wild type. For the mutant, there was little restitution to peak 1. Almost free DNA component in UV17 mutant, therefore, was merely degraded into shorter pieces. Restoration of DNA-membrane complex from free DNA derived from gamma-ray induced double strand scission involved closely in the repair of gamma-induced damage and survival. (N.K.)

Photomorphogenic responses to UV radiation III: a comparative study of UVB effects on anthocyanin and flavonoid accumulation in wild-type and aurea mutant of tomato (Lycopersicon esculentum Mill.)

International Nuclear Information System (INIS)

Brandt, K.; Giannini, A.; Lercari, B.

1995-01-01

The UV-mediated induction of anthocyanin and UV-absorbing compounds was characterized in etiolated hypocotyls of wild-type and aurea (au) mutant tomato seedlings. Ultraviolet radiation induced significant increases of anthocyanin and UV-absorbing compounds in hypocotyls of the au mutant and of its isogenic wild-type, but the differences in the time courses of UV-induced pigment accumulation indicate that different photoregulatory mechanisms are involved for each of these two groups of pigments. It appears that prolonged presence of adequate levels of UVB (290-320 nm) energy and consequently the action of a specific UVB photoreceptor are indispensable for the photoinduction of anthocyanin accumulation in UV-irradiated hypocotyl of the au mutant that is missing the labile phytochrome pool. The large difference found between the wild-type and the au mutant strongly indicate the involvement of labile phytochrome as the primary functional photoreceptor for the photoinduction of anthocyanin accumulation in wild-type tomato hypocotyls. (author)

Function and Evolutionary Origin of Unicellular Camera-Type Eye Structure

KAUST Repository

Hayakawa, Shiho

2015-03-03

The ocelloid is an extraordinary eyespot organelle found only in the dinoflagellate family Warnowiaceae. It contains retina- and lens-like structures called the retinal body and the hyalosome. The ocelloid has been an evolutionary enigma because of its remarkable resemblance to the multicellular camera-type eye. To determine if the ocelloid is functionally photoreceptive, we investigated the warnowiid dinoflagellate Erythropsidinium. Here, we show that the morphology of the retinal body changed depending on different illumination conditions and the hyalosome manifests the refractile nature. Identifying a rhodopsin gene fragment in Erythropsidinium ESTs that is expressed in the retinal body by in situ hybridization, we also show that ocelloids are actually light sensitive photoreceptors. The rhodopsin gene identified is most closely related to bacterial rhodopsins. Taken together, we suggest that the ocelloid is an intracellular camera-type eye, which might be originated from endosymbiotic origin. © 2015 Hayakawa et al.

Function and Evolutionary Origin of Unicellular Camera-Type Eye Structure

KAUST Repository

Hayakawa, Shiho; Takaku, Yasuharu; Hwang, Jung Shan; Horiguchi, Takeo; Suga, Hiroshi; Gehring, Walter; Ikeo, Kazuho; Gojobori, Takashi

2015-01-01

The ocelloid is an extraordinary eyespot organelle found only in the dinoflagellate family Warnowiaceae. It contains retina- and lens-like structures called the retinal body and the hyalosome. The ocelloid has been an evolutionary enigma because of its remarkable resemblance to the multicellular camera-type eye. To determine if the ocelloid is functionally photoreceptive, we investigated the warnowiid dinoflagellate Erythropsidinium. Here, we show that the morphology of the retinal body changed depending on different illumination conditions and the hyalosome manifests the refractile nature. Identifying a rhodopsin gene fragment in Erythropsidinium ESTs that is expressed in the retinal body by in situ hybridization, we also show that ocelloids are actually light sensitive photoreceptors. The rhodopsin gene identified is most closely related to bacterial rhodopsins. Taken together, we suggest that the ocelloid is an intracellular camera-type eye, which might be originated from endosymbiotic origin. © 2015 Hayakawa et al.

Genetic recombination of tick-borne flaviviruses among wild-type strains.

Science.gov (United States)

Norberg, Peter; Roth, Anette; Bergström, Tomas

2013-06-05

Genetic recombination has been suggested to occur in mosquito-borne flaviviruses. In contrast, tick-borne flaviviruses have been thought to evolve in a clonal manner, although recent studies suggest that recombination occurs also for these viruses. We re-analyzed the data and found that previous conclusions on wild type recombination were probably falsely drawn due to misalignments of nucleotide sequences, ambiguities in GenBank sequences, or different laboratory culture histories suggestive of recombination events in laboratory. To evaluate if reliable predictions of wild type recombination of tick-borne flaviviruses can be made, we analyzed viral strains sequenced exclusively for this study, and other flavivirus sequences retrieved from GenBank. We detected genetic signals supporting recombination between viruses within the three clades of TBEV-Eu, TBEV-Sib and TBEV-Fe, respectively. Our results suggest that the tick-borne encephalitis viruses may undergo recombination under natural conditions, but that geographic barriers restrict most recombination events to involve only closely genetically related viruses. Copyright © 2013 Elsevier Inc. All rights reserved.

Wild-Type MIC Distributions for Aminoglycoside and Cyclic Polypeptide Antibiotics Used for Treatment of Mycobacterium tuberculosis Infections▿

Science.gov (United States)

Juréen, P.; Ängeby, K.; Sturegård, E.; Chryssanthou, E.; Giske, C. G.; Werngren, J.; Nordvall, M.; Johansson, A.; Kahlmeter, G.; Hoffner, S.; Schön, T.

2010-01-01

The aminoglycosides and cyclic polypeptides are essential drugs in the treatment of multidrug-resistant tuberculosis, underscoring the need for accurate and reproducible drug susceptibility testing (DST). The epidemiological cutoff value (ECOFF) separating wild-type susceptible strains from non-wild-type strains is an important but rarely used tool for indicating susceptibility breakpoints against Mycobacterium tuberculosis. In this study, we established wild-type MIC distributions on Middlebrook 7H10 medium for amikacin, kanamycin, streptomycin, capreomycin, and viomycin using 90 consecutive clinical isolates and 21 resistant strains. Overall, the MIC variation between and within runs did not exceed ±1 MIC dilution step, and validation of MIC values in Bactec 960 MGIT demonstrated good agreement. Tentative ECOFFs defining the wild type were established for all investigated drugs, including amikacin and viomycin, which currently lack susceptibility breakpoints for 7H10. Five out of seven amikacin- and kanamycin-resistant isolates were classified as susceptible to capreomycin according to the current critical concentration (10 mg/liter) but were non-wild type according to the ECOFF (4 mg/liter), suggesting that the critical concentration may be too high. All amikacin- and kanamycin-resistant isolates were clearly below the ECOFF for viomycin, and two of them were below the ECOFF for streptomycin, indicating that these two drugs may be considered for treatment of amikacin-resistant strains. Pharmacodynamic indices (peak serum concentration [Cmax]/MIC) were more favorable for amikacin and viomycin compared to kanamycin and capreomycin. In conclusion, our data emphasize the importance of establishing wild-type MIC distributions for improving the quality of drug susceptibility testing against Mycobacterium tuberculosis. PMID:20237102

Research on the ultrafast fluorescence property of thylakoid membranes of the wild-type and mutant rice

Science.gov (United States)

Ren, Zhao-Yu; Xu, Xiao-Ming; Wang, Shui-Cai; Xin, Yue-Yong; He, Jun-Fang; Hou, Xun

2003-10-01

A high yielding rice variety mutant (Oryza sativa L., Zhenhui 249) with low chlorophyll b (Chl b) has been discovered in natural fields. It has a quality character controlled by a pair of recessive genes (nuclear gene). The partial loss of Chl b in content affects the efficiency of light harvest in a light harvest complex (LHC), thus producing the difference of the exciting energy transfer and the efficiency of photochemistry conversion between the mutant and wild-type rice in photosynthetic unit. The efficiency of utilizing light energy is higher in the mutant than that in the wild-type rice relatively. For further discussion of the above-mentioned difference and learning about the mechanism of the increase in the photochemical efficiency of the mutant, the pico-second resolution fluorescence spectrum measurement with delay-frame-scanning single photon counting technique is adopted. Thylakoid membranes of the mutant and the wild-type rice are excited by an Ar+ laser with a pulse width of 120 ps, repetition rate of 4 MHz and wavelength of 514 nm. Compared with the time and spectrum property of exciting fluorescence, conclusions of those ultrafast dynamic experiments are: 1) The speeds of the exciting energy transferred in photo-system I are faster than that in photo-system II in both samples. 2) The speeds of the exciting energy transfer of mutant sample are faster than those of the wild-type. This might be one of the major reasons why the efficiency of photosynthesis is higher in mutant than that in the wild-type rice.

Structural and functional characterization of the product of disease-related factor H gene conversion.

Science.gov (United States)

Herbert, Andrew P; Kavanagh, David; Johansson, Conny; Morgan, Hugh P; Blaum, Bärbel S; Hannan, Jonathan P; Barlow, Paul N; Uhrín, Dušan

2012-03-06

Numerous complement factor H (FH) mutations predispose patients to atypical hemolytic uremic syndrome (aHUS) and other disorders arising from inadequately regulated complement activation. No unifying structural or mechanistic consequences have been ascribed to these mutants beyond impaired self-cell protection. The S1191L and V1197A mutations toward the C-terminus of FH, which occur in patients singly or together, arose from gene conversion between CFH encoding FH and CFHR1 encoding FH-related 1. We show that neither single nor double mutations structurally perturbed recombinant proteins consisting of the FH C-terminal modules, 19 and 20 (FH19-20), although all three FH19-20 mutants were poor, compared to wild-type FH19-20, at promoting hemolysis of C3b-coated erythrocytes through competition with full-length FH. Indeed, our new crystal structure of the S1191L mutant of FH19-20 complexed with an activation-specific complement fragment, C3d, was nearly identical to that of the wild-type FH19-20:C3d complex, consistent with mutants binding to C3b with wild-type-like affinity. The S1191L mutation enhanced thermal stability of module 20, whereas the V1197A mutation dramatically decreased it. Thus, although mutant proteins were folded at 37 °C, they differ in conformational rigidity. Neither single substitutions nor double substitutions increased measurably the extent of FH19-20 self-association, nor did these mutations significantly affect the affinity of FH19-20 for three glycosaminoglycans, despite critical roles of module 20 in recognizing polyanionic self-surface markers. Unexpectedly, FH19-20 mutants containing Leu1191 self-associated on a heparin-coated surface to a higher degree than on surfaces coated with dermatan or chondroitin sulfates. Thus, potentially disease-related functional distinctions between mutants, and between FH and FH-related 1, may manifest in the presence of specific glycosaminoglycans.

Crystallization and preliminary X-ray diffraction analysis of the wild-type haloalkane dehalogenase DhaA and its variant DhaA13 complexed with different ligands

International Nuclear Information System (INIS)

Stsiapanava, Alena; Chaloupkova, Radka; Fortova, Andrea; Brynda, Jiri; Weiss, Manfred S.; Damborsky, Jiri; Kuta Smatanova, Ivana

2011-01-01

Crystals of the wild-type haloalkane dehalogenase DhaA derived from R. rhodochrous NCIMB 13064 and of its catalytically inactive variant DhaA13 were grown in the presence of various ligands and diffraction data were collected to high and atomic resolution. Haloalkane dehalogenases make up an important class of hydrolytic enzymes which catalyse the cleavage of carbon–halogen bonds in halogenated aliphatic compounds. There is growing interest in these enzymes owing to their potential use in environmental and industrial applications. The haloalkane dehalogenase DhaA from Rhodococcus rhodochrous NCIMB 13064 can slowly detoxify the industrial pollutant 1,2,3-trichloropropane (TCP). Structural analysis of this enzyme complexed with target ligands was conducted in order to obtain detailed information about the structural limitations of its catalytic properties. In this study, the crystallization and preliminary X-ray analysis of complexes of wild-type DhaA with 2-propanol and with TCP and of complexes of the catalytically inactive variant DhaA13 with the dye coumarin and with TCP are described. The crystals of wild-type DhaA were plate-shaped and belonged to the triclinic space group P1, while the variant DhaA13 can form prism-shaped crystals belonging to the orthorhombic space group P2 1 2 1 2 1 as well as plate-shaped crystals belonging to the triclinic space group P1. Diffraction data for crystals of wild-type DhaA grown from crystallization solutions with different concentrations of 2-propanol were collected to 1.70 and 1.26 Å resolution, respectively. A prism-shaped crystal of DhaA13 complexed with TCP and a plate-shaped crystal of the same variant complexed with the dye coumarin diffracted X-rays to 1.60 and 1.33 Å resolution, respectively. A crystal of wild-type DhaA and a plate-shaped crystal of DhaA13, both complexed with TCP, diffracted to atomic resolutions of 1.04 and 0.97 Å, respectively

Distinct Phylogeographic Structures of Wild Radish (Raphanus sativus L. var. raphanistroides Makino in Japan.

Directory of Open Access Journals (Sweden)

Qingxiang Han

Full Text Available Coastal plants with simple linear distribution ranges along coastlines provide a suitable system for improving our understanding of patterns of intra-specific distributional history and genetic variation. Due to the combination of high seed longevity and high dispersibility of seeds via seawater, we hypothesized that wild radish would poorly represent phylogeographic structure at the local scale. On the other hand, we also hypothesized that wild radish populations might be geographically differentiated, as has been exhibited by their considerable phenotypic variations along the islands of Japan. We conducted nuclear DNA microsatellite loci and chloroplast DNA haplotype analyses for 486 samples and 144 samples, respectively, from 18 populations to investigate the phylogeographic structure of wild radish in Japan. Cluster analysis supported the existence of differential genetic structures between the Ryukyu Islands and mainland Japan populations. A significant strong pattern of isolation by distance and significant evidence of a recent bottleneck were detected. The chloroplast marker analysis resulted in the generation of eight haplotypes, of which two haplotypes (A and B were broadly distributed in most wild radish populations. High levels of variation in microsatellite loci were identified, whereas cpDNA displayed low levels of genetic diversity within populations. Our results indicate that the Kuroshio Current would have contributed to the sculpting of the phylogeographic structure by shaping genetic gaps between isolated populations. In addition, the Tokara Strait would have created a geographic barrier between the Ryukyu Islands and mainland Japan. Finally, extant habitat disturbances (coastal erosion, migration patterns (linear expansion, and geographic characteristics (small islands and sea currents have influenced the expansion and historical population dynamics of wild radish. Our study is the first to record the robust phylogeographic

Nitroaromatic detection and infrared communication from wild-type plants using plant nanobionics

Science.gov (United States)

Wong, Min Hao; Giraldo, Juan P.; Kwak, Seon-Yeong; Koman, Volodymyr B.; Sinclair, Rosalie; Lew, Tedrick Thomas Salim; Bisker, Gili; Liu, Pingwei; Strano, Michael S.

2017-02-01

Plant nanobionics aims to embed non-native functions to plants by interfacing them with specifically designed nanoparticles. Here, we demonstrate that living spinach plants (Spinacia oleracea) can be engineered to serve as self-powered pre-concentrators and autosamplers of analytes in ambient groundwater and as infrared communication platforms that can send information to a smartphone. The plants employ a pair of near-infrared fluorescent nanosensors--single-walled carbon nanotubes (SWCNTs) conjugated to the peptide Bombolitin II to recognize nitroaromatics via infrared fluorescent emission, and polyvinyl-alcohol functionalized SWCNTs that act as an invariant reference signal--embedded within the plant leaf mesophyll. As contaminant nitroaromatics are transported up the roots and stem into leaf tissues, they accumulate in the mesophyll, resulting in relative changes in emission intensity. The real-time monitoring of embedded SWCNT sensors also allows residence times in the roots, stems and leaves to be estimated, calculated to be 8.3 min (combined residence times of root and stem) and 1.9 min mm-1 leaf, respectively. These results demonstrate the ability of living, wild-type plants to function as chemical monitors of groundwater and communication devices to external electronics at standoff distances.

Maintenance erlotinib in advanced nonsmall cell lung cancer: cost-effectiveness in EGFR wild-type across Europe

Directory of Open Access Journals (Sweden)

Walleser S

2012-09-01

Full Text Available Silke Walleser,1 Joshua Ray,2 Helge Bischoff,3 Alain Vergnenègre,4 Hubertus Rosery,5 Christos Chouaid,6 David Heigener,7 Javier de Castro Carpeño,8 Marcello Tiseo,9 Stefan Walzer21Health Economic Consultancy, Renens, Switzerland; 2F Hoffmann-La Roche Pharmaceuticals AG, Basel, Switzerland; 3Thoracic Hospital of Heidelberg, Heidelberg, Germany; 4Limoges University Hospital, Limoges, France; 5Assessment-in-Medicine GmbH, Loerrach, Germany; 6Hospital Saint Antoine, Paris, France; 7Hospital Grosshansdorf, Grosshansdorf, Germany; 8University Hospital La Paz, Madrid, Spain; 9University Hospital of Parma, Parma, ItalyBackground: First-line maintenance erlotinib in patients with locally advanced or metastatic nonsmall cell lung cancer (NSCLC has demonstrated significant overall survival and progression-free survival benefits compared with best supportive care plus placebo, irrespective of epidermal growth factor receptor (EGFR status (SATURN trial. The cost-effectiveness of first-line maintenance erlotinib in the overall SATURN population has been assessed and published recently, but analyses according to EGFR mutation status have not been performed yet, which was the rationale for assessing the cost-effectiveness of first-line maintenance erlotinib specifically in EGFR wild-type metastatic NSCLC.Methods: The incremental cost per life-year gained of first-line maintenance erlotinib compared with best supportive care in patients with EGFR wild-type stable metastatic NSCLC was assessed for five European countries (the United Kingdom, Germany, France, Spain, and Italy with an area-under-the-curve model consisting of three health states (progression-free survival, progressive disease, death. Log-logistic survival functions were fitted to Phase III patient-level data (SATURN to model progression-free survival and overall survival. The first-line maintenance erlotinib therapy cost (modeled for time to treatment cessation, medication cost in later lines, and

Panitumumab and pegylated liposomal doxorubicin in platinum-resistant epithelial ovarian cancer with KRAS wild-type

DEFF Research Database (Denmark)

Steffensen, Karina Dahl; Waldstrøm, Marianne; Pallisgård, Niels

2013-01-01

OBJECTIVE: The increasing number of negative trials for ovarian cancer treatment has prompted an evaluation of new biologic agents, which in combination with chemotherapy may improve survival. The aim of this study was to investigate the response rate in platinum-resistant, KRAS wild-type ovarian...... cancer patients treated with pegylated liposomal doxorubicin (PLD) supplemented with panitumumab. PATIENTS AND METHODS: Major eligibility criteria were relapsed ovarian/fallopian/peritoneal cancer patients with platinum-resistant disease, measurable disease by GCIG CA125 criteria and KRAS wild-type...

Automatic Detection of Wild-type Mouse Cranial Sutures

DEFF Research Database (Denmark)

Ólafsdóttir, Hildur; Darvann, Tron Andre; Hermann, Nuno V.

, automatic detection of the cranial sutures becomes important. We have previously built a craniofacial, wild-type mouse atlas from a set of 10 Micro CT scans using a B-spline-based nonrigid registration method by Rueckert et al. Subsequently, all volumes were registered nonrigidly to the atlas. Using......, the observer traced the sutures on each of the mouse volumes as well. The observer outperforms the automatic approach by approximately 0.1 mm. All mice have similar errors while the suture error plots reveal that suture 1 and 2 are cumbersome, both for the observer and the automatic approach. These sutures can...

Liver steatosis study_PFAA treated Wild type and PPAR KO mouse data

Data.gov (United States)

U.S. Environmental Protection Agency — Data set 1 consists of the experimental data for the Wild Type and PPAR KO animal study and includes data used to prepare Figures 1-4 and Table 1 of the Das et al,...

Coexistence of Cluster Structure and Mean-field-type Structure in Medium-weight Nuclei

International Nuclear Information System (INIS)

Taniguchi, Yasutaka; Horiuchi, Hisashi; Kimura, Masaaki

2006-01-01

We have studied the coexistence of cluster structure and mean-field-type structure in 20Ne and 40Ca using Antisymmetrized Molecular Dynamics (AMD) + Generator Coordinate Method (GCM). By energy variation with new constraint for clustering, we calculate cluster structure wave function. Superposing cluster structure wave functions and mean-field-type structure wave function, we found that 8Be-12C, α-36Ar and 12C-28Si cluster structure are important components of K π = 0 3 + band of 20Ne, that of normal deformed band of 40Ca and that of super deformed band of 40Ca, respectively

Dose selenomethionine have radio-protective effect on cell lines with wild type p53?

International Nuclear Information System (INIS)

Tsuji, K.; Hagihira, T.; Ohnishi, K.; Ohnishi, T.; Matsumoto, H.

2003-01-01

Full text: Selenium compounds are known to have cancer preventive effects. It is reported recently that selenium in the form of selenomethionine (SeMet) can protect cells with wild type p53 from UV-induced cell killing by activating the DNA repair mechanism of p53 tumor suppressor protein via redox factor Ref1 by reducing p53 cysteine residue 275 and 277. In contrast, SeMet has no protective effect on UV-induced cell killing in p53-null cells. If SeMet also has protective effect in cells with wild type p53 on cell killing by photon irradiation, SeMet can be used as normal tissue radio-protector. We examined the effect of SeMet on cell killing by X-ray irradiation in several cell lines with different p53 status at exponentially growing phase. Cell lines used in this experiment were as follows: H1299/neo; human lung cancer cell line of p53 null type tranfected with control vector with no p53, H1299/wp53; wild type p53 transfected counterpart. A172/neo; human glioblastoma cell line with wild type p53, A172/mp53-248; mp53-248 (248-mutant, ARG >TRP) transfected counterpart. SAS/neo; human tongue cancer cell line with wild type p53, and SAS/mp53-248; mp53-248 transfected counterpart. Cells were subcultured at monolayer in D-MEM containing 10% FBS. Survivals of the cells were determined by colony forming ability. Ten-MV linac X-ray was used to irradiate the cells. Exponentially growing cells were incubated with 20μM of SeMet for 15 hours before irradiation. After 24 hours exposure of SeMet, cells were incubated up to two weeks in growth medium for colony formation. Twenty-four hours exposure of 20μM of SeMet had no cytotoxicity on these cell lines. SeMet had no modification effect on cell killing by photon irradiation in H1299/neo, H1299/wp53, SAS/neo, SAS/mp53-248, and A172/mp53-248. On the other hand, SeMet sensitized A172/neo in radiation cell killing. The effects of p53 on interaction of SeMet and photon irradiation differ according to cell lines

Spatial encoding in spinal sensorimotor circuits differs in different wild type mice strains

Directory of Open Access Journals (Sweden)

Schouenborg Jens

2008-05-01

Full Text Available Abstract Background Previous studies in the rat have shown that the spatial organisation of the receptive fields of nociceptive withdrawal reflex (NWR system are functionally adapted through experience dependent mechanisms, termed somatosensory imprinting, during postnatal development. Here we wanted to clarify 1 if mice exhibit a similar spatial encoding of sensory input to NWR as previously found in the rat and 2 if mice strains with a poor learning capacity in various behavioural tests, associated with deficient long term potention, also exhibit poor adaptation of NWR. The organisation of the NWR system in two adult wild type mouse strains with normal long term potentiation (LTP in hippocampus and two adult wild type mouse strains exhibiting deficiencies in corresponding LTP were used and compared to previous results in the rat. Receptive fields of reflexes in single hindlimb muscles were mapped with CO2 laser heat pulses. Results While the spatial organisation of the nociceptive receptive fields in mice with normal LTP were very similar to those in rats, the LTP impaired strains exhibited receptive fields of NWRs with aberrant sensitivity distributions. However, no difference was found in NWR thresholds or onset C-fibre latencies suggesting that the mechanisms determining general reflex sensitivity and somatosensory imprinting are different. Conclusion Our results thus confirm that sensory encoding in mice and rat NWR is similar, provided that mice strains with a good learning capability are studied and raise the possibility that LTP like mechanisms are involved in somatosensory imprinting.

ELECTROPHORETIC MOBILITIES OF ESCHERICHIA COLI 0157:H7 AND WILD-TYPE ESCHERICHIA COLI STRAINS

Science.gov (United States)

The electrophoretic mobility (EPM) of a number of human-virulent and "wild-type" Escherichia coli strains in phosphate buffered water was measured. The impact of pH, ionic strength, cation type (valence) and concentration, and bacterial strain on the EPM was investigated. Resul...

Crystal Structure of the Human, FIC-Domain Containing Protein HYPE and Implications for Its Functions

Science.gov (United States)

Bunney, Tom D.; Cole, Ambrose R.; Broncel, Malgorzata; Esposito, Diego; Tate, Edward W.; Katan, Matilda

2014-01-01

Summary Protein AMPylation, the transfer of AMP from ATP to protein targets, has been recognized as a new mechanism of host-cell disruption by some bacterial effectors that typically contain a FIC-domain. Eukaryotic genomes also encode one FIC-domain protein, HYPE, which has remained poorly characterized. Here we describe the structure of human HYPE, solved by X-ray crystallography, representing the first structure of a eukaryotic FIC-domain protein. We demonstrate that HYPE forms stable dimers with structurally and functionally integrated FIC-domains and with TPR-motifs exposed for protein-protein interactions. As HYPE also uniquely possesses a transmembrane helix, dimerization is likely to affect its positioning and function in the membrane vicinity. The low rate of autoAMPylation of the wild-type HYPE could be due to autoinhibition, consistent with the mechanism proposed for a number of putative FIC AMPylators. Our findings also provide a basis to further consider possible alternative cofactors of HYPE and distinct modes of target-recognition. PMID:25435325

Anthelmintic effect of Psidium guajava and Tagetes erecta on wild-type and Levamisole-resistant Caenorhabditis elegans strains.

Science.gov (United States)

Piña-Vázquez, Denia M; Mayoral-Peña, Zyanya; Gómez-Sánchez, Maricela; Salazar-Olivo, Luis A; Arellano-Carbajal, Fausto

2017-04-18

Psidium guajava and Tagetes erecta have been used traditionally to treat gastrointestinal parasites, but their active metabolites and mechanisms of action remain largely unknown. To evaluate the anthelmintic potential of Psidium guajava and Tagetes erecta extracts on Levamisole-sensitive and Levamisole-resistant strains of the model nematode Caenorhabditis elegans. Aqueous extracts of Psidium guajava (PGE) and Tagetes erecta (TEE) were assayed on locomotion and egg-laying behaviors of the wild-type (N2) and Levamisole-resistant (CB193) strains of Caenorhabditis elegans. Both extracts paralyzed wild-type and Levamisole-resistant nematodes in a dose-dependent manner. In wild-type worms, TEE 25mg/mL induced a 75% paralysis after 8h of treatment and PGE 25mg/mL induced a 100% paralysis after 4h of treatment. PGE exerted a similar paralyzing effect on N2 wild-type and CB193 Levamisole-resistant worms, while TEE only partially paralyzed CB193 worms. TEE 25mg/mL decreased N2 egg-laying by 65% with respect to the untreated control, while PGE did it by 40%. Psidium guajava leaves and Tagetes erecta flower-heads possess hydrosoluble compounds that block the motility of Caenorhabditis elegans by a mechanism different to that of the anthelmintic drug Levamisole. Effects are also observable on oviposition, which was diminished in the wild-type worms. The strong anthelmintic effects in crude extracts of these plants warrants future work to identify their active compounds and to elucidate their molecular mechanisms of action. Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.

Capacity for cooperative binding of thyroid hormone (T3) receptor dimers defines wild type T3 response elements.

Science.gov (United States)

Brent, G A; Williams, G R; Harney, J W; Forman, B M; Samuels, H H; Moore, D D; Larsen, P R

1992-04-01

Thyroid hormone response elements (T3REs) have been identified in a variety of promoters including those directing expression of rat GH (rGH), alpha-myosin heavy chain (rMHC), and malic enzyme (rME). A detailed biochemical and genetic analysis of the rGH element has shown that it consists of three hexamers related to the consensus [(A/G)GGT(C/A)A]. We have extended this analysis to the rMHC and rME elements. Binding of highly purified thyroid hormone receptor (T3R) to T3REs was determined using the gel shift assay, and thyroid hormone (T3) induction was measured in transient tranfections. We show that the wild type version of each of the three elements binds T3R dimers cooperatively. Mutational analysis of the rMHC and rME elements identified domains important for binding T3R dimers and allowed a direct determination of the relationship between T3R binding and function. In each element two hexamers are required for dimer binding, and mutations that interfere with dimer formation significantly reduce T3 induction. Similar to the rGH element, the rMHC T3RE contains three hexameric domains arranged as a direct repeat followed by an inverted copy, although the third domain is weaker than in rGH. All three are required for full function and T3R binding. The rME T3RE is a two-hexamer direct repeat T3RE, which also binds T3R monomer and dimer. Across a series of mutant elements, there was a strong correlation between dimer binding in vitro and function in vivo for rMHC (r = 0.99, P less than 0.01) and rME (r = 0.67, P less than 0.05) T3REs. Our results demonstrate a similar pattern of T3R dimer binding to a diverse array of hexameric sequences and arrangements in three wild type T3REs. Addition of nuclear protein enhanced T3R binding but did not alter the specificity of binding to wild type or mutant elements. Binding of purified T3R to T3REs was highly correlated with function, both with and without the addition of nuclear protein. T3R dimer formation is the common

Comparison of the nucleotide sequence of wild-type hepatitis - A virus and its attenuated candidate vaccine derivative

International Nuclear Information System (INIS)

Cohen, J.I.; Rosenblum, B.; Ticehurst, J.R.; Daemer, R.; Feinstone, S.; Purcell, R.H.

1987-01-01

Development of attenuated mutants for use as vaccines is in progress for other viruses, including influenza, rotavirus, varicella-zoster, cytomegalovirus, and hepatitis-A virus (HAV). Attenuated viruses may be derived from naturally occurring mutants that infect human or nonhuman hosts. Alternatively, attenuated mutants may be generated by passage of wild-type virus in cell culture. Production of attenuated viruses in cell culture is a laborious and empiric process. Despite previous empiric successes, understanding the molecular basis for attenuation of vaccine viruses could facilitate future development and use of live-virus vaccines. Comparison of the complete nucleotide sequences of wild-type (virulent) and vaccine (attenuated) viruses has been reported for polioviruses and yellow fever virus. Here, the authors compare the nucleotide sequence of wild-type HAV HM-175 with that of a candidate vaccine derivative

Comparison of the crystal structure and function to wild-type and His25Ala mutant human heme oxygenase-1.

Science.gov (United States)

Zhou, Wen-Pu; Zhong, Wen-Wei; Zhang, Xue-Hong; Ding, Jian-Ping; Zhang, Zi-Li; Xia, Zhen-Wei

2009-03-01

Human heme oxygenase-1 (hHO-1) is a rate-limiting enzyme in heme metabolism. It regulates serum bilirubin level. Site-directed mutagenesis studies indicate that the proximal residue histidine 25 (His25) plays a key role in hHO-1 activity. A highly purified hHO-1 His25Ala mutant was generated and crystallized with a new expression system. The crystal structure of the mutant was determined by X-ray diffraction technology and molecular replacement at the resolution of 2.8 A, and the model of hHO-1 His25Ala mutant was refined. The final crystallographic and free R factors were 0.245 and 0.283, respectively. The standard bond length deviation was 0.007 A, and the standard bond angle deviation was 1.3 degrees . The mutation of His25 to Ala led to an empty pocket underneath the ferric ion in the heme, leading to loss of binding iron ligand. Although this did not cause an overall structural change, the enzymatic activity of the mutant hHO-1 was reduced by 90%. By supplementing imidazole, the HO-1 activity was restored approximately 90% to its normal level. These data suggest that Ala25 remains unchanged in the structure compared to His25, but the important catalytic function of hHO-1 is lost. Thus, it appears that His25 is a crucial residue for proper hHO-1 catalysis.

Structural Features Reminiscent of ATP-Driven Protein Translocases Are Essential for the Function of a Type III Secretion-Associated ATPase.

Science.gov (United States)

Kato, Junya; Lefebre, Matthew; Galán, Jorge E

2015-09-01

Many bacterial pathogens and symbionts utilize type III secretion systems to interact with their hosts. These machines have evolved to deliver bacterial effector proteins into eukaryotic target cells to modulate a variety of cellular functions. One of the most conserved components of these systems is an ATPase, which plays an essential role in the recognition and unfolding of proteins destined for secretion by the type III pathway. Here we show that structural features reminiscent of other ATP-driven protein translocases are essential for the function of InvC, the ATPase associated with a Salmonella enterica serovar Typhimurium type III secretion system. Mutational and functional analyses showed that a two-helix-finger motif and a conserved loop located at the entrance of and within the predicted pore formed by the hexameric ATPase are essential for InvC function. These findings provide mechanistic insight into the function of this highly conserved component of type III secretion machines. Type III secretion machines are essential for the virulence or symbiotic relationships of many bacteria. These machines have evolved to deliver bacterial effector proteins into host cells to modulate cellular functions, thus facilitating bacterial colonization and replication. An essential component of these machines is a highly conserved ATPase, which is necessary for the recognition and secretion of proteins destined to be delivered by the type III secretion pathway. Using modeling and structure and function analyses, we have identified structural features of one of these ATPases from Salmonella enterica serovar Typhimurium that help to explain important aspects of its function. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

The fusion protein of wild-type canine distemper virus is a major determinant of persistent infection

International Nuclear Information System (INIS)

Plattet, Philippe; Rivals, Jean-Paul; Zuber, BenoIt; Brunner, Jean-Marc; Zurbriggen, Andreas; Wittek, Riccardo

2005-01-01

The wild-type A75/17 canine distemper virus (CDV) strain induces a persistent infection in the central nervous system but infects cell lines very inefficiently. In contrast, the genetically more distant Onderstepoort CDV vaccine strain (OP-CDV) induces extensive syncytia formation. Here, we investigated the roles of wild-type fusion (F WT ) and attachment (H WT ) proteins in Vero cells expressing, or not, the canine SLAM receptor by transfection experiments and by studying recombinants viruses expressing different combinations of wild-type and OP-CDV glycoproteins. We show that low fusogenicity is not due to a defect of the envelope proteins to reach the cell surface and that H WT determines persistent infection in a receptor-dependent manner, emphasizing the role of SLAM as a potent enhancer of fusogenicity. However, importantly, F WT reduced cell-to-cell fusion independently of the cell surface receptor, thus demonstrating that the fusion protein of the neurovirulent A75/17-CDV strain plays a key role in determining persistent infection

Crystallization and preliminary X-ray diffraction analysis of the wild-type haloalkane dehalogenase DhaA and its variant DhaA13 complexed with different ligands.

Science.gov (United States)

Stsiapanava, Alena; Chaloupkova, Radka; Fortova, Andrea; Brynda, Jiri; Weiss, Manfred S; Damborsky, Jiri; Smatanova, Ivana Kuta

2011-02-01

Haloalkane dehalogenases make up an important class of hydrolytic enzymes which catalyse the cleavage of carbon-halogen bonds in halogenated aliphatic compounds. There is growing interest in these enzymes owing to their potential use in environmental and industrial applications. The haloalkane dehalogenase DhaA from Rhodococcus rhodochrous NCIMB 13064 can slowly detoxify the industrial pollutant 1,2,3-trichloropropane (TCP). Structural analysis of this enzyme complexed with target ligands was conducted in order to obtain detailed information about the structural limitations of its catalytic properties. In this study, the crystallization and preliminary X-ray analysis of complexes of wild-type DhaA with 2-propanol and with TCP and of complexes of the catalytically inactive variant DhaA13 with the dye coumarin and with TCP are described. The crystals of wild-type DhaA were plate-shaped and belonged to the triclinic space group P1, while the variant DhaA13 can form prism-shaped crystals belonging to the orthorhombic space group P2(1)2(1)2(1) as well as plate-shaped crystals belonging to the triclinic space group P1. Diffraction data for crystals of wild-type DhaA grown from crystallization solutions with different concentrations of 2-propanol were collected to 1.70 and 1.26 Å resolution, respectively. A prism-shaped crystal of DhaA13 complexed with TCP and a plate-shaped crystal of the same variant complexed with the dye coumarin diffracted X-rays to 1.60 and 1.33 Å resolution, respectively. A crystal of wild-type DhaA and a plate-shaped crystal of DhaA13, both complexed with TCP, diffracted to atomic resolutions of 1.04 and 0.97 Å, respectively.

Expression of wild-type and mutant medium-chain acyl-CoA dehydrogenase (MCAD) cDNA in eucaryotic cells

DEFF Research Database (Denmark)

Jensen, T G; Andresen, B S; Bross, P

1992-01-01

An effective EBV-based expression system for eucaryotic cells has been developed and used for the study of the mitochondrial enzyme medium-chain acyl-CoA dehydrogenase (MCAD). 1325 bp of PCR-generated MCAD cDNA, containing the entire coding region, was placed between the SV40 early promoter...... and polyadenylation signals in the EBV-based vector. Both wild-type MCAD cDNA and cDNA containing the prevalent disease-causing mutation A to G at position 985 of the MCAD cDNA were tested. In transfected COS-7 cells, the steady state amount of mutant MCAD protein was consistently lower than the amount of wild......-type human enzyme. The enzyme activity in extracts from cells harbouring the wild-type MCAD cDNA was dramatically higher than in the controls (harbouring the vector without the MCAD gene) while only a slightly higher activity was measured with the mutant MCAD. The mutant MCAD present behaves like wild...

The AtNFXL1 gene functions as a signaling component of the type A trichothecene-dependent response

Science.gov (United States)

Asano, Tomoya; Yasuda, Michiko; Nakashita, Hideo; Kimura, Makoto; Yamaguchi1, Kazuo

2008-01-01

Phytopathogenic Fusarium species produce the trichothecene family of phytotoxins, which function as a virulence factor during infection of plants. Trichothecenes are classifiable into four major groups by their chemical structures. Recently, the AtNFXL1 gene was reported as a type A trichothecene T-2 toxin-inducible gene. The AtNFXL1 gene encodes a putative transcription factor with similarity to the human transcription repressor NF-X1. The atnfxl1 mutant exhibited hypersensitivity phenotype to T-2 toxin but not to type B deoxynivalenol (DON) in comparison with wild type when Arabidopsis thaliana grew on agar medium containing trichothecenes. The absence or presence of a carbonyl group at the C8 position distinguishes type A and type B. Growth defect by another type A trichothecene diacetoxyscirpenol (DAS), was weakly enhanced in the atnfxl1 mutant. Diacetoxyscirpenol is distinguishable from T-2 toxin only by the absence of an isovaleryl group at the C8 position. Correspondingly, the AtNFXL1 promoter activity was apparently induced in T-2 toxin-treated and DAS-treated plants. In contrast, DON failed to induce the AtNFXL1 promoter activity. Consequently, the AtNFXL1 gene functions as a signaling component of the type A trichothecene-dependent response in Arabidopsis. In addition, the C8 position of trichothecenes might be closely related to the function of AtNFXL1 gene. PMID:19704430

The correlation between the use of personal protective equipment and level wild-type p53 of dental technicians in Surabaya

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Puspa Dila Rohmaniar

2017-03-01

Full Text Available Background: Exposure of metals among dental technicians that come from the working environment can lead to the formation reactive oxygen species (ROS. ROS can cause mutations in the p53 gene (p53. The mutation is transversion mutation GuanineThymine. p53 mutations can lead to low expression of the wild-type p53 protein (p53. Wild-type p53 involved in many biological processes such as regulation of genes involved in cell cycle, cell growth after DNA damage, and apoptosis. However, exposure to metals among dental technicians can be prevented through the use of personal protective equipment (PPE during work. Purpose: The purpose of this study was to analyze the correlation between the use of personal protective equipment to wild-type p53 protein levels among dental technicians in Surabaya. Method: This study was observational analytic with cross sectional approach. 40 samples were taken by random sampling. Data were retrieved through interviews and observations. Wild-type p53 was analyzed from saliva with indirect ELISA method. Analysis of data used Kolmogorov Smirnov normality test and a Pearson correlation test. Value significance was p<0.05 (95% confidence level. Result: There was a significant association between the use of personal protective equipment with wild-type p53 levels with p=0.002 Conclusion: The use PPE properly is positively correlated with the wild-type p53 protein levels of dental technicians in Surabaya.

Scanning Ultrasound (SUS Causes No Changes to Neuronal Excitability and Prevents Age-Related Reductions in Hippocampal CA1 Dendritic Structure in Wild-Type Mice.

Directory of Open Access Journals (Sweden)

Robert John Hatch

Full Text Available Scanning ultrasound (SUS is a noninvasive approach that has recently been shown to ameliorate histopathological changes and restore memory functions in an Alzheimer's disease mouse model. Although no overt neuronal damage was reported, the short- and long-term effects of SUS on neuronal excitability and dendritic tree morphology had not been investigated. To address this, we performed patch-clamp recordings from hippocampal CA1 pyramidal neurons in wild-type mice 2 and 24 hours after a single SUS treatment, and one week and 3 months after six weekly SUS treatments, including sham treatments as controls. In both treatment regimes, no changes in CA1 neuronal excitability were observed in SUS-treated neurons when compared to sham-treated neurons at any time-point. For the multiple treatment groups, we also determined the dendritic morphology and spine densities of the neurons from which we had recorded. The apical trees of sham-treated neurons were reduced at the 3 month time-point when compared to one week; however, surprisingly, no longitudinal change was detected in the apical dendritic trees of SUS-treated neurons. In contrast, the length and complexity of the basal dendritic trees were not affected by SUS treatment at either time-point. The apical dendritic spine densities were reduced, independent of the treatment group, at 3 months compared to one week. Collectively, these data suggest that ultrasound can be employed to prevent an age-associated loss of dendritic structure without impairing neuronal excitability.

Effect of uremia on HDL composition, vascular inflammation, and atherosclerosis in wild-type mice

DEFF Research Database (Denmark)

Bang, Christian A; Bro, Susanne; Bartels, Emil D

2007-01-01

Wild-type mice normally do not develop atherosclerosis, unless fed cholic acid. Uremia is proinflammatory and increases atherosclerosis 6- to 10-fold in apolipoprotein E-deficient mice. This study examined the effect of uremia on lipoproteins, vascular inflammation, and atherosclerosis in wild...... in cholic acid-fed sham mice. The results suggest that moderate uremia neither induces aortic inflammation nor atherosclerosis in C57BL/6J mice despite increased LDL/HDL cholesterol ratio and altered HDL composition....

SDHA loss of function mutations in a subset of young adult wild-type gastrointestinal stromal tumors

International Nuclear Information System (INIS)

Italiano, Antoine; Chen, Chun-Liang; Sung, Yun-Shao; Singer, Samuel; DeMatteo, Ronald P; LaQuaglia, Michael P; Besmer, Peter; Socci, Nicholas; Antonescu, Cristina R

2012-01-01

A subset of KIT/PDGFRA wild-type gastrointestinal stromal tumors (WT GIST) have been associated with alteration of the succinate dehydrogenase (SDH) complex II function. A recent report identified four non-syndromic, KIT/PDGFRA WT GIST harboring compound heterozygous or homozygous mutations in SDHA encoding the main subunit of the SDH complex II. Next generation sequencing was applied on five pediatric and one young adult WT GIST, by whole exome capture and SOLiD 3-plus system sequencing. The putative mutations were first confirmed by Sanger sequencing and then screened on a larger panel of 11 pediatric and young adult WT GIST, including 5 in the context of Carney triad. A germline p.Arg31X nonsense SDHA mutation was identified in one of the six cases tested by SOLiD platform. An additional p.D38V missense mutation in SDHA exon 2 was identified by Sanger sequencing in the extended KIT/PDGFRA WT GIST patients cohort. Western blotting showed loss of SDHA expression in the two cases harboring SDHA mutations, while expression being retained in the other WT GIST tumors. Results were further confirmed by immunohistochemistry for both SDHA and SDHB, which showed a concurrent loss of expression of both proteins in SDHA-mutant lesions, while the remaining WT tumors showed only loss of SDHB expression. Germline and/or somatic aberrations of SDHA occur in a small subset of KIT/PDGFRA WT GISTs, outside the Carney’s triad and are associated with loss of both SDHA and SDHB protein expression. Mutations of the SDH complex II are more particularly associated with KIT/PDGFRA WT GIST occurring in young adults. Although pediatric GIST consistently display alterations of SDHB protein expression, further molecular studies are needed to identify the crucial genes involved in their tumorigenesis

Atomic force microscopy study of the structure function relationships of the biofilm-forming bacterium Streptococcus mutans

Science.gov (United States)

Cross, Sarah E.; Kreth, Jens; Zhu, Lin; Qi, Fengxia; Pelling, Andrew E.; Shi, Wenyuan; Gimzewski, James K.

2006-02-01

Atomic force microscopy (AFM) has garnered much interest in recent years for its ability to probe the structure, function and cellular nanomechanics inherent to specific biological cells. In particular, we have used AFM to probe the important structure-function relationships of the bacterium Streptococcus mutans. S. mutans is the primary aetiological agent in human dental caries (tooth decay), and is of medical importance due to the virulence properties of these cells in biofilm initiation and formation, leading to increased tolerance to antibiotics. We have used AFM to characterize the unique surface structures of distinct mutants of S. mutans. These mutations are located in specific genes that encode surface proteins, thus using AFM we have resolved characteristic surface features for mutant strains compared to the wild type. Ultimately, our characterization of surface morphology has shown distinct differences in the local properties displayed by various S. mutans strains on the nanoscale, which is imperative for understanding the collective properties of these cells in biofilm formation.

Nitroaromatic detection and infrared communication from wild-type plants using plant nanobionics.

Science.gov (United States)

Wong, Min Hao; Giraldo, Juan P; Kwak, Seon-Yeong; Koman, Volodymyr B; Sinclair, Rosalie; Lew, Tedrick Thomas Salim; Bisker, Gili; Liu, Pingwei; Strano, Michael S

2017-02-01

Plant nanobionics aims to embed non-native functions to plants by interfacing them with specifically designed nanoparticles. Here, we demonstrate that living spinach plants (Spinacia oleracea) can be engineered to serve as self-powered pre-concentrators and autosamplers of analytes in ambient groundwater and as infrared communication platforms that can send information to a smartphone. The plants employ a pair of near-infrared fluorescent nanosensors-single-walled carbon nanotubes (SWCNTs) conjugated to the peptide Bombolitin II to recognize nitroaromatics via infrared fluorescent emission, and polyvinyl-alcohol functionalized SWCNTs that act as an invariant reference signal-embedded within the plant leaf mesophyll. As contaminant nitroaromatics are transported up the roots and stem into leaf tissues, they accumulate in the mesophyll, resulting in relative changes in emission intensity. The real-time monitoring of embedded SWCNT sensors also allows residence times in the roots, stems and leaves to be estimated, calculated to be 8.3 min (combined residence times of root and stem) and 1.9 min mm -1 leaf, respectively. These results demonstrate the ability of living, wild-type plants to function as chemical monitors of groundwater and communication devices to external electronics at standoff distances.

Wild-type minimal inhibitory concentration distributions in bacteria of animal origin in Argentina

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Florencia L Pantozzi

Full Text Available The aim of this study was to determine the antimicrobial resistance profiles of indicator bacteria isolated from domestic animal feces. Minimal inhibitory concentration (MIC was determined by agar dilution. Interpretative criteria on the basis of wild-type MIC distributions and epidemiological cutoff values (ECOFF or ECV were used according to the 'European Committee on Antimicrobial Susceptibility Testing' (EUCAST data. Results from 237 isolates of Escherichia coli showed reduced susceptibility for ampicillin, streptomycin and tetracycline, the antimicrobials commonly used in intensive breeding of pigs and hens. Regarding all the species of the genus Enterococcus spp., there are only ECOFF or ECV for vancomycin. Of the 173 Enterococcus spp. isolated, only one showed reduced susceptibility to vancomycin and was classified as 'non-wild-type' (NWT population. This is the first report in Argentina showing data of epidemiological cutoff values in animal bacteria.

A cerebellar learning model of vestibulo-ocular reflex adaptation in wild-type and mutant mice.

Science.gov (United States)

Clopath, Claudia; Badura, Aleksandra; De Zeeuw, Chris I; Brunel, Nicolas

2014-05-21

Mechanisms of cerebellar motor learning are still poorly understood. The standard Marr-Albus-Ito theory posits that learning involves plasticity at the parallel fiber to Purkinje cell synapses under control of the climbing fiber input, which provides an error signal as in classical supervised learning paradigms. However, a growing body of evidence challenges this theory, in that additional sites of plasticity appear to contribute to motor adaptation. Here, we consider phase-reversal training of the vestibulo-ocular reflex (VOR), a simple form of motor learning for which a large body of experimental data is available in wild-type and mutant mice, in which the excitability of granule cells or inhibition of Purkinje cells was affected in a cell-specific fashion. We present novel electrophysiological recordings of Purkinje cell activity measured in naive wild-type mice subjected to this VOR adaptation task. We then introduce a minimal model that consists of learning at the parallel fibers to Purkinje cells with the help of the climbing fibers. Although the minimal model reproduces the behavior of the wild-type animals and is analytically tractable, it fails at reproducing the behavior of mutant mice and the electrophysiology data. Therefore, we build a detailed model involving plasticity at the parallel fibers to Purkinje cells' synapse guided by climbing fibers, feedforward inhibition of Purkinje cells, and plasticity at the mossy fiber to vestibular nuclei neuron synapse. The detailed model reproduces both the behavioral and electrophysiological data of both the wild-type and mutant mice and allows for experimentally testable predictions. Copyright © 2014 the authors 0270-6474/14/347203-13$15.00/0.

Engineering of red cells of Arabidopsis thaliana and comparative genome-wide gene expression analysis of red cells versus wild-type cells.

Science.gov (United States)

Shi, Ming-Zhu; Xie, De-Yu

2011-04-01

We report metabolic engineering of Arabidopsis red cells and genome-wide gene expression analysis associated with anthocyanin biosynthesis and other metabolic pathways between red cells and wild-type (WT) cells. Red cells of A. thaliana were engineered for the first time from the leaves of production of anthocyanin pigment 1-Dominant (pap1-D). These red cells produced seven anthocyanin molecules including a new one that was characterized by LC-MS analysis. Wild-type cells established as a control did not produce anthocyanins. A genome-wide microarray analysis revealed that nearly 66 and 65% of genes in the genome were expressed in the red cells and wild-type cells, respectively. In comparison with the WT cells, 3.2% of expressed genes in the red cells were differentially expressed. The expression levels of 14 genes involved in the biosynthetic pathway of anthocyanin were significantly higher in the red cells than in the WT cells. Microarray and RT-PCR analyses demonstrated that the TTG1-GL3/TT8-PAP1 complex regulated the biosynthesis of anthocyanins. Furthermore, most of the genes with significant differential expression levels in the red cells versus the WT cells were characterized with diverse biochemical functions, many of which were mapped to different metabolic pathways (e.g., ribosomal protein biosynthesis, photosynthesis, glycolysis, glyoxylate metabolism, and plant secondary metabolisms) or organelles (e.g., chloroplast). We suggest that the difference in gene expression profiles between the two cell lines likely results from cell types, the overexpression of PAP1, and the high metabolic flux toward anthocyanins.

Structural and functional analysis of the ASM p.Ala359Asp mutant that causes acid sphingomyelinase deficiency.

Science.gov (United States)

Acuña, Mariana; Castro-Fernández, Víctor; Latorre, Mauricio; Castro, Juan; Schuchman, Edward H; Guixé, Victoria; González, Mauricio; Zanlungo, Silvana

2016-10-21

Niemann-Pick disease (NPD) type A and B are recessive hereditary disorders caused by deficiency in acid sphingomyelinase (ASM). The p.Ala359Asp mutation has been described in several patients but its functional and structural effects in the protein are unknown. In order to characterize this mutation, we modeled the three-dimensional ASM structure using the recent available crystal of the mammalian ASM as a template. We found that the p.Ala359Asp mutation is localized in the hydrophobic core and far from the sphingomyelin binding site. However, energy function calculations using statistical potentials indicate that the mutation causes a decrease in ASM stability. Therefore, we investigated the functional effect of the p.Ala359Asp mutation in ASM expression, secretion, localization and activity in human fibroblasts. We found a 3.8% residual ASM activity compared to the wild-type enzyme, without changes in the other parameters evaluated. These results support the hypothesis that the p.Ala359Asp mutation causes structural alterations in the hydrophobic environment where ASM is located, decreasing its enzymatic activity. A similar effect was observed in other previously described NPDB mutations located outside the active site of the enzyme. This work shows the first full size ASM mutant model describe at date, providing a complete analysis of the structural and functional effects of the p.Ala359Asp mutation over the stability and activity of the enzyme. Copyright © 2016 Elsevier Inc. All rights reserved.

Gene therapy restores auditory and vestibular function in a mouse model of Usher syndrome type 1c.

Science.gov (United States)

Pan, Bifeng; Askew, Charles; Galvin, Alice; Heman-Ackah, Selena; Asai, Yukako; Indzhykulian, Artur A; Jodelka, Francine M; Hastings, Michelle L; Lentz, Jennifer J; Vandenberghe, Luk H; Holt, Jeffrey R; Géléoc, Gwenaëlle S

2017-03-01

Because there are currently no biological treatments for hearing loss, we sought to advance gene therapy approaches to treat genetic deafness. We focused on Usher syndrome, a devastating genetic disorder that causes blindness, balance disorders and profound deafness, and studied a knock-in mouse model, Ush1c c.216G>A, for Usher syndrome type IC (USH1C). As restoration of complex auditory and balance function is likely to require gene delivery systems that target auditory and vestibular sensory cells with high efficiency, we delivered wild-type Ush1c into the inner ear of Ush1c c.216G>A mice using a synthetic adeno-associated viral vector, Anc80L65, shown to transduce 80-90% of sensory hair cells. We demonstrate recovery of gene and protein expression, restoration of sensory cell function, rescue of complex auditory function and recovery of hearing and balance behavior to near wild-type levels. The data represent unprecedented recovery of inner ear function and suggest that biological therapies to treat deafness may be suitable for translation to humans with genetic inner ear disorders.

Iron and zinc complexation in wild-type and ferritin-expressing wheat grain: implications for mineral transport into developing grain

DEFF Research Database (Denmark)

Neal, Andrew L; Geraki, Kalotina; Borg, Søren

2013-01-01

of modified complexation of both metals in transgenic grain overexpressing wheat ferritin. For zinc, there is a consistent doubling of the number of complexing phosphorus atoms. Although there is some EXAFS evidence for iron phytate in ferritin-expressing grain, there is also evidence of a structure lacking......We have used synchrotron-based X-ray fluorescence and absorption techniques to establish both metal distribution and complexation in mature wheat grains. In planta, extended X-ray absorption fine structure (EXAFS) spectroscopy reveals iron phytate and zinc phytate structures in aleurone cells...... of ferritin-expressing grains is quite different from that in wild-type grain. This may explain why the raised levels of minerals transported to the developing grain accumulate within the crease region of the transgenic grain....

Structural characterization and mutational assessment of podocin - a novel drug target to nephrotic syndrome - an in silico approach.

Science.gov (United States)

Tabassum, Asra; Rajeshwari, Tadigadapa; Soni, Nidhi; Raju, D S B; Yadav, Mukesh; Nayarisseri, Anuraj; Jahan, Parveen

2014-03-01

Non-synonymous single nucleotide changes (nSNC) are coding variants that introduce amino acid changes in their corresponding proteins. They can affect protein function; they are believed to have the largest impact on human health compared with SNCs in other regions of the genome. Such a sequence alteration directly affects their structural stability through conformational changes. Presence of these conformational changes near catalytic site or active site may alter protein function and as a consequence receptor-ligand complex interactions. The present investigation includes assessment of human podocin mutations (G92C, P118L, R138Q, and D160G) on its structure. Podocin is an important glomerular integral membrane protein thought to play a key role in steroid resistant nephrotic syndrome. Podocin has a hairpin like structure with 383 amino acids, it is an integral protein homologous to stomatin, and acts as a molecular link in a stretch-sensitive system. We modeled 3D structure of podocin by means of Modeller and validated via PROCHECK to get a Ramachandran plot (88.5% in most favored region), main chain, side chain, bad contacts, gauche and pooled standard deviation. Further, a protein engineering tool Triton was used to induce mutagenesis corresponding to four variants G92C, P118L, R138Q and D160G in the wild type. Perusal of energies of wild and mutated type of podocin structures confirmed that mutated structures were thermodynamically more stable than wild type and therefore biological events favored synthesis of mutated forms of podocin than wild type. As a conclusive part, two mutations G92C (-8179.272 kJ/mol) and P118L (-8136.685 kJ/mol) are more stable and probable to take place in podocin structure over wild podocin structure (-8105.622 kJ/mol). Though there is lesser difference in mutated and wild type (approximately, 74 and 35 kJ/mol), it may play a crucial role in deciding why mutations are favored and occur at the genetic level.

Structural and functional cardiac changes in myotonic dystrophy type 1: a cardiovascular magnetic resonance study

Directory of Open Access Journals (Sweden)

Hermans Mieke CE

2012-07-01

Full Text Available Abstract Background Myotonic dystrophy type 1 (MD1 is a neuromuscular disorder with potential involvement of the heart and increased risk of sudden death. Considering the importance of cardiomyopathy as a predictor of prognosis, we aimed to systematically evaluate and describe structural and functional cardiac alterations in patients with MD1. Methods Eighty MD1 patients underwent physical examination, electrocardiography (ECG, echocardiography and cardiovascular magnetic resonance (CMR. Blood samples were taken for determination of NT-proBNP plasma levels and CTG repeat length. Results Functional and structural abnormalities were detected in 35 patients (44%. Left ventricular systolic dysfunction was found in 20 cases, left ventricular dilatation in 7 patients, and left ventricular hypertrophy in 6 patients. Myocardial fibrosis was seen in 10 patients (12.5%. In general, patients had low left ventricular mass indexes. Right ventricular involvement was uncommon and only seen together with left ventricular abnormalities. Functional or structural cardiac involvement was associated with age (p = 0.04, male gender (p Conclusions CMR can be useful to detect early structural and functional myocardial abnormalities in patients with MD1. Myocardial involvement is strongly associated with conduction abnormalities, but a normal ECG does not exclude myocardial alterations. These findings lend support to the hypothesis that MD1 patients have a complex cardiac phenotype, including both myocardial and conduction system alteration.

Interleukin-1 receptor type I gene-deficient mice are less susceptible to Staphylococcus epidermidis biomaterial-associated infection than are wild-type mice

NARCIS (Netherlands)

Boelens, J. J.; van der Poll, T.; Zaat, S. A.; Murk, J. L.; Weening, J. J.; Dankert, J.

2000-01-01

Elevated concentrations of interleukin-1 (IL-1) were found in tissue surrounding biomaterials infected with Staphylococcus epidermidis. To determine the role of IL-1 in biomaterial-associated infection (BAI), IL-1 receptor type I-deficient (IL-1R(-/-)) and wild-type mice received subcutaneous

Effects of phorbol ester on mitogen-activated protein kinase kinase activity in wild-type and phorbol ester-resistant EL4 thymoma cells.

Science.gov (United States)

Gause, K C; Homma, M K; Licciardi, K A; Seger, R; Ahn, N G; Peterson, M J; Krebs, E G; Meier, K E

1993-08-05

Phorbol ester-sensitive and -resistant EL4 thymoma cell lines differ in their ability to activate mitogen-activated protein kinase (MAPK) in response to phorbol ester. Treatment of wild-type EL4 cells with phorbol ester results in the rapid activations of MAPK and pp90rsk kinase, a substrate for MAPK, while neither kinase is activated in response to phorbol ester in variant EL4 cells. This study examines the activation of MAPK kinase (MAPKK), an activator of MAPK, in wild-type and variant EL4 cells. Phosphorylation of a 40-kDa substrate, identified as MAPK, was observed following in vitro phosphorylation reactions using cytosolic extracts or Mono Q column fractions prepared from phorbol ester-treated wild-type EL4 cells. MAPKK activity coeluted with a portion of the inactive MAPK upon Mono Q anion-exchange chromatography, permitting detection of the MAPKK activity in fractions containing both kinases. This MAPKK activity was present in phorbol ester-treated wild-type cells, but not in phorbol ester-treated variant cells or in untreated wild-type or variant cells. The MAPKK from wild-type cells was able to activate MAPK prepared from either wild-type or variant cells. MAPKK activity could be stimulated in both wildtype and variant EL4 cells in response to treatment of cells with okadaic acid. These results indicate that the failure of variant EL4 cells to activate MAP kinase in response to phorbol ester is due to a failure to activate MAPKK. Therefore, the step that confers phorbol ester resistance to variant EL4 cells lies between the activation of protein kinase C and the activation of MAPKK.

Collagens--structure, function, and biosynthesis.

Science.gov (United States)

Gelse, K; Pöschl, E; Aigner, T

2003-11-28

The extracellular matrix represents a complex alloy of variable members of diverse protein families defining structural integrity and various physiological functions. The most abundant family is the collagens with more than 20 different collagen types identified so far. Collagens are centrally involved in the formation of fibrillar and microfibrillar networks of the extracellular matrix, basement membranes as well as other structures of the extracellular matrix. This review focuses on the distribution and function of various collagen types in different tissues. It introduces their basic structural subunits and points out major steps in the biosynthesis and supramolecular processing of fibrillar collagens as prototypical members of this protein family. A final outlook indicates the importance of different collagen types not only for the understanding of collagen-related diseases, but also as a basis for the therapeutical use of members of this protein family discussed in other chapters of this issue.

Phase II marker-driven trial of panitumumab and chemotherapy in KRAS wild-type biliary tract cancer

DEFF Research Database (Denmark)

Jensen, L H; Lindebjerg, J; Ploen, J

2012-01-01

BACKGROUND: Combination chemotherapy has proven beneficial in biliary tract cancer and further improvements may be achieved by individualizing treatment based on biomarkers and by adding biological agents. We report the effect of chemotherapy with panitumumab as first-line therapy for KRAS wild....... Combination chemotherapy with panitumumab in patients with KRAS wild-type tumors met the efficacy criteria for future testing in a randomized trial....

Citrulline diet supplementation improves specific age-related raft changes in wild-type rodent hippocampus.

Science.gov (United States)

Marquet-de Rougé, Perrine; Clamagirand, Christine; Facchinetti, Patricia; Rose, Christiane; Sargueil, Françoise; Guihenneuc-Jouyaux, Chantal; Cynober, Luc; Moinard, Christophe; Allinquant, Bernadette

2013-10-01

The levels of molecules crucial for signal transduction processing change in the brain with aging. Lipid rafts are membrane microdomains involved in cell signaling. We describe here substantial biophysical and biochemical changes occurring within the rafts in hippocampus neurons from aging wild-type rats and mice. Using continuous sucrose density gradients, we observed light-, medium-, and heavy raft subpopulations in young adult rodent hippocampus neurons containing very low levels of amyloid precursor protein (APP) and almost no caveolin-1 (CAV-1). By contrast, old rodents had a homogeneous age-specific high-density caveolar raft subpopulation containing significantly more cholesterol (CHOL), CAV-1, and APP. C99-APP-Cter fragment detection demonstrates that the first step of amyloidogenic APP processing takes place in this caveolar structure during physiological aging of the rat brain. In this age-specific caveolar raft subpopulation, levels of the C99-APP-Cter fragment are exponentially correlated with those of APP, suggesting that high APP concentrations may be associated with a risk of large increases in beta-amyloid peptide levels. Citrulline (an intermediate amino acid of the urea cycle) supplementation in the diet of aged rats for 3 months reduced these age-related hippocampus raft changes, resulting in raft patterns tightly close to those in young animals: CHOL, CAV-1, and APP concentrations were significantly lower and the C99-APP-Cter fragment was less abundant in the heavy raft subpopulation than in controls. Thus, we report substantial changes in raft structures during the aging of rodent hippocampus and describe new and promising areas of investigation concerning the possible protective effect of citrulline on brain function during aging.

Structure, function and folding of phosphoglycerate kinase are strongly perturbed by macromolecular crowding.

Science.gov (United States)

Samiotakis, Antonios; Dhar, Apratim; Ebbinghaus, Simon; Nienhaus, Lea; Homouz, Dirar; Gruebele, Martin; Cheung, Margaret

2010-10-01

We combine experiment and computer simulation to show how macromolecular crowding dramatically affects the structure, function and folding landscape of phosphoglycerate kinase (PGK). Fluorescence labeling shows that compact states of yeast PGK are populated as the amount of crowding agents (Ficoll 70) increases. Coarse-grained molecular simulations reveal three compact ensembles: C (crystal structure), CC (collapsed crystal) and Sph (spherical compact). With an adjustment for viscosity, crowded wild type PGK and fluorescent PGK are about 15 times or more active in 200 mg/ml Ficoll than in aqueous solution. Our results suggest a new solution to the classic problem of how the ADP and diphosphoglycerate binding sites of PGK come together to make ATP: rather than undergoing a hinge motion, the ADP and substrate sites are already located in proximity under crowded conditions that mimic the in vivo conditions under which the enzyme actually operates.

A ten-week biochemistry lab project studying wild-type and mutant bacterial alkaline phosphatase.

Science.gov (United States)

Witherow, D Scott

2016-11-12

This work describes a 10-week laboratory project studying wild-type and mutant bacterial alkaline phosphatase, in which students purify, quantitate, and perform kinetic assays on wild-type and selected mutants of the enzyme. Students also perform plasmid DNA purification, digestion, and gel analysis. In addition to simply learning important techniques, students acquire novel biochemical data in their kinetic analysis of mutant enzymes. The experiments are designed to build on students' work from week to week in a way that requires them to apply quantitative analysis and reasoning skills, reinforcing traditional textbook biochemical concepts. Students are assessed through lab reports focused on journal style writing, quantitative and conceptual question sheets, and traditional exams. © 2016 by The International Union of Biochemistry and Molecular Biology, 44(6):555-564, 2016. © 2016 The International Union of Biochemistry and Molecular Biology.

A multiplex reverse transcription-nested polymerase chain reaction for detection and differentiation of wild-type and vaccine strains of canine distemper virus

Directory of Open Access Journals (Sweden)

Cui Shang-jin

2010-05-01

Full Text Available Abstract A multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR method was developed for the detection and differentiation of wild-type and vaccine strains of canine distemper virus (CDV. A pair of primers (P1 and P4 specific for CDV corresponding to the highly conserved region of the CDV genome were used as a common primer pair in the first-round PCR of the nested PCR. Primers P2 specific for CDV wild-type strains, were used as the forward primer together with the common reverse primer P4 in the second round of nested PCR. Primers P3, P5 specific for CDV wild-type strain or vaccine strain, were used as the forward primer together with the common reverse primer P4+P6 in the second round of nested PCR. A fragment of 177 bp was amplified from vaccine strain genomic RNA, and a fragment of 247 bp from wild-type strain genomic RNA in the RT-nPCR, and two fragments of 247 bp and 177 bp were amplified from the mixed samples of vaccine and wild-type strains. No amplification was achieved for uninfected cells, or cells infected with Newcastle disease virus (NDV, canine parvovirus (CPV, canine coronavirus (CCV, rabies virus (RV, or canine adenovirus (CAV. The RT-nPCR method was used to detect 30 field samples suspected of canine distemper from Heilongjiang and Jilin Provinces, and 51 samples in Shandong province. As a result of 30 samples, were found to be wild-type-like, and 5 to be vaccine-strain-like. The RT-nPCR method can be used to effectively detect and differentiate wild-type CDV-infected dogs from dogs vaccinated with CDV vaccine, and thus can be used in clinical detection and epidemiological surveillance.

A multiplex reverse transcription-nested polymerase chain reaction for detection and differentiation of wild-type and vaccine strains of canine distemper virus

Science.gov (United States)

2010-01-01

A multiplex reverse transcription-nested polymerase chain reaction (RT-nPCR) method was developed for the detection and differentiation of wild-type and vaccine strains of canine distemper virus (CDV). A pair of primers (P1 and P4) specific for CDV corresponding to the highly conserved region of the CDV genome were used as a common primer pair in the first-round PCR of the nested PCR. Primers P2 specific for CDV wild-type strains, were used as the forward primer together with the common reverse primer P4 in the second round of nested PCR. Primers P3, P5 specific for CDV wild-type strain or vaccine strain, were used as the forward primer together with the common reverse primer P4+P6 in the second round of nested PCR. A fragment of 177 bp was amplified from vaccine strain genomic RNA, and a fragment of 247 bp from wild-type strain genomic RNA in the RT-nPCR, and two fragments of 247 bp and 177 bp were amplified from the mixed samples of vaccine and wild-type strains. No amplification was achieved for uninfected cells, or cells infected with Newcastle disease virus (NDV), canine parvovirus (CPV), canine coronavirus (CCV), rabies virus (RV), or canine adenovirus (CAV). The RT-nPCR method was used to detect 30 field samples suspected of canine distemper from Heilongjiang and Jilin Provinces, and 51 samples in Shandong province. As a result of 30 samples, were found to be wild-type-like, and 5 to be vaccine-strain-like. The RT-nPCR method can be used to effectively detect and differentiate wild-type CDV-infected dogs from dogs vaccinated with CDV vaccine, and thus can be used in clinical detection and epidemiological surveillance. PMID:20433759

Microbial Community and Functional Structure Significantly Varied among Distinct Types of Paddy Soils But Responded Differently along Gradients of Soil Depth Layers

Directory of Open Access Journals (Sweden)

Ren Bai

2017-05-01

Full Text Available Paddy rice fields occupy broad agricultural area in China and cover diverse soil types. Microbial community in paddy soils is of great interest since many microorganisms are involved in soil functional processes. In the present study, Illumina Mi-Seq sequencing and functional gene array (GeoChip 4.2 techniques were combined to investigate soil microbial communities and functional gene patterns across the three soil types including an Inceptisol (Binhai, an Oxisol (Leizhou, and an Ultisol (Taoyuan along four profile depths (up to 70 cm in depth in mesocosm incubation columns. Detrended correspondence analysis revealed that distinctly differentiation in microbial community existed among soil types and profile depths, while the manifest variance in functional structure was only observed among soil types and two rice growth stages, but not across profile depths. Along the profile depth within each soil type, Acidobacteria, Chloroflexi, and Firmicutes increased whereas Cyanobacteria, β-proteobacteria, and Verrucomicrobia declined, suggesting their specific ecophysiological properties. Compared to bacterial community, the archaeal community showed a more contrasting pattern with the predominant groups within phyla Euryarchaeota, Thaumarchaeota, and Crenarchaeota largely varying among soil types and depths. Phylogenetic molecular ecological network (pMEN analysis further indicated that the pattern of bacterial and archaeal communities interactions changed with soil depth and the highest modularity of microbial community occurred in top soils, implying a relatively higher system resistance to environmental change compared to communities in deeper soil layers. Meanwhile, microbial communities had higher connectivity in deeper soils in comparison with upper soils, suggesting less microbial interaction in surface soils. Structure equation models were developed and the models indicated that pH was the most representative characteristics of soil type and

Microbial Community and Functional Structure Significantly Varied among Distinct Types of Paddy Soils But Responded Differently along Gradients of Soil Depth Layers.

Science.gov (United States)

Bai, Ren; Wang, Jun-Tao; Deng, Ye; He, Ji-Zheng; Feng, Kai; Zhang, Li-Mei

2017-01-01

Paddy rice fields occupy broad agricultural area in China and cover diverse soil types. Microbial community in paddy soils is of great interest since many microorganisms are involved in soil functional processes. In the present study, Illumina Mi-Seq sequencing and functional gene array (GeoChip 4.2) techniques were combined to investigate soil microbial communities and functional gene patterns across the three soil types including an Inceptisol (Binhai), an Oxisol (Leizhou), and an Ultisol (Taoyuan) along four profile depths (up to 70 cm in depth) in mesocosm incubation columns. Detrended correspondence analysis revealed that distinctly differentiation in microbial community existed among soil types and profile depths, while the manifest variance in functional structure was only observed among soil types and two rice growth stages, but not across profile depths. Along the profile depth within each soil type, Acidobacteria , Chloroflexi , and Firmicutes increased whereas Cyanobacteria , β -proteobacteria , and Verrucomicrobia declined, suggesting their specific ecophysiological properties. Compared to bacterial community, the archaeal community showed a more contrasting pattern with the predominant groups within phyla Euryarchaeota , Thaumarchaeota , and Crenarchaeota largely varying among soil types and depths. Phylogenetic molecular ecological network (pMEN) analysis further indicated that the pattern of bacterial and archaeal communities interactions changed with soil depth and the highest modularity of microbial community occurred in top soils, implying a relatively higher system resistance to environmental change compared to communities in deeper soil layers. Meanwhile, microbial communities had higher connectivity in deeper soils in comparison with upper soils, suggesting less microbial interaction in surface soils. Structure equation models were developed and the models indicated that pH was the most representative characteristics of soil type and

Mid-aged and aged wild-type and progestin receptor knockout (PRKO) mice demonstrate rapid progesterone and 3alpha,5alpha-THP-facilitated lordosis.

Science.gov (United States)

Frye, C A; Sumida, K; Lydon, J P; O'Malley, B W; Pfaff, D W

2006-05-01

Progesterone (P) and its 5alpha-reduced metabolite, 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha,5alpha-THP), facilitate sexual behavior of rodents via agonist-like actions at intracellular progestin receptors (PRs) and membrane GABA(A)/benzodiazepine receptor complexes (GBRs), respectively. Given that ovarian secretion of progestins declines with aging, whether or not senescent mice are responsive to progestins was of interest. Homozygous PR knockout (PRKO) or wild-type mice that were between 10-12 (mid-aged) or 20-24 (aged) months of age were administered P or 3alpha,5alpha-THP, and the effect on lordosis were examined. Effects of a progestin-priming regimen that enhances PR-mediated (experiment 1) or more rapid, PR-independent effects of progestins (experiments 2 and 3) on sexual behavior were examined. Levels of P, 3alpha,5alpha-THP, and muscimol binding were examined in tissues from aged mice (experiment 4). Wild-type, but not PRKO, mice were responsive when primed with 17beta-estradiol (E(2); 0.5 microg) and administered P (500 microg, subcutaneously). Mid-aged wild-type mice demonstrated greater increases in lordosis 6 h later compared to their pre-P, baseline test than did aged wild-type mice (experiment 1). Lordosis of younger and older wild-type, but not PRKO, mice was significantly increased within 5 min of intravenous (IV) administration of P (100 ng), compared with E(2)-priming alone (experiment 2). However, wild-type and PRKO mice demonstrated significant increases in lordosis 5 min after IV administration of 3alpha,5alpha-THP, an effect which was more pronounced in mid-aged than in aged animals (100 ng-experiment 3). In tissues from aged wild-type and PRKO mice, levels of P, 3alpha,5alpha-THP, and muscimol binding were increased by P administration (experiment 4). PR binding was lower in the cortex of PRKO than that of wild-type mice. Mid-aged and aged PRKO and wild-type mice demonstrated rapid P or 3alpha,5alpha-THP-facilitated lordosis that may be

Linezolid-Dependent Function and Structure Adaptation of Ribosomes in a Staphylococcus epidermidis Strain Exhibiting Linezolid Dependence

Science.gov (United States)

Kokkori, Sofia; Apostolidi, Maria; Tsakris, Athanassios; Pournaras, Spyros

2014-01-01

Linezolid-dependent growth was recently reported in Staphylococcus epidermidis clinical strains carrying mutations associated with linezolid resistance. To investigate this unexpected behavior at the molecular level, we isolated active ribosomes from one of the linezolid-dependent strains and we compared them with ribosomes isolated from a wild-type strain. Both strains were grown in the absence and presence of linezolid. Detailed biochemical and structural analyses revealed essential differences in the function and structure of isolated ribosomes which were assembled in the presence of linezolid. The catalytic activity of peptidyltransferase was found to be significantly higher in the ribosomes derived from the linezolid-dependent strain. Interestingly, the same ribosomes exhibited an abnormal ribosomal subunit dissociation profile on a sucrose gradient in the absence of linezolid, but the profile was restored after treatment of the ribosomes with an excess of the antibiotic. Our study suggests that linezolid most likely modified the ribosomal assembly procedure, leading to a new functional ribosomal population active only in the presence of linezolid. Therefore, the higher growth rate of the partially linezolid-dependent strains could be attributed to the functional and structural adaptations of ribosomes to linezolid. PMID:24890589

Molecular characterization and functional analysis of pteridine reductase in wild-type and antimony-resistant Leishmania lines.

Science.gov (United States)

de Souza Moreira, Douglas; Ferreira, Rafael Fernandes; Murta, Silvane M F

2016-01-01

Pteridine reductase (PTR1) is an NADPH-dependent reductase that participates in the salvage of pteridines, which are essential to maintain growth of Leishmania. In this study, we performed the molecular characterization of ptr1 gene in wild-type (WTS) and SbIII-resistant (SbR) lines from Leishmania guyanensis (Lg), Leishmania amazonensis (La), Leishmania braziliensis (Lb) and Leishmania infantum (Li), evaluating the chromosomal location, mRNA levels of the ptr1 gene and PTR1 protein expression. PFGE results showed that the ptr1 gene is located in a 797 kb chromosomal band in all Leishmania lines analyzed. Interestingly, an additional chromosomal band of 1070 kb was observed only in LbSbR line. Northern blot results showed that the levels of ptr1 mRNA are increased in the LgSbR, LaSbR and LbSbR lines. Western blot assays using the polyclonal anti-LmPTR1 antibody demonstrated that PTR1 protein is more expressed in the LgSbR, LaSbR and LbSbR lines compared to their respective WTS counterparts. Nevertheless, no difference in the level of mRNA and protein was observed between the LiWTS and LiSbR lines. Functional analysis of PTR1 enzyme was performed to determine whether the overexpression of ptr1 gene in the WTS L. braziliensis and L. infantum lines would change the SbIII-resistance phenotype of transfected parasites. Western blot results showed that the expression level of PTR1 protein was increased in the transfected parasites compared to the non-transfected ones. IC50 analysis revealed that the overexpression of ptr1 gene in the WTS L. braziliensis line increased 2-fold the SbIII-resistance phenotype compared to the non-transfected counterpart. Furthermore, the overexpression of ptr1 gene in the WTS L. infantum line did not change the SbIII-resistance phenotype. These results suggest that the PTR1 enzyme may be implicated in the SbIII-resistance phenotype in L. braziliensis line. Copyright © 2015 Elsevier Inc. All rights reserved.

Common variants of the BRCA1 wild-type allele modify the risk of breast cancer in BRCA1 mutation carriers.

OpenAIRE

Cox, D. G.; Simard, J.; Sinnett, D.; Hamdi, Y.; Soucy, P.; Ouimet, M.; Barjhoux, L.; Verny-Pierre, C.; McGuffog, L.; Healey, S.; Szabo, C.; Greene, M. H.; Mai, P. L.; Andrulis, I. L.; Thomassen, M.

2011-01-01

Mutations in the BRCA1 gene substantially increase a woman's lifetime risk of breast cancer. However, there is great variation in this increase in risk with several genetic and non-genetic modifiers identified. The BRCA1 protein plays a central role in DNA repair, a mechanism that is particularly instrumental in safeguarding cells against tumorigenesis. We hypothesized that polymorphisms that alter the expression and/or function of BRCA1 carried on the wild-type (non-mutated) copy of the BRCA...

Structural basis for type VI secreted peptidoglycan dl-endopeptidase function, specificity and neutralization in Serratia marcescens

Energy Technology Data Exchange (ETDEWEB)

Srikannathasan, Velupillai; English, Grant [University of Dundee, Dundee DD1 5EH, Scotland (United Kingdom); Bui, Nhat Khai [Newcastle University, Newcastle upon Tyne NE2 4HH (United Kingdom); Trunk, Katharina; O’Rourke, Patrick E. F.; Rao, Vincenzo A. [University of Dundee, Dundee DD1 5EH, Scotland (United Kingdom); Vollmer, Waldemar [Newcastle University, Newcastle upon Tyne NE2 4HH (United Kingdom); Coulthurst, Sarah J., E-mail: s.j.coulthurst@dundee.ac.uk; Hunter, William N., E-mail: s.j.coulthurst@dundee.ac.uk [University of Dundee, Dundee DD1 5EH, Scotland (United Kingdom)

2013-12-01

Crystal structures of type VI secretion system-associated immunity proteins, a peptidoglycan endopeptidase and a complex of the endopeptidase and its cognate immunity protein are reported together with assays of endopeptidase activity and functional assessment. Some Gram-negative bacteria target their competitors by exploiting the type VI secretion system to extrude toxic effector proteins. To prevent self-harm, these bacteria also produce highly specific immunity proteins that neutralize these antagonistic effectors. Here, the peptidoglycan endopeptidase specificity of two type VI secretion-system-associated effectors from Serratia marcescens is characterized. These small secreted proteins, Ssp1 and Ssp2, cleave between γ-d-glutamic acid and l-meso-diaminopimelic acid with different specificities. Ssp2 degrades the acceptor part of cross-linked tetratetrapeptides. Ssp1 displays greater promiscuity and cleaves monomeric tripeptides, tetrapeptides and pentapeptides and dimeric tetratetra and tetrapenta muropeptides on both the acceptor and donor strands. Functional assays confirm the identity of a catalytic cysteine in these endopeptidases and crystal structures provide information on the structure–activity relationships of Ssp1 and, by comparison, of related effectors. Functional assays also reveal that neutralization of these effectors by their cognate immunity proteins, which are called resistance-associated proteins (Raps), contributes an essential role to cell fitness. The structures of two immunity proteins, Rap1a and Rap2a, responsible for the neutralization of Ssp1 and Ssp2-like endopeptidases, respectively, revealed two distinct folds, with that of Rap1a not having previously been observed. The structure of the Ssp1–Rap1a complex revealed a tightly bound heteromeric assembly with two effector molecules flanking a Rap1a dimer. A highly effective steric block of the Ssp1 active site forms the basis of effector neutralization. Comparisons with Ssp2–Rap2

Structural basis for type VI secreted peptidoglycan dl-endopeptidase function, specificity and neutralization in Serratia marcescens

International Nuclear Information System (INIS)

Srikannathasan, Velupillai; English, Grant; Bui, Nhat Khai; Trunk, Katharina; O’Rourke, Patrick E. F.; Rao, Vincenzo A.; Vollmer, Waldemar; Coulthurst, Sarah J.; Hunter, William N.

2013-01-01

Crystal structures of type VI secretion system-associated immunity proteins, a peptidoglycan endopeptidase and a complex of the endopeptidase and its cognate immunity protein are reported together with assays of endopeptidase activity and functional assessment. Some Gram-negative bacteria target their competitors by exploiting the type VI secretion system to extrude toxic effector proteins. To prevent self-harm, these bacteria also produce highly specific immunity proteins that neutralize these antagonistic effectors. Here, the peptidoglycan endopeptidase specificity of two type VI secretion-system-associated effectors from Serratia marcescens is characterized. These small secreted proteins, Ssp1 and Ssp2, cleave between γ-d-glutamic acid and l-meso-diaminopimelic acid with different specificities. Ssp2 degrades the acceptor part of cross-linked tetratetrapeptides. Ssp1 displays greater promiscuity and cleaves monomeric tripeptides, tetrapeptides and pentapeptides and dimeric tetratetra and tetrapenta muropeptides on both the acceptor and donor strands. Functional assays confirm the identity of a catalytic cysteine in these endopeptidases and crystal structures provide information on the structure–activity relationships of Ssp1 and, by comparison, of related effectors. Functional assays also reveal that neutralization of these effectors by their cognate immunity proteins, which are called resistance-associated proteins (Raps), contributes an essential role to cell fitness. The structures of two immunity proteins, Rap1a and Rap2a, responsible for the neutralization of Ssp1 and Ssp2-like endopeptidases, respectively, revealed two distinct folds, with that of Rap1a not having previously been observed. The structure of the Ssp1–Rap1a complex revealed a tightly bound heteromeric assembly with two effector molecules flanking a Rap1a dimer. A highly effective steric block of the Ssp1 active site forms the basis of effector neutralization. Comparisons with Ssp2–Rap2

[Study on identification of wild Dipsacus asper and cultivated Dipsacus asper at different growth years by organization structure].

Science.gov (United States)

Yang, Jun; Peng, Hua-Sheng; Xuan, Yan; Qu, Jie

2012-12-01

To study the regular pattern of growing development and the structural changes of the main root of Dipsacus asper; Find the basis for distinguishing between wild and cultivated Dipsacus asper as well as identifying cultivated Dipsacus asper at different growth years. Continuous sectional sampling and histological observation was conducted on the main roots of annual and biennial cultivated Dipsacus asper and on the main roots of bolting and non-bolting plants of wild Dipsacus asper; Histological observation was carried out respectively on the main roots of one to four-year-old cultivated Dipsacus asper. It was discovered for the first time that there was structure of growth rings in the main roots of Dipsacus asper. And the ways in which the vessels of the growth rings were arranged differed between wild and cultivated Dipsacus asper, the vessels of growth rings in wild Dipsacus asper were arranged tangentially while those in cultivated Dipsacus asper were arranged as clusters. Pith existed in the upper part of roots of wild Dipsacus asper,while it was not found in the cultivated ones, and this could be adopted as a preliminary method to distinguish the wild Dipsacus asper from cultivatedone. The structures of growth rings in each main root were in consistency from top to bottom, and the thickening of roots mainly depended on the growth of xylem in the first year. There were particular relations and regular patterns between the structure of main roots of one to four-year-old cultivated Dipsacus asper and the ages of their growth, which could serve as the primary basis for identifying their growth years. Methods to distinguish wild Dipsacus asper from cultivated ones at different ages by organization structure are established, which could provide the basis for identification of the medicinal material of Dipsacus asper.

Wild harvest

NARCIS (Netherlands)

Cruz-Garcia, G.S.; Struik, P.C.; Johnson, D.E.

2016-01-01

Rice fields provide not only a staple food but are also bio-diverse and multi-functional ecosystems. Wild food plants are important elements of biodiversity in rice fields and are critical components to the subsistence of poor farmers. The spatial and seasonal distribution of wild food plants

Roentgenological structural types of the unaltered breast

International Nuclear Information System (INIS)

Kolganova, I.P.; Zal'tsman, I.N.

1981-01-01

The authors present a roentgenoanatomical analysis of normal breast specimens and mammograms of 324 healthy women aged 31 to 60. Four roentgenological structural types of the unaltered breast have been singled out: linear-reticular, lamellar-cellular, large focal, and a single polygonal shadow. These structural types were detected in age groups from 31 to 40, from 41 to 50, and from 51 to 60 with various frequency. Each type corresponds to a certain morphological and functional state of the breast. The frequency of the 2nd and 4th types decreases and of the 1st type increases with advancing age [ru

Roentgenological structural types of the unaltered breast

Energy Technology Data Exchange (ETDEWEB)

Kolganova, I P; Zal' tsman, I N [Akademiya Meditsinskikh Nauk RSFSR, Moscow. Pervyj Moskovskij Meditsinskij Inst.

1981-11-01

The authors present a roentgenoanatomical analysis of normal breast specimens and mammograms of 324 healthy women aged 31 to 60. Four roentgenological structural types of the unaltered breast have been singled out: linear-reticular, lamellar-cellular, large focal, and a single polygonal shadow. These structural types were detected in age groups from 31 to 40, from 41 to 50, and from 51 to 60 with various frequency. Each type corresponds to a certain morphological and functional state of the breast. The frequency of the 2nd and 4th types decreases and of the 1st type increases with advancing age.

Changes in fatty acid content and composition between wild type and CsHMA3 overexpressing Camelina sativa under heavy-metal stress.

Science.gov (United States)

Park, Won; Feng, Yufeng; Kim, Hyojin; Suh, Mi Chung; Ahn, Sung-Ju

2015-09-01

Under heavy-metal stress, CsHMA3 overexpressing transgenic Camelina plants displayed not only a better quality, but also a higher quantity of unsaturated fatty acids in their seeds compared with wild type. Camelina sativa L. belongs to the Brassicaceae family and is frequently used as a natural vegetable oil source, as its seeds contain a high content of fatty acids. In this study, we observed that, when subjected to heavy metals (Cd, Co, Zn and Pb), the seeds of CsHMA3 (Heavy-Metal P1B-ATPase 3) transgenic lines retained their original golden yellow color and smooth outline, unlike wild-type seeds. Furthermore, we investigated the fatty acids content and composition of wild type and CsHMA3 transgenic lines after heavy metal treatments compared to the control. The results showed higher total fatty acid amounts in seeds of CsHMA3 transgenic lines compared with those in wild-type seeds under heavy-metal stresses. In addition, the compositions of unsaturated fatty acids-especially 18:1 (oleic acid), 18:2 (linoleic acid; only in case of Co treatment), 18:3 (linolenic acid) and 20:1 (eicosenoic acid)-in CsHMA3 overexpressing transgenic lines treated with heavy metals were higher than those of wild-type seeds under the same conditions. Furthermore, reactive oxygen species (ROS) contents in wild-type leaves and roots when treated with heavy metal were higher than in CsHMA3 overexpressing transgenic lines. These results indicate that overexpression of CsHMA3 affects fatty acid composition and content-factors that are responsible for the fuel properties of biodiesel-and can alleviate ROS accumulation caused by heavy-metal stresses in Camelina. Due to these factors, we propose that CsHMA3 transgenic Camelina can be used for phytoremediation of metal-contaminated soil as well as for oil production.

The Use of EGFR Tyrosine Kinase Inhibitors in EGFR Wild-Type Non-Small-Cell Lung Cancer.

Science.gov (United States)

Stinchcombe, Thomas E

2016-04-01

The objective response rate and progression-free survival observed with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) in patients with metastatic epidermal growth factor receptor (EGFR) wild-type non-small cell lung cancer (NSCLC) are modest. The adverse events associated with EGFR TKIs are manageable but they must be considered in the context of the limited efficacy. The development of anti-PD-1 immunotherapy as second-line therapy has reduced the role of EGFR TKIs in EGFR wild-type NSCLC. Recently, there has been increased recognition of the benefit of the earlier integration of palliative care and symptom management, and this is reasonable alternative to treatment with an EGFR TKI for many patients. My practice pattern for patients with EGFR wild-type NSCLC is platinum-based chemotherapy as first-line therapy, immunotherapy as second-line therapy, and single-agent chemotherapy as third-line therapy for patients with preserved performance status who want to pursue further therapy. Only a small proportion of patients are eligible for fourth-line therapy, and I prefer to enroll them in clinical trials rather than use EGFR TKIs. I suspect that the use of EGFR TKIs in clinical use and as a comparator arm for clinical trials will continue to decline over the next several years.

Structure and functions of exopolysaccharide produced by gut commensal Lactobacillus reuteri 100-23.

Science.gov (United States)

Sims, Ian M; Frese, Steven A; Walter, Jens; Loach, Diane; Wilson, Michelle; Appleyard, Kay; Eason, Jocelyn; Livingston, Megan; Baird, Margaret; Cook, Gregory; Tannock, Gerald W

2011-07-01

Lactobacillus reuteri strain 100-23 together with a Lactobacillus-free mouse model, provides a system with which the molecular traits underpinning bacterial commensalism in vertebrates can be studied. A polysaccharide was extracted from sucrose-containing liquid cultures of strain 100-23. Chemical analysis showed that this exopolysaccharide was a levan (β-2, 6-linked fructan). Mutation of the fructosyl transferase (ftf) gene resulted in loss of exopolysaccharide production. The ftf mutant was able to colonise the murine gastrointestinal tract in the absence of competition, but colonisation was impaired in competition with the wild type. Biofilm formation by the mutant on the forestomach epithelial surface was not impaired and the matrix between cells was indistinguishable from that of the wild type in electron micrographs. Colonisation of the mouse gut by the wild-type strain led to increased proportions of regulatory T cells (Foxp3+) in the spleen, whereas colonisation by the ftf mutant did not. Survival of the mutant in sucrose-containing medium was markedly reduced relative to the wild type. Comparison of the genomic ftf loci of strain 100-23 with other L. reuteri strains suggested that the ftf gene was acquired by lateral gene transfer early in the evolution of the species and subsequently diversified at accelerated rates. Levan production by L. reuteri 100-23 may represent a function acquired by the bacterial species for life in moderate to high-sucrose extra-gastrointestinal environments that has subsequently been diverted to novel uses, including immunomodulation, that aid in colonisation of the murine gut.

Establishment of new transmissible and drug-sensitive human immunodeficiency virus type 1 wild types due to transmission of nucleoside analogue-resistant virus.

Science.gov (United States)

de Ronde, A; van Dooren, M; van Der Hoek, L; Bouwhuis, D; de Rooij, E; van Gemen, B; de Boer, R; Goudsmit, J

2001-01-01

Sequence analysis of human immunodeficiency virus type 1 (HIV-1) from 74 persons with acute infections identified eight strains with mutations in the reverse transcriptase (RT) gene at positions 41, 67, 68, 70, 215, and 219 associated with resistance to the nucleoside analogue zidovudine (AZT). Follow-up of the fate of these resistant HIV-1 strains in four newly infected individuals revealed that they were readily replaced by sensitive strains. The RT of the resistant viruses changed at amino acid 215 from tyrosine (Y) to aspartic acid (D) or serine (S), with asparagine (N) as a transient intermediate, indicating the establishment of new wild types. When we introduced these mutations and the original threonine (T)-containing wild type into infectious molecular clones and assessed their competitive advantage in vitro, the order of fitness was in accord with the in vivo observations: 215Y types with D, S, or N residues at position 215 may be warranted in order to estimate the threat to long-term efficacy of regimens including nucleoside analogues.

Biological activity and safety of adenoviral vector-expressed wild-type p53 after intratumoral injection in melanoma and breast cancer patients with p53-overexpressing tumors

NARCIS (Netherlands)

Dummer, R; Bergh, J; Karlsson, Y; Horovitz, JA; Mulder, NH; Huinin, DT; Burg, G; Hofbauer, G; Osanto, S

p53 mutations are common genetic alterations in human cancer. Gene transfer of a wild-type (wt) p53 gene reverses the loss of normal p53 function in vitro and in vivo. A phase I dose escalation study of single intratumoral (i.t.) injection of a replication-defective adenoviral expression vector

Black bear parathyroid hormone has greater anabolic effects on trabecular bone in dystrophin-deficient mice than in wild type mice.

Science.gov (United States)

Gray, Sarah K; McGee-Lawrence, Meghan E; Sanders, Jennifer L; Condon, Keith W; Tsai, Chung-Jui; Donahue, Seth W

2012-09-01

Duchenne muscular dystrophy (DMD) is an X-linked neuromuscular disease that has deleterious consequences in muscle and bone, leading to decreased mobility, progressive osteoporosis, and premature death. Patients with DMD experience a higher-than-average fracture rate, particularly in the proximal and distal femur and proximal tibia. The dystrophin-deficient mdx mouse is a model of DMD that demonstrates muscle degeneration and fibrosis and osteoporosis. Parathyroid hormone, an effective anabolic agent for post-menopausal and glucocorticoid-induced osteoporosis, has not been explored for DMD. Black bear parathyroid hormone (bbPTH) has been implicated in the maintenance of bone properties during extended periods of disuse (hibernation). We cloned bbPTH and found 9 amino acid residue differences from human PTH. Apoptosis was mitigated and cAMP was activated by bbPTH in osteoblast cultures. We administered 28nmol/kg of bbPTH 1-84 to 4-week old male mdx and wild type mice via daily (5×/week) subcutaneous injection for 6 weeks. Vehicle-treated mdx mice had 44% lower trabecular bone volume fraction than wild type mice. No changes were found in femoral cortical bone geometry or mechanical properties with bbPTH treatment in wild type mice, and only medio-lateral moment of inertia changed with bbPTH treatment in mdx femurs. However, μCT analyses of the trabecular regions of the distal femur and proximal tibia showed marked increases in bone volume fraction with bbPTH treatment, with a greater anabolic response (7-fold increase) in mdx mice than wild type mice (2-fold increase). Trabecular number increased in mdx long bone, but not wild type bone. Additionally, greater osteoblast area and decreased osteoclast area were observed with bbPTH treatment in mdx mice. The heightened response to PTH in mdx bone compared to wild type suggests a link between dystrophin deficiency, altered calcium signaling, and bone. These findings support further investigation of PTH as an anabolic

Host and Pathogen Copper-Transporting P-Type ATPases Function Antagonistically during Salmonella Infection.

Science.gov (United States)

Ladomersky, Erik; Khan, Aslam; Shanbhag, Vinit; Cavet, Jennifer S; Chan, Jefferson; Weisman, Gary A; Petris, Michael J

2017-09-01

Copper is an essential yet potentially toxic trace element that is required by all aerobic organisms. A key regulator of copper homeostasis in mammalian cells is the copper-transporting P-type ATPase ATP7A, which mediates copper transport from the cytoplasm into the secretory pathway, as well as copper export across the plasma membrane. Previous studies have shown that ATP7A-dependent copper transport is required for killing phagocytosed Escherichia coli in a cultured macrophage cell line. In this investigation, we expanded on these studies by generating Atp7a LysMcre mice, in which the Atp7a gene was specifically deleted in cells of the myeloid lineage, including macrophages. Primary macrophages isolated from Atp7a LysMcre mice exhibit decreased copper transport into phagosomal compartments and a reduced ability to kill Salmonella enterica serovar Typhimurium compared to that of macrophages isolated from wild-type mice. The Atp7a LysMcre mice were also more susceptible to systemic infection by S Typhimurium than wild-type mice. Deletion of the S Typhimurium copper exporters, CopA and GolT, was found to decrease infection in wild-type mice but not in the Atp7a LysMcre mice. These studies suggest that ATP7A-dependent copper transport into the phagosome mediates host defense against S Typhimurium, which is counteracted by copper export from the bacteria via CopA and GolT. These findings reveal unique and opposing functions for copper transporters of the host and pathogen during infection. Copyright © 2017 American Society for Microbiology.

Effects of different types of moderate severity disturbance on forest structural complexity and ecosystem functioning: A story of ice and fire

Science.gov (United States)

Fahey, R. T.; Atkins, J.; Gough, C. M.; Hardiman, B. S.; Haber, L.; Stuart-Haentjens, E.; David, O.; Campbell, J. L.; Rustad, L.; Duffy, M.

2017-12-01

Disturbances that alter the structure and function of forest ecosystems occur along a continuum of severity. In contrast to the extremes of the disturbance gradient (i.e., stand-replacing disturbance and small gap formation), moderate severity disturbances are poorly understood, even though they make up the majority of the gradient and their spatial extent (and likely overall importance to regional disturbance regimes) often exceeds that of more severe disturbances. Moderate severity disturbances originate from a variety of causes, such as fires, ice storms, or pest and pathogen outbreaks, and each of these could reshape structure and function in different ways. Observational data from a limited number of sites shows that moderate disturbance can increase ecosystem complexity, but the generality of this effect has not been tested across a broad range of disturbance types and severities. Here, we utilize data from a set of five case studies of experimental or natural moderate disturbance to assess the effects of different types and severities of disturbance on forest canopy structural complexity (CSC) and the relationship of canopy structure with ecosystem functioning. Using pre- and post-disturbance measures of CSC derived from aerial and terrestrial LiDAR, UAV imagery, and Landsat data we quantified changes in CSC following an experimental ice storm, a low-severity surface fire, Beech Bark Disease and Hemlock Wooly Adelgid outbreaks, and experimental accelerated succession. Our initial findings indicate that different disturbance types have highly variable effects on CSC, and also that progressive increases in disturbance severity alter CSC differently among disturbance types. Differential effects of variable disturbance types on CSC has implications for the carbon cycle, as forest structure is strongly linked with both growth-limiting resource (e.g., nutrients and light) acquisition and net primary productivity. Understanding how different types and severities of

Effects of age and liquid holding on the UV-radiation sensitivities of wild-type and mutant Caenorhabditis elegans dauer larvae

Energy Technology Data Exchange (ETDEWEB)

Hartman, P S

1984-01-01

The dauer larva is a facultative developmental stage in the life cycle of the nematode Caenorhabditis elegans. Dauer larvae, which can survive under starvation for over 60 days, resume normal development when feeding is resumed. Wild-type (N2) and 4 radiation-sensitive (rad) mutant dauer larvae were tested for their abilities to develop into adults after UV-irradiation. The rad-3 mutant was over 30 times as sensitive as N2; rad-1, rad-2 and rad-7 mutants were not hypersensitive. Irradiation also delayed development in survivors. Wild-type dauer larvae did not differ in radiation sensitivity from 0 through 50 days of age. There was no liquid holding recovery (LHR); that is, survival did not increase when wild-type dauer larvae were held in buffer after irradiation. (orig.). 28 refs.; 4 figs.

PBI creams: a spontaneously mutated mouse strain showing wild animal-type reactivity.

Science.gov (United States)

Hendrie, C A; Van Driel, K S; Talling, J C; Inglis, I R

2001-01-01

PBI creams are mice derived from warfarin-resistant wild stock that has been maintained under laboratory conditions since the 1970s. This study compares their behaviour to that of laboratory mice and wild house and wood mice. Animals were tested in a black/white box and a 2.64x1.4 m runway. In the black/white box, the behaviour of PBI creams was not significantly different from that of house mice and differed most from that of laboratory mice. Notably, the PBI creams showed the greatest activity and escape-orientated behaviours. When animals were approached by the experimenter in the open runway test, the PBI creams had higher flight speeds than both house and wood mice, whilst laboratory mice failed to respond. In the closed runway test where the animals could not escape, the PBI creams, house mice and wood mice all turned and attempted to run past the approaching experimenter, whilst the laboratory mice again failed to react. At the end of this test session, the time taken to catch each animal was recorded. It took less than 5 s to catch laboratory mice but significantly longer to catch the wild strains and the PBI creams (90-100 s for the latter). In these tests, the PBI creams showed wild animal-type reactivity, and as this behaviour has been retained in the laboratory colony for over 30 years, these animals may be useful in the study of the physiological and genetic basis of fear/anxiety in mice.

Genotyping assay for differentiation of wild-type and vaccine viruses in subjects immunized with live attenuated influenza vaccine.

Directory of Open Access Journals (Sweden)

Victoria Matyushenko

Full Text Available Live attenuated influenza vaccines (LAIVs are considered as safe and effective tool to control influenza in different age groups, especially in young children. An important part of the LAIV safety evaluation is the detection of vaccine virus replication in the nasopharynx of the vaccinees, with special attention to a potential virus transmission to the unvaccinated close contacts. Conducting LAIV clinical trials in some geographical regions with year-round circulation of influenza viruses warrants the development of robust and reliable tools for differentiating vaccine viruses from wild-type influenza viruses in nasal pharyngeal wash (NPW specimens of vaccinated subjects. Here we report the development of genotyping assay for the detection of wild-type and vaccine-type influenza virus genes in NPW specimens of young children immunized with Russian-backbone seasonal trivalent LAIV using Sanger sequencing from newly designed universal primers. The new primer set allowed amplification and sequencing of short fragments of viral genes in NPW specimens and appeared to be more sensitive than conventional real-time RT-PCR protocols routinely used for the detection and typing/subtyping of influenza virus in humans. Furthermore, the new assay is capable of defining the origin of wild-type influenza virus through BLAST search with the generated sequences of viral genes fragments.

The Genetic Structure of Wild Orobanche cumana Wallr. (Orobanchaceae Populations in Eastern Bulgaria Reflects Introgressions from Weedy Populations

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Rocío Pineda-Martos

2014-01-01

Full Text Available Orobanche cumana is a holoparasitic plant naturally distributed from central Asia to south-eastern Europe, where it parasitizes wild Asteraceae species. It is also an important parasitic weed of sunflower crops. The objective of this research was to investigate genetic diversity, population structure, and virulence on sunflower of O. cumana populations parasitizing wild plants in eastern Bulgaria. Fresh tissue of eight O. cumana populations and mature seeds of four of them were collected in situ on wild hosts. Genetic diversity and population structure were studied with SSR markers and compared to weedy populations. Two main gene pools were identified in Bulgarian populations, with most of the populations having intermediate characteristics. Cross-inoculation experiments revealed that O. cumana populations collected on wild species possessed similar ability to parasitize sunflower to those collected on sunflower. The results were explained on the basis of an effective genetic exchange between populations parasitizing sunflower crops and those parasitizing wild species. The occurrence of bidirectional gene flow may have an impact on wild populations, as new physiological races continuously emerge in weedy populations. Also, genetic variability of wild populations may favour the ability of weedy populations to overcome sunflower resistance mechanisms.

The Genetic Structure of Wild Orobanche cumana Wallr. (Orobanchaceae) Populations in Eastern Bulgaria Reflects Introgressions from Weedy Populations

Science.gov (United States)

Pineda-Martos, Rocío; Pujadas-Salvà, Antonio J.; Fernández-Martínez, José M.; Stoyanov, Kiril; Pérez-Vich, Begoña

2014-01-01

Orobanche cumana is a holoparasitic plant naturally distributed from central Asia to south-eastern Europe, where it parasitizes wild Asteraceae species. It is also an important parasitic weed of sunflower crops. The objective of this research was to investigate genetic diversity, population structure, and virulence on sunflower of O. cumana populations parasitizing wild plants in eastern Bulgaria. Fresh tissue of eight O. cumana populations and mature seeds of four of them were collected in situ on wild hosts. Genetic diversity and population structure were studied with SSR markers and compared to weedy populations. Two main gene pools were identified in Bulgarian populations, with most of the populations having intermediate characteristics. Cross-inoculation experiments revealed that O. cumana populations collected on wild species possessed similar ability to parasitize sunflower to those collected on sunflower. The results were explained on the basis of an effective genetic exchange between populations parasitizing sunflower crops and those parasitizing wild species. The occurrence of bidirectional gene flow may have an impact on wild populations, as new physiological races continuously emerge in weedy populations. Also, genetic variability of wild populations may favour the ability of weedy populations to overcome sunflower resistance mechanisms. PMID:25143963

Unique Safety Issues Associated with Virus Vectored Vaccines: Potential for and Theoretical Consequences of Recombination with Wild Type Virus Strains

Science.gov (United States)

Condit, Richard C.; Williamson, Anna-Lise; Sheets, Rebecca; Seligman, Stephen J.; Monath, Thomas P.; Excler, Jean-Louis; Gurwith, Marc; Bok, Karin; Robertson, James S.; Kim, Denny; Hendry, Michael; Singh, Vidisha; Mac, Lisa M.; Chen, Robert T.

2016-01-01

In 2003 and 2013, the World Health Organization convened informal consultations on characterization and quality aspects of vaccines based on live virus vectors. In the resulting reports, one of several issues raised for future study was the potential for recombination of virus-vectored vaccines with wild type pathogenic virus strains. This paper presents an assessment of this issue formulated by the Brighton Collaboration. To provide an appropriate context for understanding the potential for recombination of virus-vectored vaccines, we review briefly the current status of virus vectored vaccines, mechanisms of recombination between viruses, experience with recombination involving live attenuated vaccines in the field, and concerns raised previously in the literature regarding recombination of virus-vectored vaccines with wild type virus strains. We then present a discussion of the major variables that could influence recombination between a virus-vectored vaccine and circulating wild type virus and the consequences of such recombination, including intrinsic recombination properties of the parent virus used as a vector; sequence relatedness of vector and wild virus; virus host range, pathogenesis and transmission; replication competency of vector in target host; mechanism of vector attenuation; additional factors potentially affecting virulence; and circulation of multiple recombinant vectors in the same target population. Finally, we present some guiding principles for vector design and testing intended to anticipate and mitigate the potential for and consequences of recombination of virus-vectored vaccines with wild type pathogenic virus strains. PMID:27346303

Exploratory biomarker analysis for treatment response in KRAS wild type metastatic colorectal cancer patients who received cetuximab plus irinotecan

International Nuclear Information System (INIS)

Kim, Seung Tae; Ahn, Tae Jin; Lee, Eunjin; Do, In-Gu; Lee, Su Jin; Park, Se Hoon; Park, Joon Oh; Park, Young Suk; Lim, Ho Yeong; Kang, Won Ki; Kim, Suk Hyeong; Lee, Jeeyun; Kim, Hee Cheol

2015-01-01

More than half of the patients selected based on KRAS mutation status fail to respond to the treatment with cetuximab in metastatic colorectal cancer (mCRC). We designed a study to identify additional biomarkers that could act as indicators for cetuximab treatment in mCRC. We investigated 58 tumor samples from wild type KRAS CRC patients treated with cetuximab plus irinotecan (CI). We conducted the genotyping for mutations in either BRAF or PIK3CA and profiled comprehensively the expression of 522 kinase genes. BRAF mutation was detected in 5.1 % (3/58) of patients. All 50 patients showed wild type PIK3CA. Gene expression patterns that categorized patients with or without the disease control to CI were compared by supervised classification analysis. PSKH1, TLK2 and PHKG2 were overexpressed significantly in patients with the disease control to IC. The higher expression value of PSKH1 (r = 0.462, p < 0.001) and TLK2 (r = 0.361, p = 0.005) had the significant correlation to prolonged PFS. The result of this work demonstrated that expression nature of kinase genes such as PSKH1, TLK2 and PHKG2 may be informative to predict the efficacy of CI in wild type KRAS CRC. Mutations in either BRAF or PIK3CA were rare subsets in wild type KRAS CRC

MLH1-deficient Colorectal Carcinoma With Wild-type BRAF and MLH1 Promoter Hypermethylation Harbor KRAS Mutations and Arise From Conventional Adenomas.

Science.gov (United States)

Farchoukh, Lama; Kuan, Shih-Fan; Dudley, Beth; Brand, Randall; Nikiforova, Marina; Pai, Reetesh K

2016-10-01

Between 10% and 15% of colorectal carcinomas demonstrate sporadic DNA mismatch-repair protein deficiency as a result of MLH1 promoter methylation and are thought to arise from sessile serrated adenomas, termed the serrated neoplasia pathway. Although the presence of the BRAF V600E mutation is indicative of a sporadic cancer, up to 30% to 50% of colorectal carcinomas with MLH1 promoter hypermethylation will lack a BRAF mutation. We report the clinicopathologic and molecular features of MLH1-deficient colorectal carcinoma with wild-type BRAF and MLH1 promoter hypermethylation (referred to as MLH1-hypermethylated BRAF wild-type colorectal carcinoma, n=36) in comparison with MLH1-deficient BRAF-mutated colorectal carcinoma (n=113) and Lynch syndrome-associated colorectal carcinoma (n=36). KRAS mutations were identified in 31% of MLH1-hypermethylated BRAF wild-type colorectal carcinomas compared with 0% of MLH1-deficient BRAF-mutated colorectal carcinomas and 37% of Lynch syndrome-associated colorectal carcinomas. When a precursor polyp was identified, MLH1-hypermethylated BRAF wild-type colorectal carcinomas arose from precursor polyps resembling conventional tubular/tubulovillous adenomas in contrast to MLH1-deficient BRAF-mutated colorectal carcinomas, which arose from precursor sessile serrated adenomas (PMLH1-hypermethylated BRAF wild-type colorectal carcinoma and MLH1-deficient BRAF-mutated colorectal carcinoma had a predilection for the right colon compared with Lynch syndrome-associated colorectal carcinoma (86% vs. 92% vs. 49%, P0.05). In conclusion, our results indicate that MLH1-hypermethylated BRAF wild-type colorectal carcinomas can harbor KRAS mutations and arise from precursor polyps resembling conventional tubular/tubulovillous adenomas.

Mineral phosphate solubilization by wild type and radiation induced mutants of pantoea dispersa and pantoea terrae

International Nuclear Information System (INIS)

Murugesan, Senthilkumar; Lee, Young Keun; Kim, Jung Hun

2009-01-01

Three mineral phosphate solubilizing (MPS) bacteria where isolated from rhizosphere soil samples of common bean and weed plants. 16S rDNA analysis indicated that the isolate P2 and P3 are closely related to Pantoea dispersa while isolate P4 is closely related to Pantoea terrae. Islates P2 and P3 recorded 381.60 μg ml -1 of tricalcium phosphate (TCP) solubilization respectively on 3 days incubation. Isolate P4 recorded the TCP solubilization of 215.85 μg ml -1 and the pH was dropped to 4.44 on 24 h incubation. Further incubation of P4 sharply decreased the available phosphorous to 28.94 μg ml -1 and pH level was raised to 6.32. Gamma radiation induced mutagenesis was carried out at LD 99 dose of the wild type strains. The total of 14 mutant clones with enhanced MPS activity and 4 clones with decreased activity were selected based on solubilization index (SI) and phosphate solubilization assay. Mutant P2-M1 recorded the highest P-solubilizing potential among any other wild or mutnat clones by releasing 504.21 μg ml -1 of phosphorous i.e. 35% higher than its wild type by the end of day 5. A comparative evaluation of TCP solubilization by wild type isolates of Pantoea and their mutants, led to select three MPS mutant clones such as P2-M1, P3-M2 and P3-M4 with a potential to release >471.67 μg ml 1 of phosphorous from TCP. These over expressing mutant clones are considered as suitable candidates for biofertilization

Fitness of crop-wild hybrid sunflower under competitive conditions: implications for crop-to-wild introgression.

Science.gov (United States)

Mercer, Kristin L; Emry, D Jason; Snow, Allison A; Kost, Matthew A; Pace, Brian A; Alexander, Helen M

2014-01-01

Understanding the likelihood and extent of introgression of novel alleles in hybrid zones requires comparison of lifetime fitness of parents and hybrid progeny. However, fitness differences among cross types can vary depending on biotic conditions, thereby influencing introgression patterns. Based on past work, we predicted that increased competition would enhance introgression between cultivated and wild sunflower (Helianthus annuus) by reducing fitness advantages of wild plants. To test this prediction, we established a factorial field experiment in Kansas, USA where we monitored the fitness of four cross types (Wild, F1, F2, and BCw hybrids) under different levels of interspecific and intraspecific competition. Intraspecific manipulations consisted both of density of competitors and of frequency of crop-wild hybrids. We recorded emergence of overwintered seeds, survival to reproduction, and numbers of seeds produced per reproductive plant. We also calculated two compound fitness measures: seeds produced per emerged seedling and seeds produced per planted seed. Cross type and intraspecific competition affected emergence and survival to reproduction, respectively. Further, cross type interacted with competitive treatments to influence all other fitness traits. More intense competition treatments, especially related to density of intraspecific competitors, repeatedly reduced the fitness advantage of wild plants when considering seeds produced per reproductive plant and per emerged seedling, and F2 plants often became indistinguishable from the wilds. Wild fitness remained superior when seedling emergence was also considered as part of fitness, but the fitness of F2 hybrids relative to wild plants more than quadrupled with the addition of interspecific competitors and high densities of intraspecific competitors. Meanwhile, contrary to prediction, lower hybrid frequency reduced wild fitness advantage. These results emphasize the importance of taking a full life cycle

Fitness of crop-wild hybrid sunflower under competitive conditions: implications for crop-to-wild introgression.

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Kristin L Mercer

Full Text Available Understanding the likelihood and extent of introgression of novel alleles in hybrid zones requires comparison of lifetime fitness of parents and hybrid progeny. However, fitness differences among cross types can vary depending on biotic conditions, thereby influencing introgression patterns. Based on past work, we predicted that increased competition would enhance introgression between cultivated and wild sunflower (Helianthus annuus by reducing fitness advantages of wild plants. To test this prediction, we established a factorial field experiment in Kansas, USA where we monitored the fitness of four cross types (Wild, F1, F2, and BCw hybrids under different levels of interspecific and intraspecific competition. Intraspecific manipulations consisted both of density of competitors and of frequency of crop-wild hybrids. We recorded emergence of overwintered seeds, survival to reproduction, and numbers of seeds produced per reproductive plant. We also calculated two compound fitness measures: seeds produced per emerged seedling and seeds produced per planted seed. Cross type and intraspecific competition affected emergence and survival to reproduction, respectively. Further, cross type interacted with competitive treatments to influence all other fitness traits. More intense competition treatments, especially related to density of intraspecific competitors, repeatedly reduced the fitness advantage of wild plants when considering seeds produced per reproductive plant and per emerged seedling, and F2 plants often became indistinguishable from the wilds. Wild fitness remained superior when seedling emergence was also considered as part of fitness, but the fitness of F2 hybrids relative to wild plants more than quadrupled with the addition of interspecific competitors and high densities of intraspecific competitors. Meanwhile, contrary to prediction, lower hybrid frequency reduced wild fitness advantage. These results emphasize the importance of taking

Fitness of Crop-Wild Hybrid Sunflower under Competitive Conditions: Implications for Crop-to-Wild Introgression

Science.gov (United States)

Mercer, Kristin L.; Emry, D. Jason; Snow, Allison A.; Kost, Matthew A.; Pace, Brian A.; Alexander, Helen M.

2014-01-01

Understanding the likelihood and extent of introgression of novel alleles in hybrid zones requires comparison of lifetime fitness of parents and hybrid progeny. However, fitness differences among cross types can vary depending on biotic conditions, thereby influencing introgression patterns. Based on past work, we predicted that increased competition would enhance introgression between cultivated and wild sunflower (Helianthus annuus) by reducing fitness advantages of wild plants. To test this prediction, we established a factorial field experiment in Kansas, USA where we monitored the fitness of four cross types (Wild, F1, F2, and BCw hybrids) under different levels of interspecific and intraspecific competition. Intraspecific manipulations consisted both of density of competitors and of frequency of crop-wild hybrids. We recorded emergence of overwintered seeds, survival to reproduction, and numbers of seeds produced per reproductive plant. We also calculated two compound fitness measures: seeds produced per emerged seedling and seeds produced per planted seed. Cross type and intraspecific competition affected emergence and survival to reproduction, respectively. Further, cross type interacted with competitive treatments to influence all other fitness traits. More intense competition treatments, especially related to density of intraspecific competitors, repeatedly reduced the fitness advantage of wild plants when considering seeds produced per reproductive plant and per emerged seedling, and F2 plants often became indistinguishable from the wilds. Wild fitness remained superior when seedling emergence was also considered as part of fitness, but the fitness of F2 hybrids relative to wild plants more than quadrupled with the addition of interspecific competitors and high densities of intraspecific competitors. Meanwhile, contrary to prediction, lower hybrid frequency reduced wild fitness advantage. These results emphasize the importance of taking a full life cycle

The JH2 domain and SH2-JH2 linker regulate JAK2 activity: A detailed kinetic analysis of wild type and V617F mutant kinase domains.

Science.gov (United States)

Sanz Sanz, Arturo; Niranjan, Yashavanthi; Hammarén, Henrik; Ungureanu, Daniela; Ruijtenbeek, Rob; Touw, Ivo P; Silvennoinen, Olli; Hilhorst, Riet

2014-10-01

JAK2 tyrosine kinase regulates many cellular functions. Its activity is controlled by the pseudokinase (JH2) domain by still poorly understood mechanisms. The V617F mutation in the pseudokinase domain activates JAK2 and causes myeloproliferative neoplasms. We conducted a detailed kinetic analysis of recombinant JAK2 tyrosine kinase domain (JH1) and wild-type and V617F tandem kinase (JH1JH2) domains using peptide microarrays to define the functions of the kinase domains. The results show that i) JAK2 follows a random Bi-Bi reaction mechanism ii) JH2 domain restrains the activity of the JH1 domain by reducing the affinity for ATP and ATP competitive inhibitors iii) V617F decreases affinity for ATP but increases catalytic activity compared to wild-type and iv) the SH2-JH2 linker region participates in controlling activity by reducing the affinity for ATP. Copyright © 2014 Elsevier B.V. All rights reserved.

Sustained miRNA-mediated Knockdown of Mutant AAT With Simultaneous Augmentation of Wild-type AAT Has Minimal Effect on Global Liver miRNA Profiles

OpenAIRE

2013-01-01

α-1 antitrypsin (AAT) deficiency can exhibit two pathologic states: a lung disease that is primarily due to the loss of AAT's antiprotease function, and a liver disease resulting from a toxic gain-of-function of the PiZ-AAT (Z-AAT) mutant protein. We have developed several recombinant adeno-associated virus (rAAV) vectors that incorporate microRNA (miRNA) sequences targeting the AAT gene while also driving the expression of miRNA-resistant wild-type AAT-PiM (M-AAT) gene, thus achieving concom...

Component analysis of cultivated ginseng, cultivated wild ginseng, and natural wild ginseng by structural parts using HPLC method

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Young-Ju,Han

2007-02-01

Full Text Available Objectives : The aim of this experiments is to provide an objective differentiation of ginseng, Korean and Chinese cultivated wild ginseng, and natural wild ginseng through components analysis of different parts of ginseng. Methods : Comparative analyses of ginsenoside-, ginsenoside-, and ginsenosides and from the root, stem, and leaves of ginseng, Korean and Chinese cultivated wild ginseng, and natural wild ginseng were conducted using HPLC. Results : 1. For content comparison of leaves, ginseng showed highest content of ginsenoside than other samples. Natural wild ginseng showed relatively high content of ginsenosides and than other samples. 2. For content comparison of the stem, ginseng and 10 years old Chinese cultivated wild ginseng didn't contain ginsenoside . Natural wild ginseng showed higher content of ginsenosides and than other samples. 3. For content comparison of the root, ginsenoside was found only in 5 and 10 years old Korean cultivated wild ginseng. 4. Distribution of contents by the parts of ginseng was similar in ginseng and Chinese cultivated wild ginseng. Conclusions : Above experiment data can be an important indicator for the identification of ginseng, Korean and Chinese cultivated wild ginseng, and natural wild ginseng.

Staurosporine scaffold-based rational discovery of the wild-type sparing reversible inhibitors of EGFR T790M gatekeeper mutant in lung cancer with analog-sensitive kinase technology.

Science.gov (United States)

Song, Xiaoyun; Liu, Xingcai; Ding, Xi

2017-04-01

The human epidermal growth factor receptor (EGFR) has been established as an attractive target for lung cancer therapy. However, an acquired EGFR T790M gatekeeper mutation is frequently observed in patients treated with first-line anticancer agents such as gefitinib and erlotinib to cause drug resistance, largely limiting the application of small-molecule kinase inhibitors in EGFR-targeted chemotherapy. Previously, the reversible pan-kinase inhibitor staurosporine and its several analogs such as Gö6976 and K252a have been reported to selectively inhibit the EGFR T790M mutant (EGFR T790M ) over wild-type kinase (EGFR WT ), suggesting that the staurosporine scaffold is potentially to develop the wild-type sparing reversible inhibitors of EGFR T790M . Here, we systematically evaluated the inhibitor response of 28 staurosporine scaffold-based compounds to EGFR T790M mutation at structural, energetic, and molecular levels by using an integrated in silico-in vitro analog-sensitive (AS) kinase technology. With the strategy, we were able to identify 4 novel wild-type sparing inhibitors UCN-01, UCN-02, AFN941, and SB-218078 with high or moderate selectivity of 30-, 45-, 5-, and 8-fold for EGFR T790M over EGFR WT , respectively, which are comparable with or even better than that of the parent compound staurosporine (24-fold). Molecular modeling and structural analysis revealed that van der Waals contacts and hydrophobic forces can form between the side chain of mutated residue Met790 and the pyrrolidinone moiety of inhibitor ligand UCN-02, which may simultaneously improve the favorable interaction energy between the kinase and inhibitor, and reduce the unfavorable desolvation penalty upon the kinase-inhibitor binding. A hydroxyl group of UCN-02 additional to staurosporine locates at the pyrrolidinone moiety, which can largely alter the electronic distribution of pyrrolidinone moiety and thus promote the intermolecular interaction with Met790 residue. This can well explain

Chronic activation of wild-type epidermal growth factor receptor and loss of Cdkn2a cause mouse glioblastoma formation.

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Acquaviva, Jaime; Jun, Hyun Jung; Lessard, Julie; Ruiz, Rolando; Zhu, Haihao; Donovan, Melissa; Woolfenden, Steve; Boskovitz, Abraham; Raval, Ami; Bronson, Roderick T; Pfannl, Rolf; Whittaker, Charles A; Housman, David E; Charest, Al

2011-12-01

Glioblastoma multiforme (GBM) is characterized by overexpression of epidermal growth factor receptor (EGFR) and loss of the tumor suppressors Ink4a/Arf. Efforts at modeling GBM using wild-type EGFR in mice have proven unsuccessful. Here, we present a unique mouse model of wild-type EGFR-driven gliomagenesis. We used a combination of somatic conditional overexpression and ligand-mediated chronic activation of EGFR in cooperation with Ink4a/Arf loss in the central nervous system of adult mice to generate tumors with the histopathologic and molecular characteristics of human GBMs. Sustained, ligand-mediated activation of EGFR was necessary for gliomagenesis, functionally substantiating the clinical observation that EGFR-positive GBMs from patients express EGFR ligands. To gain a better understanding of the clinically disappointing EGFR-targeted therapies for GBM, we investigated the molecular responses to EGFR tyrosine kinase inhibitor (TKI) treatment in this model. Gefitinib treatment of primary GBM cells resulted in a robust apoptotic response, partially conveyed by mitogen-activated protein kinase (MAPK) signaling attenuation and accompanied by BIM(EL) expression. In human GBMs, loss-of-function mutations in the tumor suppressor PTEN are a common occurrence. Elimination of PTEN expression in GBM cells posttumor formation did not confer resistance to TKI treatment, showing that PTEN status in our model is not predictive. Together, these findings offer important mechanistic insights into the genetic determinants of EGFR gliomagenesis and sensitivity to TKIs and provide a robust discovery platform to better understand the molecular events that are associated with predictive markers of TKI therapy.

Genetic diversity and structure analysis in wild and landraces of barley from Jordan by using ISJ markers

International Nuclear Information System (INIS)

Baloch, A. W.; Balogh, M. J.; Baloch, M.; Baloch, I. A.

2016-01-01

The present experiment was carried out to estimate genetic diversity and genetic structure in cultivated and wild barley populations collected from Jordan which is considered as primary gene pool of barley. In a total, 94 cultivated barley accessions composed of 4 populations and 52 wild barley accessions consisted of 3 populations were used for genetic analysis using 7 Intron Splice Junction (ISJ) markers. The genetic diversity index (He) of cultivated barley ranged between 0.049 and 0.060; whereas that of wild barley populations ranged between 0.084 and 0.146, suggesting that wild resources of barley harbored greater genetic diversity than its domesticated counterpart, reflecting that barley domestication occurred with genetic bottleneck. Analysis of molecular variance showed high genetic variations among rather than within populations, referring that high genetic differentiation of barley populations caused by genetic and geographical separation of the populations in the harsh growing conditions of Fertile Crescent. Principal coordinate, clustering and structure analysis not only separated cultivated and wild barley, but also each single population, showing their genetic basis and original sample site. The obtained Results also revealed that there is lesser genetic communication between cultivated and wild barley under natural environments. The current findings can better be exploited for collection and utilization of plant germplasms. (author)

Identification of Potential Wild Herbal as parts of Landscape Elements

Science.gov (United States)

Sulistyantara, Bambang; Mentari, Nio

2017-10-01

Many landscape plants can grow on their own without cultivated by humans. They are type of plants that can be found anywhere, so they can be categorized as wild plants. The economic value of wild plants are easy to obtain and their maintenance costs are low. Because wild plants not widely known even a just a few of people that aware of their existence, it is necessary to do a study to learn the potential of the wild plants to be used as an element of landscape. This research aims to identify the species that have potential to be used in landscape design, to describe the benefits of the their implementation as a landscape element, and to recommend the wild plants that have functional value and visual. This research used a scoring method based on the functional and visual criteria, and questionnaires were conducted to 50 students of Landscape Architecture IPB who have completed Landscape Plants courses. Based on the research, there are 150 species of wild plants that found in the study site, and 60 of them are recommended as landscape elements. Then all of the species were arranged as a recommendations book so they can be used as alternative landscape plants.

Clavulanic acid production by the MMS 150 mutant obtained from wild type Streptomyces clavuligerus ATCC 27064

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Eliton da Silva Vasconcelos

2013-12-01

Full Text Available Clavulanic acid (CA is a powerful inhibitor of the beta-lactamases, enzymes produced by bacteria resistants to penicillin and cefalosporin. This molecule is produced industrially by strains of Streptomyces clavuligerus in complex media which carbon and nitrogen resources are supplied by inexpensive compounds still providing high productivity. The genetic production improvement using physical and chemical mutagenic agents is an important strategy in programs of industrial production development of bioactive metabolites. However, parental strains are susceptible to loss of their original productivity due genetic instability phenomenona. In this work, some S. clavuligerus mutant strains obtained by treatment with UV light and with MMS are compared with the wild type (Streptomyces clavuligerus ATCC 27064. The results indicated that the random mutations originated some strains with different phenotypes, most divergent demonstrated by the mutants strains named AC116, MMS 150 and MMS 54, that exhibited lack of pigmentation in their mature spores. Also, the strain MMS 150 presented a larger production of CA when cultivated in semi-synthetics media. Using other media, the wild type strain obtained a larger CA production. Besides, using the modifed complex media the MMS 150 strain showed changes in its lipolitic activity and a larger production of CA. The studies also allowed finding the best conditions for a lipase activity exhibited by wild type S. clavuligerus and the MMS150 mutant.

Clavulanic acid production by the MMS 150 mutant obtained from wild type Streptomyces clavuligerus ATCC 27064.

Science.gov (United States)

da Silva Vasconcelos, Eliton; de Lima, Vanderlei Aparecido; Goto, Leandro Seiji; Cruz-Hernández, Isara Lourdes; Hokka, Carlos Osamu

2013-12-01

Clavulanic acid (CA) is a powerful inhibitor of the beta-lactamases, enzymes produced by bacteria resistants to penicillin and cefalosporin. This molecule is produced industrially by strains of Streptomyces clavuligerus in complex media which carbon and nitrogen resources are supplied by inexpensive compounds still providing high productivity. The genetic production improvement using physical and chemical mutagenic agents is an important strategy in programs of industrial production development of bioactive metabolites. However, parental strains are susceptible to loss of their original productivity due genetic instability phenomenona. In this work, some S. clavuligerus mutant strains obtained by treatment with UV light and with MMS are compared with the wild type (Streptomyces clavuligerus ATCC 27064). The results indicated that the random mutations originated some strains with different phenotypes, most divergent demonstrated by the mutants strains named AC116, MMS 150 and MMS 54, that exhibited lack of pigmentation in their mature spores. Also, the strain MMS 150 presented a larger production of CA when cultivated in semi-synthetics media. Using other media, the wild type strain obtained a larger CA production. Besides, using the modifed complex media the MMS 150 strain showed changes in its lipolitic activity and a larger production of CA. The studies also allowed finding the best conditions for a lipase activity exhibited by wild type S. clavuligerus and the MMS150 mutant.

Craniofacial Statistical Deformation Models of Wild-type mice and Crouzon mice

DEFF Research Database (Denmark)

Ólafsdóttir, Hildur; Darvann, Tron Andre; Ersbøll, Bjarne Kjær

2007-01-01

Crouzon syndrome is characterised by the premature fusion of cranial sutures and synchondroses leading to craniofacial growth disturbances. The gene causing the syndrome was discovered approximately a decade ago and recently the first mouse model of the syndrome was generated. In this study, a set...... of Micro CT scannings of the heads of wild-type (normal) mice and Crouzon mice were investigated. Statistical deformation models were built to assess the anatomical differences between the groups, as well as the within-group anatomical variation. Following the approach by Rueckert et al. we built an atlas...

Phosphorylated EGFR expression may predict outcome of EGFR-TKIs therapy for the advanced NSCLC patients with wild-type EGFR

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Wang Fen

2012-08-01

Full Text Available Abstract Background EGFR mutation is a strong predictive factor of EGFR-TKIs therapy. However, at least 10% of patients with EGFR wild-type are responsive to TKIs, suggesting that other determinants of outcome besides EGFR mutation might exist. We hypothesized that activation of phosphorylated EGFR could be a potential predictive biomarker to EGFR-TKIs treatment among patients in wild-type EGFR. Method Total of 205 stage IIIb and IV NSCLC patients, tissue samples of whom were available for molecular analysis, were enrolled in this study. The phosphorylation of EGFR at tyrosine 1068 (pTyr1068 and 1173 (pTyr1173 were assessed by immunohistochemistry, and EGFR mutations were detected by denaturing high performance liquid chromatograph (DHPLC. Results Among 205 patients assessable for EGFR mutation and phosphorylation analysis, 92 (44.9% were EGFR mutant and 165 patients (57.6% had pTyr1173 expression. Superior progression-free survival (PFS was seen after EGFR-TKIs therapy in patients with pTyr1068 expression compared to pTyr1068 negative ones (median PFS 7.0 months vs. 1.2 months, P P = 0.016. In subgroup of patients with wild-type EGFR, pTyr1068 expression positive ones had a significantly prolonged PFS (4.2 months vs.1.2 months P  Conclusion pTyr1068 may be a predictive biomarker for screening the population for clinical response to EGFR-TKIs treatment; especially for patients with wild-type EGFR.

Differentiation between probiotic and wild-type Bacillus cereus isolates by antibiotic susceptibility test and Fourier transform infrared spectroscopy (FT-IR).

Science.gov (United States)

Mietke, Henriette; Beer, W; Schleif, Julia; Schabert, G; Reissbrodt, R

2010-05-30

Animal feed often contains probiotic Bacillus strains used as feed additives. Spores of the non-pathogenic B. cereus var. toyoi (product name Toyocerin) are used. Distinguishing between toxic wild-type Bacillus cereus strains and this probiotic strain is essential for evaluating the quality and risk of feed. Bacillus cereus CIP 5832 (product name Paciflor was used as probiotic strain until 2001. The properties of the two probiotic strains are quite similar. Differentiating between probiotic strains and wild-type B. cereus strains is not easy. ss-lactam antibiotics such as penicillin and cefamandole exhibit an inhibition zone in the agar diffusion test of probiotic B. cereus strains which are not seen for wild-type strains. Therefore, performing the agar diffusion test first may make sense before FT-IR testing. When randomly checking these strains by Fourier transform infrared spectroscopy (FT-IR), the probiotic B. cereus strains were separated from wild-type B. cereus/B. thuringiensis/B. mycoides/B. weihenstephanensis strains by means of hierarchical cluster analysis. The discriminatory information was contained in the spectral windows 3000-2800 cm(-1) ("fatty acid region"), 1200-900 cm(-1) ("carbohydrate region") and 900-700 cm(-1) ("fingerprint region"). It is concluded that FT-IR spectroscopy can be used for the rapid quality control and risk analysis of animal feed containing probiotic B. cereus strains. (c) 2010 Elsevier B.V. All rights reserved.

Glucocorticoid-regulated and constitutive trafficking of proteolytically processed cell surface-associated glycoproteins in wild type and variant rat hepatoma cells

International Nuclear Information System (INIS)

Amacher, S.L.; Goodman, L.J.; Bravo, D.A.; Wong, K.Y.; Goldfine, I.D.; Hawley, D.M.; Firestone, G.L.

1989-01-01

Glucocorticoids regulate the trafficking of mouse mammary tumor virus (MMTV) glycoproteins to the cell surface in the rat hepatoma cell line M1.54, but not in the immunoselected sorting variant CR4. To compare the localization of MMTV glycoproteins to another proteolytically processed glycoprotein, both wild type M1.54 cells and variant CR4 cells were transfected with a human insulin receptor (hIR) expression vector, pRSVhIR. The production of cell surface hIR was monitored in dexamethasone-treated and -untreated wild type M1.54 and variant CR4 cells by indirect immunofluorescence, direct plasma membrane immunoprecipitation, and by [125I] insulin binding. In both wild type and variant rat hepatoma cells, hIR were localized at the cell surface in the presence or in the absence of 1 microM dexamethasone. In contrast, the glucocorticoid-regulated trafficking of cell surface MMTV glycoproteins occurred only in wild type M1.54 cells. We conclude that the hIR, which undergoes posttranslational processing reactions similar to MMTV glycoproteins, does not require glucocorticoids to be transported to the plasma membrane and is representative of a subset of cell surface glycoproteins whose trafficking is constitutive in rat hepatoma cells. Thus, MMTV glycoproteins and hIR provide specific cell surface markers to characterize the glucocorticoid-regulated and constitutive sorting pathways

Spatiotemporal trends in Canadian domestic wild boar production and habitat predict wild pig distribution

DEFF Research Database (Denmark)

Michel, Nicole; Laforge, Michel; van Beest, Floris

2017-01-01

eradication of wild pigs is rarely feasible after establishment over large areas, effective management will depend on strengthening regulations and enforcement of containment practices for Canadian domestic wild boar farms. Initiation of coordinated provincial and federal efforts to implement population...... wild boar and test the propagule pressure hypothesis to improve predictive ability of an existing habitat-based model of wild pigs. We reviewed spatiotemporal patterns in domestic wild boar production across ten Canadian provinces during 1991–2011 and evaluated the ability of wild boar farm...... distribution to improve predictive models of wild pig occurrence using a resource selection probability function for wild pigs in Saskatchewan. Domestic wild boar production in Canada increased from 1991 to 2001 followed by sharp declines in all provinces. The distribution of domestic wild boar farms in 2006...

Growth hormone receptor deficiency in mice results in reduced systolic blood pressure and plasma renin, increased aortic eNOS expression, and altered cardiovascular structure and function

DEFF Research Database (Denmark)

Egecioglu, E; Andersson, I J; Bollano, E

2007-01-01

To study the role of the growth hormone receptor (GHR) in the development of cardiovascular structure and function, female GHR gene-disrupted or knockout (KO) and wild-type (WT) mice at age 18 wk were used. GHR KO mice had lower plasma renin levels (12 +/- 2 vs. 20 +/- 4 mGU/ml, P < 0.05) and inc....... These data suggest an important role for an intact GH/IGF-I axis in the maintenance of a normal cardiovascular system....

Hierarchical structure of the otolith of adult wild carp

International Nuclear Information System (INIS)

Li Zhuo; Gao Yonghua; Feng Qingling

2009-01-01

The otolith of adult wild carp contains a pair of asterisci, a pair of lappilli and a pair of sagittae. Current research works are mainly restricted to the field of the daily ring structure. The purpose of this work is to explore the structural characteristics of carp's otolith in terms of hierarchy from nanometer to millimeter scale by transmission election microscope (TEM) and scanning electron microscope (SEM). Based on the observation, carp's lapillus is composed of ordered aragonite crystals. Seven hierarchical levels of the microstructure were proposed and described with the scheme representing a complete organization in detail. SEM studies show not only the clear daily growth increment, but also the morphology within the single daily increment. The domain structure of crystal orientation in otolith was observed for the first time. Furthermore, TEM investigation displays that the lapillus is composed of aragonite crystals with nanometer scale. Four hierarchical levels of the microstructure of the sagitta are also proposed. The asteriscus which is composed of nanometer scale vaterite crystals is considered to have a uniform structure.

Hierarchical structure of the otolith of adult wild carp

Energy Technology Data Exchange (ETDEWEB)

Li Zhuo; Gao Yonghua [State key laboratory of new ceramics and fine processing, Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Feng Qingling, E-mail: biomater@mail.tsinghua.edu.cn [State key laboratory of new ceramics and fine processing, Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China)

2009-04-30

The otolith of adult wild carp contains a pair of asterisci, a pair of lappilli and a pair of sagittae. Current research works are mainly restricted to the field of the daily ring structure. The purpose of this work is to explore the structural characteristics of carp's otolith in terms of hierarchy from nanometer to millimeter scale by transmission election microscope (TEM) and scanning electron microscope (SEM). Based on the observation, carp's lapillus is composed of ordered aragonite crystals. Seven hierarchical levels of the microstructure were proposed and described with the scheme representing a complete organization in detail. SEM studies show not only the clear daily growth increment, but also the morphology within the single daily increment. The domain structure of crystal orientation in otolith was observed for the first time. Furthermore, TEM investigation displays that the lapillus is composed of aragonite crystals with nanometer scale. Four hierarchical levels of the microstructure of the sagitta are also proposed. The asteriscus which is composed of nanometer scale vaterite crystals is considered to have a uniform structure.

Genome Sequences of Three Vaccine Strains and Two Wild-Type Canine Distemper Virus Strains from a Recent Disease Outbreak in South Africa.

Science.gov (United States)

Loots, Angelika K; Du Plessis, Morné; Dalton, Desiré Lee; Mitchell, Emily; Venter, Estelle H

2017-07-06

Canine distemper virus causes global multihost infectious disease. This report details complete genome sequences of three vaccine and two new wild-type strains. The wild-type strains belong to the South African lineage, and all three vaccine strains to the America 1 lineage. This constitutes the first genomic sequences of this virus from South Africa. Copyright © 2017 Loots et al.

Identification of concomitant infection with Chlamydia trachomatis IncA-negative mutant and wild-type strains by genomic, transcriptional, and biological characterizations.

Science.gov (United States)

Suchland, Robert J; Jeffrey, Brendan M; Xia, Minsheng; Bhatia, Ajay; Chu, Hencelyn G; Rockey, Daniel D; Stamm, Walter E

2008-12-01

Clinical isolates of Chlamydia trachomatis that lack IncA on their inclusion membrane form nonfusogenic inclusions and have been associated with milder, subclinical infections in patients. The molecular events associated with the generation of IncA-negative strains and their roles in chlamydial sexually transmitted infections are not clear. We explored the biology of the IncA-negative strains by analyzing their genomic structure, transcription, and growth characteristics in vitro and in vivo in comparison with IncA-positive C. trachomatis strains. Three clinical samples were identified that contained a mixture of IncA-positive and -negative same-serovar C. trachomatis populations, and two more such pairs were found in serial isolates from persistently infected individuals. Genomic sequence analysis of individual strains from each of two serovar-matched pairs showed that these pairs were very similar genetically. In contrast, the genome sequence of an unmatched IncA-negative strain contained over 5,000 nucleotide polymorphisms relative to the genome sequence of a serovar-matched but otherwise unlinked strain. Transcriptional analysis, in vitro culture kinetics, and animal modeling demonstrated that IncA-negative strains isolated in the presence of a serovar-matched wild-type strain are phenotypically more similar to the wild-type strain than are IncA-negative strains isolated in the absence of a serovar-matched wild-type strain. These studies support a model suggesting that a change from an IncA-positive strain to the previously described IncA-negative phenotype may involve multiple steps, the first of which involves a translational inactivation of incA, associated with subsequent unidentified steps that lead to the observed decrease in transcript level, differences in growth rate, and differences in mouse infectivity.

Characterization of two second-site mutations preventing wild type protein aggregation caused by a dominant negative PMA1 mutant.

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Pilar Eraso

Full Text Available The correct biogenesis and localization of Pma1 at the plasma membrane is essential for yeast growth. A subset of PMA1 mutations behave as dominant negative because they produce aberrantly folded proteins that form protein aggregates, which in turn provoke the aggregation of the wild type protein. One approach to understand this dominant negative effect is to identify second-site mutations able to suppress the dominant lethal phenotype caused by those mutant alleles. We isolated and characterized two intragenic second-site suppressors of the PMA1-D378T dominant negative mutation. We present here the analysis of these new mutations that are located along the amino-terminal half of the protein and include a missense mutation, L151F, and an in-frame 12bp deletion that eliminates four residues from Cys409 to Ala412. The results show that the suppressor mutations disrupt the interaction between the mutant and wild type enzymes, and this enables the wild type Pma1 to reach the plasma membrane.

Detection and differentiation of wild-type and vaccine strains of canine distemper virus by a duplex reverse transcription polymerase chain reaction.

Science.gov (United States)

Dong, X Y; Li, W H; Zhu, J L; Liu, W J; Zhao, M Q; Luo, Y W; Chen, J D

2015-01-01

Canine distemper virus (CDV) is the cause of canine distemper (CD) which is a severe and highly contagious disease in dogs. In the present study, a duplex reverse transcription polymerase chain reaction (RT-PCR) method was developed for the detection and differentiation of wild-type and vaccine strains of CDV. Four primers were designed to detect and discriminate the two viruses by generating 638- and 781-bp cDNA products, respectively. Furthermore, the duplex RT-PCR method was used to detect 67 field samples suspected of CD from Guangdong province in China. Results showed that, 33 samples were to be wild-type-like. The duplex RT-PCR method exhibited high specificity and sensitivity which could be used to effectively detect and differentiate wild-type and vaccine CDV, indicating its use for clinical detection and epidemiological surveillance.

Structural and functional salivary disorders in type 2 diabetic patients.

Science.gov (United States)

Carda, Carmen; Mosquera-Lloreda, Nezly; Salom, Lucas; Gomez de Ferraris, Maria Elsa; Peydró, Amando

2006-07-01

Diabetes mellitus type 2 is the most common metabolic disorder and it causes an important morbimortality. The structural modifications in the parotid gland (sialosis) had already been described in these patients and could result in variations in the salivary composition, as well as an increase in periodontal and dental pathology. To compare the biochemical findings in the saliva and to correlate these biochemical disturbances with the morphologic findings previously described. Clinical information were gathered about 33 patients, 17 had type 2 diabetes. Samples of whole saliva were obtained for biochemical analysis and serum samples to determine metabolic control. In the diabetics saliva we found urea and total proteins increased and reduced levels of microalbumina. Salivary glucose was only augmented in patients with poor metabolic control. Clinical symptoms of xerostomia were present in 76,4% and dental and periodontal disease in 100%. The parotid gland was characterised by the presence of small acini, lipid intracytoplasmic droplets, as well as adipose stroma infiltration. The acinar cytoqueratins expression was heterogeneous and very positive in the hyperplasic ducts. These biochemical disorders in the saliva of the type 2 diabetic patients would be related with the structural changes previously observed in parotid glands.

Comparative Study on Growth Performance of Transgenic (Over-Expressed OsNHX1 and Wild-Type Nipponbare under Different Salinity Regimes

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Nurul Kahrani ISHAK

2015-11-01

Full Text Available Transgenic Nipponbare which over-expressed a Na+/H+ antiporter gene OsNHX1 was used to compare its growth performance, water status and photosynthetic efficiency with its wild type under varying salinity regimes. Chlorophyll content, quantum yield and photosynthetic rate were measured to assess the impact of salinity stress on photosynthetic efficiency for transgenic and wild-type Nipponbare. Effects of salinity on water status and gas exchange to both lines were studied by measuring water use efficiency, instantaneous transpiration rate and stomatal conductance. Dry shoot weight and leaf area were determined after three months of growth to assess the impacts of salinity on the growth of those two lines. Our study showed that both lines were affected by salinity stress, however, the transgenic line showed higher photosynthetic efficiency, better utilization of water, and better growth due to low transpiration rate and stomatal conductance. Reduction of photosynthetic efficiency exhibited by the wild-type Nipponbare was correlated to its poor growth under salinity stress.

Transcriptional regulatory program in wild-type and retinoblastoma gene-deficient mouse embryonic fibroblasts during adipocyte differentiation

DEFF Research Database (Denmark)

Hakim-Weber, Robab; Krogsdam, Anne-M; Jørgensen, Claus

2011-01-01

Although many molecular regulators of adipogenesis have been identified a comprehensive catalogue of components is still missing. Recent studies showed that the retinoblastoma protein (pRb) was expressed in the cell cycle and late cellular differentiation phase during adipogenesis. To investigate...... this dual role of pRb in the early and late stages of adipogenesis we used microarrays to perform a comprehensive systems-level analysis of the common transcriptional program of the classic 3T3-L1 preadipocyte cell line, wild-type mouse embryonic fibroblasts (MEFs), and retinoblastoma gene-deficient MEFs...... of experimental data and computational analyses pinpointed a feedback-loop between Pparg and Foxo1.To analyze the effects of the retinoblastoma protein at the transcriptional level we chose a perturbated system (Rb-/- MEFs) for comparison to the transcriptional program of wild-type MEFs. Gene ontology analysis...

Discovery of an inhibitor of the production of the Pseudomonas aeruginosa virulence factor pyocyanin in wild-type cells

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Bernardas Morkunas

2016-07-01

Full Text Available Pyocyanin is a small molecule produced by Pseudomonas aeruginosa that plays a crucial role in the pathogenesis of infections by this notorious opportunistic pathogen. The inhibition of pyocyanin production has been identified as an attractive antivirulence strategy for the treatment of P. aeruginosa infections. Herein, we report the discovery of an inhibitor of pyocyanin production in cultures of wild-type P. aeruginosa which is based around a 4-alkylquinolin-2(1H-one scaffold. To the best of our knowledge, this is the first reported example of pyocyanin inhibition by a compound based around this molecular framework. The compound may therefore be representative of a new structural sub-class of pyocyanin inhibitors, which could potentially be exploited in in a therapeutic context for the development of critically needed new antipseudomonal agents. In this context, the use of wild-type cells in this study is notable, since the data obtained are of direct relevance to native situations. The compound could also be of value in better elucidating the role of pyocyanin in P. aeruginosa infections. Evidence suggests that the active compound reduces the level of pyocyanin production by inhibiting the cell–cell signalling mechanism known as quorum sensing. This could have interesting implications; quorum sensing regulates a range of additional elements associated with the pathogenicity of P. aeruginosa and there is a wide range of other potential applications where the inhibition of quorum sensing is desirable.

Anti-tumor activity of high-dose EGFR tyrosine kinase inhibitor and sequential docetaxel in wild type EGFR non-small cell lung cancer cell nude mouse xenografts

OpenAIRE

Tang, Ning; Zhang, Qianqian; Fang, Shu; Han, Xiao; Wang, Zhehai

2016-01-01

Treatment of non-small-cell lung cancer (NSCLC) with wild-type epidermal growth factor receptor (EGFR) is still a challenge. This study explored antitumor activity of high-dose icotinib (an EGFR tyrosine kinase inhibitor) plus sequential docetaxel against wild-type EGFR NSCLC cells-generated nude mouse xenografts. Nude mice were subcutaneously injected with wild-type EGFR NSCLC A549 cells and divided into different groups for 3-week treatment. Tumor xenograft volumes were monitored and record...

Diversity Partitioning of Wild Bee Assemblages (Hymenoptera: Apoidea, Apiformes and Species Preferences for Three Types of Refuge Habitats in an Agricultural Landscape in Poland

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Banaszak Józef

2014-09-01

Full Text Available Abstract Patterns in bee assemblages consisting of 52 core (most abundant species in farmland in the Wielkopolska region of W Poland were analysed. The entomological material was assessed during earlier research in 1978-1993 from 18 plots in three habitat types: shelterbelts, roadsides and forest patches. At the scale of the refuge habitat size analysed here, an increase in area only slightly enhanced bee species richness. The bee assemblage structures of roadsides and forest patches differ significantly, but their indicator species do not form any well-defined ecological groups. In non-linear forest patches, the bee community structure was more homogeneous than on roadsides. These two habitat types differed significantly in their species composition. Nine significant indicator species were found, but they did not share any ecological characteristics. Three factors were found to affect significantly the responses of individual bee species in the agricultural landscape: the degree of isolation of the refuge habitat, the edge ratio, and roadsides as a refuge habitat type. A large part of the regional diversity is due to the heterogeneity of habitats within the landscape. Habitat area has little influence on the diversity of wild bees, at least within the size range analysed here. We concluded from this study that, regardless of the habitat type, the density of bees from the summer phenological period is affected by the number of food plant species. Point forest patches are habitats where summer species from the genus Andrena and the cleptoparasitic genera Nomada and Sphecodes achieve their highest abundances. Roadsides negatively affected abundances of wild bees and there were no characteristic species for this type of habitat. We hypothesised that this might be related to the specific ecological part played by this type of habitat.

Annotating and quantifying pri-miRNA transcripts using RNA-Seq data of wild type and serrate-1 globular stage embryos of Arabidopsis thaliana

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Daniel Lepe-Soltero

2017-12-01

Full Text Available The genome annotation for the model plant Arabidopsis thaliana does not include the primary transcripts from which MIRNAs are processed. Here we present and analyze the raw mRNA sequencing data from wild type and serrate-1 globular stage embryos of A. thaliana, ecotype Columbia. Because SERRATE is required for pri-miRNA processing, these precursors accumulate in serrate-1 mutants, facilitating their detection using standard RNA-Seq protocols. We first use the mapping of the RNA-Seq reads to the reference genome to annotate the potential primary transcripts of MIRNAs expressed in the embryo. We then quantify these pri-miRNAs in wild type and serrate-1 mutants. Finally, we use differential expression analysis to determine which are up-regulated in serrate-1 compared to wild type, to select the best candidates for bona fide pri-miRNAs expressed in the globular stage embryos. In addition, we analyze a previously published RNA-Seq dataset of wild type and dicer-like 1 mutant embryos at the globular stage [1]. Our data are interpreted and discussed in a separate article [2].

Annotating and quantifying pri-miRNA transcripts using RNA-Seq data of wild type and serrate-1 globular stage embryos of Arabidopsis thaliana.

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Lepe-Soltero, Daniel; Armenta-Medina, Alma; Xiang, Daoquan; Datla, Raju; Gillmor, C Stewart; Abreu-Goodger, Cei

2017-12-01

The genome annotation for the model plant Arabidopsis thaliana does not include the primary transcripts from which MIRNAs are processed. Here we present and analyze the raw mRNA sequencing data from wild type and serrate-1 globular stage embryos of A. thaliana , ecotype Columbia. Because SERRATE is required for pri-miRNA processing, these precursors accumulate in serrate-1 mutants, facilitating their detection using standard RNA-Seq protocols. We first use the mapping of the RNA-Seq reads to the reference genome to annotate the potential primary transcripts of MIRNAs expressed in the embryo. We then quantify these pri-miRNAs in wild type and serrate-1 mutants. Finally, we use differential expression analysis to determine which are up-regulated in serrate-1 compared to wild type, to select the best candidates for bona fide pri-miRNAs expressed in the globular stage embryos. In addition, we analyze a previously published RNA-Seq dataset of wild type and dicer-like 1 mutant embryos at the globular stage [1]. Our data are interpreted and discussed in a separate article [2].

Genotype-temperature interaction in the regulation of development, growth, and morphometrics in wild-type, and growth-hormone transgenic coho salmon.

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Mare Lõhmus

2010-04-01

Full Text Available The neuroendocrine system is an important modulator of phenotype, directing cellular genetic responses to external cues such as temperature. Behavioural and physiological processes in poikilothermic organisms (e.g. most fishes, are particularly influenced by surrounding temperatures.By comparing the development and growth of two genotypes of coho salmon (wild-type and transgenic with greatly enhanced growth hormone production at six different temperatures, ranging between 8 degrees and 18 degrees C, we observed a genotype-temperature interaction and possible trend in directed neuroendocrine selection. Differences in growth patterns of the two genotypes were compared by using mathematical models, and morphometric analyses of juvenile salmon were performed to detect differences in body shape. The maximum hatching and alevin survival rates of both genotypes occurred at 12 degrees C. At lower temperatures, eggs containing embryos with enhanced GH production hatched after a shorter incubation period than wild-type eggs, but this difference was not apparent at and above 16 degrees C. GH transgenesis led to lower body weights at the time when the yolk sack was completely absorbed compared to the wild genotype. The growth of juvenile GH-enhanced salmon was to a greater extent stimulated by higher temperatures than the growth of the wild-type. Increased GH production significantly influenced the shape of the salmon growth curves.Growth hormone overexpression by transgenesis is able to stimulate the growth of coho salmon over a wide range of temperatures. Temperature was found to affect growth rate, survival, and body morphology between GH transgenic and wild genotype coho salmon, and differential responses to temperature observed between the genotypes suggests they would experience different selective forces should they ever enter natural ecosystems. Thus, GH transgenic fish would be expected to differentially respond and adapt to shifts in environmental

Attenuated, oncolytic, but not wild-type measles virus infection has pleiotropic effects on human neutrophil function.

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Zhang, Yu; Patel, Bella; Dey, Aditi; Ghorani, Ehsan; Rai, Lena; Elham, Mohammed; Castleton, Anna Z; Fielding, Adele K

2012-02-01

We previously showed that neutrophils play a role in regression of human tumor xenografts in immunodeficient mice following oncolytic vaccine measles virus (MV-Vac) treatment. In this study, we sought, using normal human neutrophils, to identify potential neutrophil-mediated mechanisms for the attenuated MV-Vac induced effects seen in vivo, by comparison with those consequent on wild-type (WT-MV) infection. Both MV-Vac and WT-MV infected and replicated within neutrophils, despite lack of SLAM expression. In both cases, neutrophils survived longer ex vivo postinfection. Furthermore, MV-Vac (but not WT-MV) infection activated neutrophils and stimulated secretion of several specific antitumor cytokines (IL-8, TNF-α, MCP-1, and IFN-α) via induction of de novo RNA and protein synthesis. In addition, MV-Vac (but not WT-MV) infection caused TRAIL secretion in the absence of de novo synthesis by triggering release of prefabricated TRAIL, via a direct effect upon degranulation. The differences between the outcome of infection by MV-Vac and WT-MV were not entirely explained by differential infection and replication of the viruses within neutrophils. To our knowledge, this is the first demonstration of potential mechanisms of oncolytic activity of an attenuated MV as compared with its WT parent. Furthermore, our study suggests that neutrophils have an important role to play in the antitumor effects of oncolytic MV.

Phosphatidylinositol 4-kinases: Function, structure, and inhibition

International Nuclear Information System (INIS)

Boura, Evzen; Nencka, Radim

2015-01-01

The phosphatidylinositol 4-kinases (PI4Ks) synthesize phosphatidylinositol 4-phosphate (PI4P), a key member of the phosphoinositide family. PI4P defines the membranes of Golgi and trans-Golgi network (TGN) and regulates trafficking to and from the Golgi. Humans have two type II PI4Ks (α and β) and two type III enzymes (α and β). Recently, the crystal structures were solved for both type II and type III kinase revealing atomic details of their function. Importantly, the type III PI4Ks are hijacked by +RNA viruses to create so-called membranous web, an extensively phosphorylated and modified membrane system dedicated to their replication. Therefore, selective and potent inhibitors of PI4Ks have been developed as potential antiviral agents. Here we focus on the structure and function of PI4Ks and their potential in human medicine

Phosphatidylinositol 4-kinases: Function, structure, and inhibition

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Boura, Evzen, E-mail: boura@uochb.cas.cz; Nencka, Radim, E-mail: nencka@uochb.cas.cz

2015-10-01

The phosphatidylinositol 4-kinases (PI4Ks) synthesize phosphatidylinositol 4-phosphate (PI4P), a key member of the phosphoinositide family. PI4P defines the membranes of Golgi and trans-Golgi network (TGN) and regulates trafficking to and from the Golgi. Humans have two type II PI4Ks (α and β) and two type III enzymes (α and β). Recently, the crystal structures were solved for both type II and type III kinase revealing atomic details of their function. Importantly, the type III PI4Ks are hijacked by +RNA viruses to create so-called membranous web, an extensively phosphorylated and modified membrane system dedicated to their replication. Therefore, selective and potent inhibitors of PI4Ks have been developed as potential antiviral agents. Here we focus on the structure and function of PI4Ks and their potential in human medicine.

Mind, brain, structure and function

Energy Technology Data Exchange (ETDEWEB)

Aleksander, I

1982-01-01

The author discusses the type of problem one encounters when trying to formalise the nature of a state structure associated with the brain and the origins of this state structure. The paper first defines in broad terms the nature of the structure function problem, and then goes on to separate out those parts of a structure that lead to the variational and adaptive nature of the state structure. It is argued that the relationship between the structure that leads to adaptation and its embedding in an external environment are crucial areas for further study. 4 references.

Sex-Specific Diurnal Immobility Induced by Forced Swim Test in Wild Type and Clock Gene Deficient Mice

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Ningyue Li

2015-03-01

Full Text Available Objective: The link between alterations in circadian rhythms and depression are well established, but the underlying mechanisms are far less elucidated. We investigated the circadian characteristics of immobility behavior in wild type (WT mice and mice with mutations in core Clock genes. Methods: All mice were tested with forced swim test (FST at 4 h intervals. Results: These experiments revealed significant diurnal rhythms associated with immobility behavior in both male and female WT mice with sex-different circadian properties. In addition, male mice showed significantly less immobility during the night phase in comparison to female mice. Female Per1Brdm1 mice also showed significant rhythmicity. However, the timing of rhythmicity was very different from that observed in female wild type mice. Male Per1Brdm1 mice showed a pattern of rhythmicity similar to that of wild type mice. Furthermore, female Per1Brdm1 mice showed higher duration of immobility in comparison to male Per1Brdm1 mice in both daytime and early night phases. Neither Per2Brdm1 nor ClockΔ19 mice showed significant rhythmicity, but both female Per2Brdm1 and ClockΔ19 mice had lower levels of immobility, compared to males. Conclusions: This study highlights the differences in the circadian characteristics of immobility induced by FST in WT, ClockΔ19, Per1, and Per2 deficient mice.

Multilocus sequence typing of commensal and enteropathogenic Escherichia coli from domestic and wild lagomorphs in Italy

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Giorgia Dotto

2015-12-01

Full Text Available The aim of the study was to determine the multilocus sequence types of Escherichia coli from diseased farm rabbits and apparently healthy wild lagomorphs, and the genetic relatedness among them. Fifty-five enteropathogenic E. coli from reared rabbits and 32 from wild rabbits and hares were characterised by multilocus sequence typing (MLST according to the Michigan State University EcMLST scheme. Isolates were differentiated into 37 sequence types (STs, which were grouped into 8 clonal complexes (CCs. The most common ST was ST140 (CC31, followed by ST238 and ST119 (CC17. MLST analysis revealed 22 novel STs. Phylogenetic analyses showed a heterogeneous distribution of STs into 3 clusters of genetically related strains. The genetic relationship among STs of different origin and the detection of new, as well as previously described STs as human pathogens, indicate a widespread distribution and adaptability of particular lineages to different hosts. These findings highlight the need for further research to improve the knowledge about E. coli populations colonising the gut of lagomorphs and their zoonotic potential.

Amyotrophic Lateral Sclerosis Type 20 - In Silico Analysis and Molecular Dynamics Simulation of hnRNPA1.

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Bruna Baumgarten Krebs

Full Text Available Amyotrophic Lateral Sclerosis (ALS is a fatal neurodegenerative disease that affects the upper and lower motor neurons. 5-10% of cases are genetically inherited, including ALS type 20, which is caused by mutations in the hnRNPA1 gene. The goals of this work are to analyze the effects of non-synonymous single nucleotide polymorphisms (nsSNPs on hnRNPA1 protein function, to model the complete tridimensional structure of the protein using computational methods and to assess structural and functional differences between the wild type and its variants through Molecular Dynamics simulations. nsSNP, PhD-SNP, Polyphen2, SIFT, SNAP, SNPs&GO, SNPeffect and PROVEAN were used to predict the functional effects of nsSNPs. Ab initio modeling of hnRNPA1 was made using Rosetta and refined using KoBaMIN. The structure was validated by PROCHECK, Rampage, ERRAT, Verify3D, ProSA and Qmean. TM-align was used for the structural alignment. FoldIndex, DICHOT, ELM, D2P2, Disopred and DisEMBL were used to predict disordered regions within the protein. Amino acid conservation analysis was assessed by Consurf, and the molecular dynamics simulations were performed using GROMACS. Mutations D314V and D314N were predicted to increase amyloid propensity, and predicted as deleterious by at least three algorithms, while mutation N73S was predicted as neutral by all the algorithms. D314N and D314V occur in a highly conserved amino acid. The Molecular Dynamics results indicate that all mutations increase protein stability when compared to the wild type. Mutants D314N and N319S showed higher overall dimensions and accessible surface when compared to the wild type. The flexibility level of the C-terminal residues of hnRNPA1 is affected by all mutations, which may affect protein function, especially regarding the protein ability to interact with other proteins.

Proteomics of Arabidopsis Seed Germination : a Comparative Study of Wild-Type and Gibberellin-Deficient Seeds

NARCIS (Netherlands)

Gallardo, K.; Job, C.; Groot, S.P.C.; Puype, M.; Vandekerckhove, J.; Job, D.

2002-01-01

We examined the role of gibberellins (GAs) in germination of Arabidopsis seeds by a proteomic approach. For that purpose, we used two systems. The first system consisted of seeds of the GA-deficient ga1 mutant, and the second corresponded to wild-type seeds incubated in paclobutrazol, a specific GA

A partial structural and functional rescue of a retinitis pigmentosa model with compacted DNA nanoparticles.

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Xue Cai

Full Text Available Previously we have shown that compacted DNA nanoparticles can drive high levels of transgene expression after subretinal injection in the mouse eye. Here we delivered compacted DNA nanoparticles containing a therapeutic gene to the retinas of a mouse model of retinitis pigmentosa. Nanoparticles containing the wild-type retinal degeneration slow (Rds gene were injected into the subretinal space of rds(+/- mice on postnatal day 5. Gene expression was sustained for up to four months at levels up to four times higher than in controls injected with saline or naked DNA. The nanoparticles were taken up into virtually all photoreceptors and mediated significant structural and biochemical rescue of the disease without histological or functional evidence of toxicity. Electroretinogram recordings showed that nanoparticle-mediated gene transfer restored cone function to a near-normal level in contrast to transfer of naked plasmid DNA. Rod function was also improved. These findings demonstrate that compacted DNA nanoparticles represent a viable option for development of gene-based interventions for ocular diseases and obviate major barriers commonly encountered with non-viral based therapies.

CD7 in acute myeloid leukemia: correlation with loss of wild-type CEBPA, consequence of epigenetic regulation

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Drexler Hans G

2010-04-01

Full Text Available Abstract Background CD7 is a negative prognostic marker in myeloid malignancies. In acute myeloid leukemia (AML, an inverse correlation exists between expression of wild-type CEBPA and CD7. Aim of this study was to find out whether C/EBPα is a negative regulator of CD7 and which other regulatory mechanisms might be involved. Results As already described for primary AML cells, the majority of AML cell lines tested were either C/EBPα+/CD7- or C/EBPα-/CD7+. However, the existence of isolated CD7+ cell lines expressing wild-type C/EBPα challenges the notion that C/EBPα acts as a unique repressor of CD7. Furthermore, ectopic expression of CEBPA did not reduce CD7 in CD7+ cells and knock-down of C/EBPα failed to induce CD7 in CD7- cells. In contrast, the DNA demethylating agent Aza-2'deoxycytidine triggered CD7 expression in CD7- AML and in T-cell lines suggesting epigenetic regulation of CD7. Bisulfite sequencing data confirmed that CpGs in the CD7 exon1 region are methylated in CD7- cell lines, and unmethylated in CD7+ cell lines. Conclusion We confirmed an inverse correlation between the expression of wild-type CEBPA and of CD7 in AML cells. Our results contradict the hypothesis that C/EBPα acts as repressor for CD7, and instead show that epigenetic mechanisms are responsible for CD7 regulation, in AML cells as well as in T-cells, the typical CD7 expressing cell type.

Shallow Boomerang-shaped Influenza Hemagglutinin G13A Mutant Structure Promotes Leaky Membrane Fusion*

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Lai, Alex L.; Tamm, Lukas K.

2010-01-01

Our previous studies showed that an angled boomerang-shaped structure of the influenza hemagglutinin (HA) fusion domain is critical for virus entry into host cells by membrane fusion. Because the acute angle of ∼105° of the wild-type fusion domain promotes efficient non-leaky membrane fusion, we asked whether different angles would still support fusion and thus facilitate virus entry. Here, we show that the G13A fusion domain mutant produces a new leaky fusion phenotype. The mutant fusion domain structure was solved by NMR spectroscopy in a lipid environment at fusion pH. The mutant adopted a boomerang structure similar to that of wild type but with a shallower kink angle of ∼150°. G13A perturbed the structure of model membranes to a lesser degree than wild type but to a greater degree than non-fusogenic fusion domain mutants. The strength of G13A binding to lipid bilayers was also intermediate between that of wild type and non-fusogenic mutants. These membrane interactions provide a clear link between structure and function of influenza fusion domains: an acute angle is required to promote clean non-leaky fusion suitable for virus entry presumably by interaction of the fusion domain with the transmembrane domain deep in the lipid bilayer. A shallower angle perturbs the bilayer of the target membrane so that it becomes leaky and unable to form a clean fusion pore. Mutants with no fixed boomerang angle interacted with bilayers weakly and did not promote any fusion or membrane perturbation. PMID:20826788

Partially dissecting the steady-state electron fluxes in Photosystem I in wild-type and pgr5 and ndh mutants of Arabidopsis

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Jiancun eKou

2015-09-01

Full Text Available Cyclic electron flux (CEF around Photosystem I (PS I is difficult to quantify. We obtained the linear electron flux (LEFO2 through both photosystems and the total electron flux through PS I (ETR1 in Arabidopsis in CO2-enriched air. DeltaFlux = ETR1 – LEFO2 is an upper estimate of CEF, which consists of two components, an antimycin A-sensitive, PGR5 (proton gradient regulation 5 protein-dependent component and an insensitive component facilitated by a chloroplastic nicotinamide adenine dinucleotide dehydrogenase-like complex (NDH. Using wild type as well as pgr5 and ndh mutants, we observed that (1 40% of the absorbed light was partitioned to PS I; (2 at high irradiance a substantial antimycin A-sensitive CEF occurred in the wild type and the ndh mutant; (3 at low irradiance a sizable antimycin A-sensitive CEF occurred in the wild type but not in the ndh mutant, suggesting an enhancing effect of NDH in low light; and (4 in the pgr5 mutant, and the wild type and ndh mutant treated with antimycin A, a residual DeltaFlux existed at high irradiance, attributable to charge recombination and/or pseudo-cyclic electron flow. Therefore, in low-light-acclimated plants exposed to high light, DeltaFlux has contributions from various paths of electron flow through PS I.

Hepcidin regulation in wild-type and Hfe knockout mice in response to alcohol consumption: evidence for an alcohol-induced hypoxic response.

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Heritage, Mandy L; Murphy, Therese L; Bridle, Kim R; Anderson, Gregory J; Crawford, Darrell H G; Fletcher, Linda M

2009-08-01

Expression of Hamp1, the gene encoding the iron regulatory peptide hepcidin, is inappropriately low in HFE-associated hereditary hemochromatosis and Hfe knockout mice (Hfe(-/-)). Since chronic alcohol consumption is also associated with disturbances in iron metabolism, we investigated the effects of alcohol consumption on hepcidin mRNA expression in Hfe(-/-) mice. Hfe(-/-) and C57BL/6 (wild-type) mice were pair-fed either an alcohol liquid diet or control diet for up to 8 weeks. The mRNA levels of hepcidin and ferroportin were measured at the mRNA level by RT-PCR and protein expression of hypoxia inducible factor-1 alpha (HIF-1alpha) was measured by western blot. Hamp1 mRNA expression was significantly decreased and duodenal ferroportin expression was increased in alcohol-fed wild-type mice at 8 weeks. Time course experiments showed that the decrease in hepcidin mRNA was not immediate, but was significant by 4 weeks. Consistent with the genetic defect, Hamp1 mRNA was decreased and duodenal ferroportin mRNA expression was increased in Hfe(-/-) mice fed on the control diet compared with wild-type animals and alcohol further exacerbated these effects. HIF-1alpha protein levels were elevated in alcohol-fed wild-type animals compared with controls. Alcohol may decrease Hamp1 gene expression independently of the HFE pathway possibly via alcohol-induced hypoxia.

Dissection of structural and functional requirements that underlie the interaction of ERdj3 protein with substrates in the endoplasmic reticulum.

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Otero, Joel H; Lizák, Beata; Feige, Matthias J; Hendershot, Linda M

2014-10-03

ERdj3, a mammalian endoplasmic reticulum (ER) Hsp40/DnaJ family member, binds unfolded proteins, transfers them to BiP, and concomitantly stimulates BiP ATPase activity. However, the requirements for ERdj3 binding to and release from substrates in cells are not well understood. We found that ERdj3 homodimers that cannot stimulate the ATPase activity of BiP (QPD mutants) bound to unfolded ER proteins under steady state conditions in much greater amounts than wild-type ERdj3. This was due to reduced release from these substrates as opposed to enhanced binding, although in both cases dimerization was strictly required for substrate binding. Conversely, heterodimers consisting of one wild-type and one mutant ERdj3 subunit bound substrates at levels comparable with wild-type ERdj3 homodimers, demonstrating that release requires only one protomer to be functional in stimulating BiP ATPase activity. Co-expressing wild-type ERdj3 and a QPD mutant, which each exclusively formed homodimers, revealed that the release rate of wild-type ERdj3 varied according to the relative half-lives of substrates, suggesting that ERdj3 release is an important step in degradation of unfolded client proteins in the ER. Furthermore, pulse-chase experiments revealed that the binding of QPD mutant homodimers remained constant as opposed to increasing, suggesting that ERdj3 does not normally undergo reiterative binding cycles with substrates. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Antibody Prevalence to Influenza Type A in Wild Boar of Northern Ukraine.

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Kovalenko, Ganna; Molozhanova, Alona; Halka, Ihor; Nychyk, Serhiy

2017-12-01

A preliminary serological survey was carried out to assess the likelihood of influenza A (IA) infection in wild boar and begin to characterize the role of wild boar in the epidemiology of the IA virus (IAV). Sera collected from 120 wild boar that were hunted in 2014 were tested. To detect antibodies to IA, a blocking the enzyme-linked immunosorbent assay (ELISA) was used. Thirty boar were collected from each of four oblasts in the north central and northwestern regions of Ukraine. Antibodies against IAV were detected in 27 samples (22.5%; 95% confidence interval 16.0-30.8) and in at least some of the wild boar from all of the four oblasts. This preliminary survey of IA antibodies in wild boar populations of northern Ukraine indicates a substantial frequency of exposure to IAV throughout the region. Infection of wild boar populations could provide an alternative or additional route for spillover from wild populations to domestic animals and humans.

Crystallization and preliminary X-ray crystallographic study of the wild type and two mutants of the CP1 hydrolytic domain from Aquifex aeolicus leucyl-tRNA synthetase

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Cura, Vincent; Olieric, Natacha; Guichard, Alexandre [Département de Biologie et Génomique Structurales, UMR 7104, Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Wang, En-Duo [State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, The Chinese Academy of Sciences, 320 Yue Yang Road, Shanghai 200031 (China); Moras, Dino [Département de Biologie et Génomique Structurales, UMR 7104, Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Eriani, Gilbert [Architecture et Réactivité de l’ARN, UPR 9002, Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 Rue René Descartes, 67084 Strasbourg (France); Cavarelli, Jean, E-mail: cava@igbmc.u-strasbg.fr [Département de Biologie et Génomique Structurales, UMR 7104, Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS/INSERM/ULP Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France)

2005-10-01

The wild-type editing CP1 domain of A. aeolicus leucyl-tRNA synthetase and two mutant CP1 domains have been overexpressed, purified and crystallized. X-ray diffraction data have been collected to 1.8 Å, which has enabled determination of the structures by molecular replacement. The editing or hydrolytic CP1 domain of leucyl-tRNA synthetase (LeuRS) hydrolyses several misactivated amino acids. The CP1 domain of Aquifex aeolicus LeuRS was expressed, purified and crystallized by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. Crystals belong to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 38.8, b = 98.4, c = 116.7 Å. Crystals diffract to beyond 1.8 Å resolution and contain two monomers in the asymmetric unit. Two CP1 mutants in which a conserved threonine residue essential for the fidelity of the hydrolytic pathway is mutated to alanine or glutamic acid have also been expressed and crystallized. Crystals of the two CP1 mutants are isomorphs of the wild type and diffract to beyond 1.9 Å resolution. All structures were solved by molecular-replacement techniques.

Crystallization and preliminary X-ray crystallographic study of the wild type and two mutants of the CP1 hydrolytic domain from Aquifex aeolicus leucyl-tRNA synthetase

International Nuclear Information System (INIS)

Cura, Vincent; Olieric, Natacha; Guichard, Alexandre; Wang, En-Duo; Moras, Dino; Eriani, Gilbert; Cavarelli, Jean

2005-01-01

The wild-type editing CP1 domain of A. aeolicus leucyl-tRNA synthetase and two mutant CP1 domains have been overexpressed, purified and crystallized. X-ray diffraction data have been collected to 1.8 Å, which has enabled determination of the structures by molecular replacement. The editing or hydrolytic CP1 domain of leucyl-tRNA synthetase (LeuRS) hydrolyses several misactivated amino acids. The CP1 domain of Aquifex aeolicus LeuRS was expressed, purified and crystallized by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. Crystals belong to space group P2 1 2 1 2 1 , with unit-cell parameters a = 38.8, b = 98.4, c = 116.7 Å. Crystals diffract to beyond 1.8 Å resolution and contain two monomers in the asymmetric unit. Two CP1 mutants in which a conserved threonine residue essential for the fidelity of the hydrolytic pathway is mutated to alanine or glutamic acid have also been expressed and crystallized. Crystals of the two CP1 mutants are isomorphs of the wild type and diffract to beyond 1.9 Å resolution. All structures were solved by molecular-replacement techniques

Mutant INS-gene induced diabetes of youth: proinsulin cysteine residues impose dominant-negative inhibition on wild-type proinsulin transport.

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Ming Liu

2010-10-01

Full Text Available Recently, a syndrome of Mutant INS-gene-induced Diabetes of Youth (MIDY, derived from one of 26 distinct mutations has been identified as a cause of insulin-deficient diabetes, resulting from expression of a misfolded mutant proinsulin protein in the endoplasmic reticulum (ER of insulin-producing pancreatic beta cells. Genetic deletion of one, two, or even three alleles encoding insulin in mice does not necessarily lead to diabetes. Yet MIDY patients are INS-gene heterozygotes; inheritance of even one MIDY allele, causes diabetes. Although a favored explanation for the onset of diabetes is that insurmountable ER stress and ER stress response from the mutant proinsulin causes a net loss of beta cells, in this report we present three surprising and interlinked discoveries. First, in the presence of MIDY mutants, an increased fraction of wild-type proinsulin becomes recruited into nonnative disulfide-linked protein complexes. Second, regardless of whether MIDY mutations result in the loss, or creation, of an extra unpaired cysteine within proinsulin, Cys residues in the mutant protein are nevertheless essential in causing intracellular entrapment of co-expressed wild-type proinsulin, blocking insulin production. Third, while each of the MIDY mutants induces ER stress and ER stress response; ER stress and ER stress response alone appear insufficient to account for blockade of wild-type proinsulin. While there is general agreement that ultimately, as diabetes progresses, a significant loss of beta cell mass occurs, the early events described herein precede cell death and loss of beta cell mass. We conclude that the molecular pathogenesis of MIDY is initiated by perturbation of the disulfide-coupled folding pathway of wild-type proinsulin.

Structural and Functional Consequences of Chaperone Site Deletion in αA-Crystallin

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Santhoshkumar, Puttur; Karmakar, Srabani; Sharma, Krishna K.

2016-01-01

The chaperone-like activity of αA-crystallin has an important role in maintaining lens transparency. Previously we identified residues 70–88 as a chaperone site in αA-crystallin. In this study, we deleted the chaperone site residues to generate αAΔ70–76 and αAΔ70–88 mutants and investigated if there are additional substrate-binding sites in αA-crystallin. Both mutant proteins when expressed in E. coli formed inclusion bodies, and on solubilizing and refolding, they exhibited similar structural properties, with a 2- to 3-fold increase in molar mass compared to the molar mass of wild-type protein. The deletion mutants were less stable than the wild-type αA-crystallin. Functionally αAΔ70–88 was completely inactive as a chaperone, while αAΔ70–76 demonstrated a 40–50% reduction in anti-aggregation activity against alcohol dehydrogenase (ADH). Deletion of residues 70–88 abolished the ADH binding sites in αA-crystallin at physiological temperature. At 45 °C, cryptic ADH binding site(s) became exposed, which contributed subtly to the chaperone-like activity of αAΔ70–88. Both of the deletion mutants were completely inactive in suppressing aggregation of βL-crystallin at 53 °C. The mutants completely lost the anti-apoptotic property that αA-crystallin exhibits while they protected ARPE-19 (a human retinal pigment epithelial cell line) and primary human lens epithelial (HLE) cells from oxidative stress. Our studies demonstrate that residues 70–88 in αA-crystallin act as a primary substrate binding site and account for the bulk of the total chaperone activity. The β3 and β4 strands in αA-crystallin comprising 70–88 residues play an important role in maintenance of the structure and in preventing aggregation of denaturing proteins. PMID:27524665

Coordination of Leaf Photosynthesis, Transpiration, and Structural Traits in Rice and Wild Relatives (Genus Oryza).

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Giuliani, Rita; Koteyeva, Nuria; Voznesenskaya, Elena; Evans, Marc A; Cousins, Asaph B; Edwards, Gerald E

2013-07-01

The genus Oryza, which includes rice (Oryza sativa and Oryza glaberrima) and wild relatives, is a useful genus to study leaf properties in order to identify structural features that control CO(2) access to chloroplasts, photosynthesis, water use efficiency, and drought tolerance. Traits, 26 structural and 17 functional, associated with photosynthesis and transpiration were quantified on 24 accessions (representatives of 17 species and eight genomes). Hypotheses of associations within, and between, structure, photosynthesis, and transpiration were tested. Two main clusters of positively interrelated leaf traits were identified: in the first cluster were structural features, leaf thickness (Thick(leaf)), mesophyll (M) cell surface area exposed to intercellular air space per unit of leaf surface area (S(mes)), and M cell size; a second group included functional traits, net photosynthetic rate, transpiration rate, M conductance to CO(2) diffusion (g(m)), stomatal conductance to gas diffusion (g(s)), and the g(m)/g(s) ratio.While net photosynthetic rate was positively correlated with gm, neither was significantly linked with any individual structural traits. The results suggest that changes in gm depend on covariations of multiple leaf (S(mes)) and M cell (including cell wall thickness) structural traits. There was an inverse relationship between Thick(leaf) and transpiration rate and a significant positive association between Thick(leaf) and leaf transpiration efficiency. Interestingly, high g(m) together with high g(m)/g(s) and a low S(mes)/g(m) ratio (M resistance to CO(2) diffusion per unit of cell surface area exposed to intercellular air space) appear to be ideal for supporting leaf photosynthesis while preserving water; in addition, thick M cell walls may be beneficial for plant drought tolerance.

STUDY DATA OF KRAS- AND RAS-UNMUTATED (WILD TYPE OF COLORECTAL CANCER

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V. A. Gorbunova

2015-01-01

Full Text Available  Analysis of latest trials, comparing treatment schemes including chemotherapy with anti-EGFR monoclonal antibodies or bevacizumab is presented in this article. The data in these trials is inconsistent, but detailed analysis of FIRE-3 trial allows to distinguish a wild-type RAS patient group that benefits most from chemotherapy with cetuximab or panitumumab as 1st line metastatic colorectal cancer treatment. A final analysis of this patient group in CALGB/SWOG 80 405 trial is pending. The RAS analysis is pivotal for choice of 1st line chemotherapy.

Collagens - structure, function and biosynthesis.

OpenAIRE

Gelse, K; Poschl, E; Aigner, T

2003-01-01

The extracellular matrix represents a complex alloy of variable members of diverse protein families defining structural integrity and various physiological functions. The most abundant family is the collagens with more than 20 different collagen types identified so far. Collagens are centrally involved in the formation of fibrillar and microfibrillar networks of the extracellular matrix, basement membranes as well as other structures of the extracellular matrix. This review focuses on the dis...

Early Transcriptional Responses of Bovine Chorioallantoic Membrane Explants to Wild Type, ΔvirB2 or ΔbtpB Brucella abortus Infection

Science.gov (United States)

Mol, Juliana P. S.; Costa, Erica A.; Carvalho, Alex F.; Sun, Yao-Hui; Tsolis, Reneé M.; Paixão, Tatiane A.; Santos, Renato L.

2014-01-01

The pathogenesis of the Brucella-induced inflammatory response in the bovine placenta is not completely understood. In this study we evaluated the role of the B. abortus Type IV secretion system and the anti-inflammatory factor BtpB in early interactions with bovine placental tissues. Transcription profiles of chorioallantoic membrane (CAM) explants inoculated with wild type (strain 2308), ΔvirB2 or ΔbtpB Brucella abortus were compared by microarray analysis at 4 hours post infection. Transcripts with significant variation (>2 fold change; Pabortus resulted in slightly more genes with decreased than increased transcription levels. Conversely, infection of trophoblastic cells with the ΔvirB2 or the ΔbtpB mutant strains, that lack a functional T4SS or that has impaired inhibition of TLR signaling, respectively, induced more upregulated than downregulated genes. Wild type Brucella abortus impaired transcription of host genes related to immune response when compared to ΔvirB and ΔbtpB mutants. Our findings suggest that proinflammatory genes are negatively modulated in bovine trophoblastic cells at early stages of infection. The virB operon and btpB are directly or indirectly related to modulation of these host genes. These results shed light on the early interactions between B. abortus and placental tissue that ultimately culminate in inflammatory pathology and abortion. PMID:25259715

Life table and male mating competitiveness of wild type and of a chromosome mutation strain of Tetranychus urticae in relation to genetic pest control

International Nuclear Information System (INIS)

Feldmann, A.M.

1981-01-01

Males of Tetranychus urticae Koch (Acarina: Tetranychidae) from a strain, homozygous for a structural chromosome mutation (T) were competed against males from a standard (wild-type) strain for mating of wild-type fermales. The T-males exhibited only a slight reduction in male mating competitiveness. The debilitating influence of ageing on male mating competitiveness was equal for males of both strains. Life-table studies on both strains showed that the net reproductive rate (R 0 ) of the T-strain was 53.3, which was higher than the R 0 -value of the standard strain (43.3). This difference was caused by the higher rate of age-dependent mortality of adult females of the standard strain. Also differences between both strains in the total sex-ratio were observed; the T-strain produced significantly fewer males and more females than the standard strain. The mean generation time of both strains was almost equal (14 days). The values of the intrinsic rate of increase (rsub(m)) for the T-strain and the standard strain were 0.286 and 0.273, respectively. The life-table data correspond well with those published elsewhere on Tetranychus urticae. The feasibility of T-strains for application in genetic pest control considering the use of structural chromosome mutations as a 'transport mechanism' for conditional lethals is discussed. (orig.)

Type checking with open type functions

DEFF Research Database (Denmark)

Schrijvers, Tom; Jones, Simon Peyton; Chakravarty, Manual

2008-01-01

We report on an extension of Haskell with open type-level functions and equality constraints that unifies earlier work on GADTs, functional dependencies, and associated types. The contribution of the paper is that we identify and characterise the key technical challenge of entailment checking; an...

High taxonomic variability despite stable functional structure across microbial communities.

Science.gov (United States)

Louca, Stilianos; Jacques, Saulo M S; Pires, Aliny P F; Leal, Juliana S; Srivastava, Diane S; Parfrey, Laura Wegener; Farjalla, Vinicius F; Doebeli, Michael

2016-12-05

Understanding the processes that are driving variation of natural microbial communities across space or time is a major challenge for ecologists. Environmental conditions strongly shape the metabolic function of microbial communities; however, other processes such as biotic interactions, random demographic drift or dispersal limitation may also influence community dynamics. The relative importance of these processes and their effects on community function remain largely unknown. To address this uncertainty, here we examined bacterial and archaeal communities in replicate 'miniature' aquatic ecosystems contained within the foliage of wild bromeliads. We used marker gene sequencing to infer the taxonomic composition within nine metabolic functional groups, and shotgun environmental DNA sequencing to estimate the relative abundances of these groups. We found that all of the bromeliads exhibited remarkably similar functional community structures, but that the taxonomic composition within individual functional groups was highly variable. Furthermore, using statistical analyses, we found that non-neutral processes, including environmental filtering and potentially biotic interactions, at least partly shaped the composition within functional groups and were more important than spatial dispersal limitation and demographic drift. Hence both the functional structure and taxonomic composition within functional groups of natural microbial communities may be shaped by non-neutral and roughly separate processes.

Identification of Quaternary Structure and Functional Domains of the CI Repressor from Bacteriophage TP901-1

DEFF Research Database (Denmark)

Pedersen, Margit; Lo Leggio, Leila; Grossmann, J. Günter

2008-01-01

is involved in the interaction with host proteins. By using small-angle X-ray scattering, we show for the first time the overall solution structure of a full-length wild-type bacteriophage repressor at low resolution revealing that the TP901-1 repressor forms a flat oligomer, most probably a trimer of dimers....

Yellow fluorescent protein phiYFPv (Phialidium): structure and structure-based mutagenesis

Energy Technology Data Exchange (ETDEWEB)

Pletneva, Nadya V.; Pletnev, Vladimir Z., E-mail: vzpletnev@gmail.com; Souslova, Ekaterina; Chudakov, Dmitry M. [Russian Academy of Sciences, Moscow (Russian Federation); Lukyanov, Sergey [Russian Academy of Sciences, Moscow (Russian Federation); Nizhny Novgorod State Medical Academy, Nizhny Novgorod (Russian Federation); Martynov, Vladimir I.; Arhipova, Svetlena; Artemyev, Igor [Russian Academy of Sciences, Moscow (Russian Federation); Wlodawer, Alexander [National Cancer Institute, Frederick, MD 21702 (United States); Dauter, Zbigniew [National Cancer Institute, Argonne, IL 60439 (United States); Pletnev, Sergei [National Cancer Institute, Argonne, IL 60439 (United States); SAIC-Frederick, 9700 South Cass Avenue, Argonne, IL 60439 (United States); Russian Academy of Sciences, Moscow (Russian Federation)

2013-06-01

The yellow fluorescent protein phiYFPv with improved folding has been developed from the spectrally identical wild-type phiYFP found in the marine jellyfish Phialidium. The yellow fluorescent protein phiYFPv (λ{sub em}{sup max} ≃ 537 nm) with improved folding has been developed from the spectrally identical wild-type phiYFP found in the marine jellyfish Phialidium. The latter fluorescent protein is one of only two known cases of naturally occurring proteins that exhibit emission spectra in the yellow–orange range (535–555 nm). Here, the crystal structure of phiYFPv has been determined at 2.05 Å resolution. The ‘yellow’ chromophore formed from the sequence triad Thr65-Tyr66-Gly67 adopts the bicyclic structure typical of fluorophores emitting in the green spectral range. It was demonstrated that perfect antiparallel π-stacking of chromophore Tyr66 and the proximal Tyr203, as well as Val205, facing the chromophore phenolic ring are chiefly responsible for the observed yellow emission of phiYFPv at 537 nm. Structure-based site-directed mutagenesis has been used to identify the key functional residues in the chromophore environment. The obtained results have been utilized to improve the properties of phiYFPv and its homologous monomeric biomarker tagYFP.

Wild-type bone marrow transplant partially reverses neuroinflammation in progranulin-deficient mice.

Science.gov (United States)

Yang, Yue; Aloi, Macarena S; Cudaback, Eiron; Josephsen, Samuel R; Rice, Samantha J; Jorstad, Nikolas L; Keene, C Dirk; Montine, Thomas J

2014-11-01

Frontotemporal dementia (FTD) is a neurodegenerative disease with devastating changes in behavioral performance and social function. Mutations in the progranulin gene (GRN) are one of the most common causes of inherited FTD due to reduced progranulin expression or activity, including in brain where it is expressed primarily by neurons and microglia. Thus, efforts aimed at enhancing progranulin levels might be a promising therapeutic strategy. Bone marrow (BM)-derived cells are able to engraft in the brain and adopt a microglial phenotype under myeloablative irradiation conditioning. This ability makes BM-derived cells a potential cellular vehicle for transferring therapeutic molecules to the central nervous system. Here, we utilized BM cells from Grn(+/+) (wild type or wt) mice labeled with green fluorescence protein for delivery of progranulin to progranulin-deficient (Grn(-/-)) mice. Our results showed that wt bone marrow transplantation (BMT) partially reconstituted progranulin in the periphery and in cerebral cortex of Grn(-/-) mice. We demonstrated a pro-inflammatory effect in vivo and in ex vivo preparations of cerebral cortex of Grn(-/-) mice that was partially to fully reversed 5 months after BMT. Our findings suggest that BMT can be administered as a stem cell-based approach to prevent or to treat neurodegenerative diseases.

Wild Type Bone Marrow Transplant Partially Reverses Neuroinflammation in Progranulin-Deficient Mice

Science.gov (United States)

Yang, Yue; Aloi, Macarena S.; Cudaback, Eiron; Josephsen, Samuel R.; Rice, Samantha J.; Jorstad, Nikolas L.; Keene, C. Dirk; Montine, Thomas J.

2014-01-01

Frontotemporal dementia (FTD) is a neurodegenerative disease with devastating changes in behavioral performance and social function. Mutations in the progranulin gene (GRN) are one of the most common causes of inherited FTD due to reduced progranulin expression or activity, including in brain where it is expressed primarily by neurons and microglia. Thus, efforts aimed at enhancing progranulin levels might be a promising therapeutic strategy. Bone marrow-derived cells are able to engraft in the brain and adopt a microglial phenotype under myeloablative irradiation conditioning. This ability makes bone marrow (BM)-derived cells a potential cellular vehicle for transferring therapeutic molecules to the central nervous system. Here, we utilized BM cells from Grn+/+ (wild type or wt) mice labeled with green fluorescence protein for delivery of progranulin to progranulin deficient (Grn−/−) mice. Our results showed that wt bone marrow transplantation (BMT) partially reconstituted progranulin in the periphery and in cerebral cortex of Grn−/− mice. We demonstrated a pro-inflammatory effect in vivo and in ex vivo preparations of cerebral cortex of Grn−/− mice that was partially to fully reversed five months after BMT. Our findings suggest that BMT can be administered as a stem cell-based approach to prevent or to treat neurodegenerative diseases. PMID:25199051

Genetic Characterization of the Hemagglutinin Genes of Wild-Type Measles Virus Circulating in China, 1993–2009

Science.gov (United States)

Zhu, Zhen; Liu, Chunyu; Mao, Naiying; Ji, Yixin; Wang, Huiling; Jiang, Xiaohong; Li, Chongshan; Tang, Wei; Feng, Daxing; Wang, Changyin; Zheng, Lei; Lei, Yue; Ling, Hua; Zhao, Chunfang; Ma, Yan; He, Jilan; Wang, Yan; Li, Ping; Guan, Ronghui; Zhou, Shujie; Zhou, Jianhui; Wang, Shuang; Zhang, Hong; Zheng, Huanying; Liu, Leng; Ma, Hemuti; Guan, Jing; Lu, Peishan; Feng, Yan; Zhang, Yanjun; Zhou, Shunde; Xiong, Ying; Ba, Zhuoma; Chen, Hui; Yang, Xiuhui; Bo, Fang; Ma, Yujie; Liang, Yong; Lei, Yake; Gu, Suyi; Liu, Wei; Chen, Meng; Featherstone, David; Jee, Youngmee; Bellini, William J.; Rota, Paul A.; Xu, Wenbo

2013-01-01

Background China experienced several large measles outbreaks in the past two decades, and a series of enhanced control measures were implemented to achieve the goal of measles elimination. Molecular epidemiologic surveillance of wild-type measles viruses (MeV) provides valuable information about the viral transmission patterns. Since 1993, virologic surveillnace has confirmed that a single endemic genotype H1 viruses have been predominantly circulating in China. A component of molecular surveillance is to monitor the genetic characteristics of the hemagglutinin (H) gene of MeV, the major target for virus neutralizing antibodies. Principal Findings Analysis of the sequences of the complete H gene from 56 representative wild-type MeV strains circulating in China during 1993–2009 showed that the H gene sequences were clustered into 2 groups, cluster 1 and cluster 2. Cluster1 strains were the most frequently detected cluster and had a widespread distribution in China after 2000. The predicted amino acid sequences of the H protein were relatively conserved at most of the functionally significant amino acid positions. However, most of the genotype H1 cluster1 viruses had an amino acid substitution (Ser240Asn), which removed a predicted N-linked glycosylation site. In addition, the substitution of Pro397Leu in the hemagglutinin noose epitope (HNE) was identified in 23 of 56 strains. The evolutionary rate of the H gene of the genotype H1 viruses was estimated to be approximately 0.76×10−3 substitutions per site per year, and the ratio of dN to dS (dN/dS) was measles in China. PMID:24073194

Food preferences of winter bird communities in different forest types.

Directory of Open Access Journals (Sweden)

Swen C Renner

Full Text Available Food availability for forest birds is a function of habitat type, forest management regime, and season. In winter, it is also impacted by variations in the weather. In the current study we assessed the food preferences of wild bird populations in two types of forest (spruce and beech during the months of November 2010 to April 2011 in the Schwäbische Alb Biodiversity Exploratory, south-western Germany. Our aim was to investigate whether local bird communities preferred fat-rich, carbohydrate-rich or wild fruits and to determine how forest structure, seasonality and local weather conditions affected food preferences. We found higher bird activity in beech forests for the eleven resident species. We observed a clear preference for fat-rich food for all birds in both forest types. Snow cover affected activity at food stations but did not affect food preferences. Periods of extreme low temperatures increased activity.

Population genetic structure in wild and aquaculture populations of Hemibarbus maculates inferred from microsatellites markers

Directory of Open Access Journals (Sweden)

Linlin Li

2017-03-01

Full Text Available The objective of this study was to investigate 4 aquaculture populations Shanghai (SH, Hangzhou (HZ, Kaihua (KH and Xianju (XJ and one wild population Yingshan (YS of spotted barbell (Hemibarbus maculates to assess their genetic diversity level and investigate the genetic structure of the populations. The dendrogram and STRUCTURE revealed that the populations XJ, KH, and HZ jointly formed one cluster, to which the populations SH and YS were sequentially adhered. The genetic diversity of the cultured populations maintained better, possible due to favourable hatchery conditions that decreased the effect of environmental selection present in wild populations. The results of the present study will contribute to the management of spotted barbell genetic resources, but also demonstrates how the genetic diversity of freshwater species is vulnerable to human activity.

Effects of animal type (wild vs. domestic) and diet alfalfa level on intake and digestibility of European adult rabbits (Oryctolagus cuniculus)

NARCIS (Netherlands)

Pinheiro, Victor; Outor-Monteiro, D.; Mourão, J.L.; Cone, J.W.; Guimaraes Dias Lourenco, Ana

2018-01-01

The aim of this study was to evaluate the effect of the level of alfalfa in the diet on feed intake and digestibility of two types of rabbits, wild (Oryctolagus cuniculus algirus) vs. domestic (O. cuniculus cuniculus). Ten wild (W; mean LW = 927 g) and 10 domestic (D; mean LW = 4,645 g) adult rabbit

The In Vivo Granulopoietic Response to Dexamethasone Injection Is Abolished in Perforin-Deficient Mutant Mice and Corrected by Lymphocyte Transfer from Nonsensitized Wild-Type Donors

Directory of Open Access Journals (Sweden)

Pedro Xavier-Elsas

2015-01-01

Full Text Available Exogenously administered glucocorticoids enhance eosinophil and neutrophil granulocyte production from murine bone-marrow. A hematological response dependent on endogenous glucocorticoids underlies bone-marrow eosinophilia induced by trauma or allergic sensitization/challenge. We detected a defect in granulopoiesis in nonsensitized, perforin-deficient mice. In steady-state conditions, perforin- (Pfp- deficient mice showed significantly decreased bone-marrow and blood eosinophil and neutrophil counts, and colony formation in response to GM-CSF, relative to wild-type controls of comparable age and/or weight. By contrast, peripheral blood or spleen total cell and lymphocyte numbers were not affected by perforin deficiency. Dexamethasone enhanced colony formation by GM-CSF-stimulated progenitors from wild-type controls, but not Pfp mice. Dexamethasone injection increased bone-marrow eosinophil and neutrophil counts in wild-type controls, but not Pfp mice. Because perforin is expressed in effector lymphocytes, we examined whether this defect would be corrected by transferring wild-type lymphocytes into perforin-deficient recipients. Short-term reconstitution of the response to dexamethasone was separately achieved for eosinophils and neutrophils by transfer of distinct populations of splenic lymphocytes from nonsensitized wild-type donors. Transfer of the same amount of splenic lymphocytes from perforin-deficient donors was ineffective. This demonstrates that the perforin-dependent, granulopoietic response to dexamethasone can be restored by transfer of innate lymphocyte subpopulations.

Role of UV-inducible proteins in repair of various wild-type Escherichia coli cells

International Nuclear Information System (INIS)

Sedliakova, M.; Slezarikova, V.; Brozmanova, J.; Masek, F.; Bayerova, V.

1980-01-01

3 wild-type strains of E. coli, namely K12 AB2497, B/r WP2 and 15 555-7, proficient in excision and post-replication repair, differ markedly in their UV resistance. To elucidate this difference, the influence was investigated of induction by application of inducing fluence (IF) before lethal fluence (LF) on repair processes after LF. In cells distinguished by low UV resistance (E. coli 15 555-7; E. coli B/r WP2), dimer excision was less complete in cultures irradiated with IF + LF than in cultures irradiated with LF only. The highly resistant E. coli K12 AB2497 performed complete excision both after IF + LF or after LF alone. All 3 types of cell survived better after IF + LF than after LF only. Because, in most strains so far investigated, the application of IF reduced dimer excision and increased survival, dimer excision per se does not appear important for survival. We conclude that the rate and completeness of dimer excision can serve as a measure of efficiency of the excision system whose action is necessary for repair of another lesion. Cells of all investigated strains could not resume DNA replication and died progressively when irradiated with LF and post-incubated with chloramphenicol (LF CAP + ). Thus, it appears that inducible proteins are necessary for repair in all wild-type E. coli cells given with potentially lethal doses of UV irradiation. (orig.)

Reduction of structured population models to threshold-type delay equations and functional differential equations: A case study

Energy Technology Data Exchange (ETDEWEB)

Smith, H.L. (Arizona State Univ., Tempe (United States))

1993-01-01

It is shown by way of a simple example that certain structured population models lead naturally to differential delay equations of the threshold type and that these equations can be transformed in a natural way to functional differential equations. The model examined can be viewed as a model of competition between adults and juveniles of a single population. The results indicate the possibility that this competition leads to instability. 28 refs., 2 figs.

Effective constraint algebras with structure functions

International Nuclear Information System (INIS)

Bojowald, Martin; Brahma, Suddhasattwa

2016-01-01

This article presents the result that fluctuations and higher moments of a state, by themselves, do not imply quantum corrections in structure functions of constrained systems. Moment corrections are isolated from other types of quantum effects, such as factor-ordering choices and regularization, by introducing a new condition with two parts: (i) having a direct (or faithful) quantization of the classical structure functions, (ii) free of factor-ordering ambiguities. In particular, it is assumed that the classical constraints can be quantized in an anomaly free way, so that properties of the resulting constraint algebras can be derived. If the two-part condition is not satisfied, effective constraints can still be evaluated, but quantum effects may be stronger. Consequences for canonical quantum gravity, whose structure functions encode space–time structure, are discussed. In particular, deformed algebras found in models of loop quantum gravity provide reliable information even in the Planck regime. (paper)

Efficient Reassignment of a Frequent Serine Codon in Wild-Type Escherichia coli.

Science.gov (United States)

Ho, Joanne M; Reynolds, Noah M; Rivera, Keith; Connolly, Morgan; Guo, Li-Tao; Ling, Jiqiang; Pappin, Darryl J; Church, George M; Söll, Dieter

2016-02-19

Expansion of the genetic code through engineering the translation machinery has greatly increased the chemical repertoire of the proteome. This has been accomplished mainly by read-through of UAG or UGA stop codons by the noncanonical aminoacyl-tRNA of choice. While stop codon read-through involves competition with the translation release factors, sense codon reassignment entails competition with a large pool of endogenous tRNAs. We used an engineered pyrrolysyl-tRNA synthetase to incorporate 3-iodo-l-phenylalanine (3-I-Phe) at a number of different serine and leucine codons in wild-type Escherichia coli. Quantitative LC-MS/MS measurements of amino acid incorporation yields carried out in a selected reaction monitoring experiment revealed that the 3-I-Phe abundance at the Ser208AGU codon in superfolder GFP was 65 ± 17%. This method also allowed quantification of other amino acids (serine, 33 ± 17%; phenylalanine, 1 ± 1%; threonine, 1 ± 1%) that compete with 3-I-Phe at both the aminoacylation and decoding steps of translation for incorporation at the same codon position. Reassignments of different serine (AGU, AGC, UCG) and leucine (CUG) codons with the matching tRNA(Pyl) anticodon variants were met with varying success, and our findings provide a guideline for the choice of sense codons to be reassigned. Our results indicate that the 3-iodo-l-phenylalanyl-tRNA synthetase (IFRS)/tRNA(Pyl) pair can efficiently outcompete the cellular machinery to reassign select sense codons in wild-type E. coli.

Generalized fluid impulse functions for oscillating marine structures

Science.gov (United States)

Janardhanan, K.; Price, W. G.; Wu, Y.

1992-03-01

A selection of generalized impulse response functions is presented for a variety of rigid and flexible marine structures (i.e. mono-hull, SWATH, floating drydock and twin dock, fixed flexible pile). These functions are determined from calculated and experimental frequency-dependent hydrodynamic data, and the characteristics of these data depend on the type of structure considered. This information is reflected in the shape and duration of the generalized impulse response functions which are pre-requisites for a generalized integro-differential mathematical model describing the dynamic behaviour of the structures to seaway excitation.

Effects of ketoconazole on the biodistribution and metabolism of [{sup 11}C]loperamide and [{sup 11}C]N-desmethyl-loperamide in wild-type and P-gp knockout mice

Energy Technology Data Exchange (ETDEWEB)

Seneca, Nicholas; Zoghbi, Sami S.; Shetty, H. Umesha; Tuan, Edward; Kannan, Pavitra; Taku, Andrew; Innis, Robert B. [Molecular Imaging Branch, National Institute of Mental Health, National Institutes of Health, Bethesda, MD 20892 (United States); Pike, Victor W. [Molecular Imaging Branch, National Institute of Mental Health, National Institutes of Health, Bethesda, MD 20892 (United States)], E-mail: pikev@mail.nih.gov

2010-04-15

Introduction: [{sup 11}C]Loperamide and [{sup 11}C]N-desmethyl-loperamide ([{sup 11}C]dLop) have been proposed as radiotracers for imaging brain P-glycoprotein (P-gp) function. A major route of [{sup 11}C]loperamide metabolism is N-demethylation to [{sup 11}C]dLop. We aimed to test whether inhibition of CYP3A4 with ketoconazole might reduce the metabolism of [{sup 11}C]loperamide and [{sup 11}C]dLop in mice, and thereby improve the quality of these radiotracers. Methods: Studies were performed in wild-type and P-gp knockout (mdr-1a/b -/-) mice. During each of seven study sessions, one pair of mice, comprising one wild-type and one knockout mouse, was pretreated with ketoconazole (50 mg/kg, ip), while another such pair was left untreated. Mice were sacrificed at 30 min after injection of [{sup 11}C]loperamide or [{sup 11}C]dLop. Whole brain and plasma samples were measured for radioactivity and analyzed with radio-high-performance liquid chromatography. Results: Ketoconazole increased the plasma concentrations of [{sup 11}C]loperamide and its main radiometabolite, [{sup 11}C]dLop, by about twofold in both wild-type and knockout mice, whereas the most polar radiometabolite was decreased threefold. Furthermore, ketoconazole increased the brain concentrations of [{sup 11}C]loperamide and the radiometabolite [{sup 11}C]dLop by about twofold in knockout mice, and decreased the brain concentrations of the major and most polar radiometabolite in wild-type and knockout mice by 82% and 49%, respectively. In contrast, ketoconazole had no effect on plasma and brain distribution of administered [{sup 11}C]dLop and its radiometabolites in either wild-type or knockout mice, except to increase the low plasma [{sup 11}C]dLop concentration. The least polar radiometabolite of [{sup 11}C]dLop was identified with LC-MS{sup n} as the N-hydroxymethyl analog of [{sup 11}C]dLop and this also behaved as a P-gp substrate. Conclusion: In this study, ketoconazole (50 mg/kg, ip) proved

Near-Infrared Spectroscopy, a Rapid Method for Predicting the Age of Male and Female Wild-Type and Wolbachia Infected Aedes aegypti.

Directory of Open Access Journals (Sweden)

Maggy T Sikulu-Lord

2016-10-01

Full Text Available Estimating the age distribution of mosquito populations is crucial for assessing their capacity to transmit disease and for evaluating the efficacy of available vector control programs. This study reports on the capacity of the near-infrared spectroscopy (NIRS technique to rapidly predict the ages of the principal dengue and Zika vector, Aedes aegypti. The age of wild-type males and females, and males and females infected with wMel and wMelPop strains of Wolbachia pipientis were characterized using this method. Calibrations were developed using spectra collected from their heads and thoraces using partial least squares (PLS regression. A highly significant correlation was found between the true and predicted ages of mosquitoes. The coefficients of determination for wild-type females and males across all age groups were R2 = 0.84 and 0.78, respectively. The coefficients of determination for the age of wMel and wMelPop infected females were 0.71 and 0.80, respectively (P< 0.001 in both instances. The age of wild-type female Ae. aegypti could be identified as < or ≥ 8 days old with an accuracy of 91% (N = 501, whereas female Ae. aegypti infected with wMel and wMelPop were differentiated into the two age groups with an accuracy of 83% (N = 284 and 78% (N = 229, respectively. Our results also indicate NIRS can distinguish between young and old male wild-type, wMel and wMelPop infected Ae. aegypti with accuracies of 87% (N = 253, 83% (N = 277 and 78% (N = 234, respectively. We have demonstrated the potential of NIRS as a predictor of the age of female and male wild-type and Wolbachia infected Ae. aegypti mosquitoes under laboratory conditions. After field validation, the tool has the potential to offer a cheap and rapid alternative for surveillance of dengue and Zika vector control programs.

Processes affecting genetic structure and conservation: a case study of wild and cultivated Brassica rapa

DEFF Research Database (Denmark)

Andersen, Naja Steen; Poulsen, Gert; Andersen, Bente Anni

2009-01-01

When planning optimal conservation strategies for wild and cultivated types of a plant species, a number of influencing biological and environmental factors should be considered from the outset. In the present study Brassica rapa was used to illustrate this: to develop Scandinavian conservation...

Bipartite structure and functional independence of adenovirus type 5 packaging elements.

OpenAIRE

Schmid, S I; Hearing, P

1997-01-01

Selectivity and polarity of adenovirus type 5 DNA packaging are believed to be directed by an interaction of putative packaging factors with the cis-acting adenovirus packaging domain located within the genomic left end (nucleotides 194 to 380). In previous studies, this packaging domain was mutationally dissected into at least seven functional elements called A repeats. These elements, albeit redundant in function, exhibit differences in the ability to support viral packaging, with elements ...

Syringolin A selectively labels the 20 S proteasome in murine EL4 and wild-type and bortezomib-adapted leukaemic cell lines.

Science.gov (United States)

Clerc, Jérôme; Florea, Bogdan I; Kraus, Marianne; Groll, Michael; Huber, Robert; Bachmann, André S; Dudler, Robert; Driessen, Christoph; Overkleeft, Herman S; Kaiser, Markus

2009-11-02

The natural product syringolin A (SylA) is a potent proteasome inhibitor with promising anticancer activities. To further investigate its potential as a lead structure, selectivity profiling with cell lysates was performed. At therapeutic concentrations, a rhodamine-tagged SylA derivative selectively bound to the 20 S proteasome active sites without detectable off-target labelling. Additional profiling with lysates of wild-type and bortezomib-adapted leukaemic cell lines demonstrated the retention of this proteasome target and subsite selectivity as well as potency even in clinically relevant cell lines. Our studies, therefore, propose that further development of SylA might indeed result in an improved small molecule for the treatment of leukaemia.

The Dynamics of an Impulsive Predator-Prey System with Stage Structure and Holling Type III Functional Response

Directory of Open Access Journals (Sweden)

Zhixiang Ju

2015-01-01

Full Text Available Based on the biological resource management of natural resources, a stage-structured predator-prey model with Holling type III functional response, birth pulse, and impulsive harvesting at different moments is proposed in this paper. By applying comparison theorem and some analysis techniques, the global attractivity of predator-extinction periodic solution and the permanence of this system are studied. At last, examples and numerical simulations are given to verify the validity of the main results.

Biosafety of Recombinant and Wild Type Nucleopolyhedroviruses as Bioinsecticides

Directory of Open Access Journals (Sweden)

Bruce D. Hammock

2007-06-01

Full Text Available The entomopathogenic Autographa californica (Speyer nucleopolyhedrovirus (AcMNPV has been genetically modified to increase its speed of kill. The potential adverse effects of a recombinant AcMNPV (AcAaIT as well as wild type AcMNPV and wild type Spodoptera littoralis NPV (SlNPV were studied. Cotton plants were treated with these viruses at concentrations that were adjusted to resemble the recommended field application rate (4 x 1012 PIBs/feddan, feddan = 4,200 m2 and 3rd instar larvae of S. littoralis were allowed to feed on the contaminated plants. SDS-PAGE, ELISA, and DNA analyses were used to confirm that larvae that fed on these plants were virus-infected. Polyhedra that were purified from the infected larvae were subjected to structural protein analysis. A 32 KDa protein was found in polyhedra that were isolated from all of the viruses. Subtle differences were found in the size and abundance of ODV proteins. Antisera against polyhedral proteins isolated from AcAaIT polyhedra were raised in rabbits. The terminal bleeds from rabbits were screened against four coating antigens (i.e., polyhedral proteins from AcAaIT, AcAaIT from field-infected larvae (AcAaIT-field, AcMNPV, and SlNPV using a two-dimensional titration method with the coated antigen format. Competitive inhibition experiments were conducted in parallel to optimize antibody and coating antigen concentrations for ELISA. The IC50 values for each combination ranged from 1.42 to 163 μg/ml. AcAaIT-derived polyhedrin gave the lowest IC50 value, followed by those of SlNPV, AcAaIT-field, and AcMNPV. The optimized ELISA system showed low cross reactivity for AcMNPV (0.87%, AcAaIT-field (1.2%, and SlNPV (4.0%. Genomic DNAs isolated from AcAaIT that were passaged in larvae of S. littoralis that were reared in the laboratory or field did not show any detectable differences. Albino rats (male and female that were treated with AcAaIT, AcMNPV or SlNPV (either orally or by intraperitoneal

Effects of social stress and intrauterine position on sexual phenotype in wild-type house mice (Mus musculus)

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William J. Zielinski; John G. Vandenbergh; Monica M. Montano

1991-01-01

Wild-type house mice were used to test the effect of intrauterine position on anogenital distance (AGD) and to verify whether crowding stress would masculinize female pups, developing at all intrauterine positions, as has been demonstrated in CF-1 mice stressed by restraint, heat, and...

Ceramic materials on perovskite-type structure for electronic applications

International Nuclear Information System (INIS)

Surowiak, Z.

2003-01-01

Ceramic materials exhibiting the perovskite-type structure constitute among others, resource base for many fields of widely understood electronics (i.e., piezoelectronics, accustoelectronics, optoelectronics, computer science, tele- and radioelectronics etc.). Most often they are used for fabrication of different type sensors (detectors), transducers, ferroelectric memories, limiters of the electronic current intensity, etc., and hence they are numbered among so-called intelligent materials. Prototype structure of this group of materials is the structure of the mineral called perovskite (CaTiO 3 ). By means of right choice of the chemical composition of ABO 3 and deforming the regular perovskite structure (m3m) more than 5000 different chemical compounds and solid solutions exhibiting the perovskite-type structure have been fabricated. The concept of perovskite functional ceramics among often things ferroelectric ceramics, pyroelectric ceramics, piezoelectric ceramics, electrostrictive ceramics, posistor ceramics, superconductive ceramics and ferromagnetic ceramics. New possibilities of application of the perovskite-type ceramics are opened by nanotechnology. (author)

Microbial mat ecosystems: Structure types, functional diversity, and biotechnological application

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Cristina M. Prieto-Barajas

2018-01-01

Full Text Available Microbial mats are horizontally stratified microbial communities, exhibiting a structure defined by physiochemical gradients, which models microbial diversity, physiological activities, and their dynamics as a whole system. These ecosystems are commonly associated with aquatic habitats, including hot springs, hypersaline ponds, and intertidal coastal zones and oligotrophic environments, all of them harbour phototrophic mats and other environments such as acidic hot springs or acid mine drainage harbour non-photosynthetic mats. This review analyses the complex structure, diversity, and interactions between the microorganisms that form the framework of different types of microbial mats located around the globe. Furthermore, the many tools that allow studying microbial mats in depth and their potential biotechnological applications are discussed.

Characterization of a sensitive mouse Aβ40 PD biomarker assay for Alzheimer's disease drug development in wild-type mice.

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Lu, Yanmei; Hoyte, Kwame; Montgomery, William H; Luk, Wilman; He, Dongping; Meilandt, William J; Zuchero, Y Joy Yu; Atwal, Jasvinder K; Scearce-Levie, Kimberly; Watts, Ryan J; DeForge, Laura E

2016-05-01

Transgenic mice that overexpress human amyloid precursor protein with Swedish or London (APPswe or APPlon) mutations have been widely used for preclinical Alzheimer's disease (AD) drug development. AD patients, however, rarely possess these mutations or overexpress APP. We developed a sensitive ELISA that specifically and accurately measures low levels of endogenous Aβ40 in mouse plasma, brain and CSF. In wild-type mice treated with a bispecific anti-TfR/BACE1 antibody, significant Aβ reductions were observed in the periphery and the brain. APPlon transgenic mice showed a slightly less reduction, whereas APPswe mice did not have any decrease. This sensitive and well-characterized mouse Aβ40 assay enables the use of wild-type mice for preclinical PK/PD and efficacy studies of potential AD therapeutics.

Testing projected wild bee distributions in agricultural habitats: predictive power depends on species traits and habitat type.

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Marshall, Leon; Carvalheiro, Luísa G; Aguirre-Gutiérrez, Jesús; Bos, Merijn; de Groot, G Arjen; Kleijn, David; Potts, Simon G; Reemer, Menno; Roberts, Stuart; Scheper, Jeroen; Biesmeijer, Jacobus C

2015-10-01

Species distribution models (SDM) are increasingly used to understand the factors that regulate variation in biodiversity patterns and to help plan conservation strategies. However, these models are rarely validated with independently collected data and it is unclear whether SDM performance is maintained across distinct habitats and for species with different functional traits. Highly mobile species, such as bees, can be particularly challenging to model. Here, we use independent sets of occurrence data collected systematically in several agricultural habitats to test how the predictive performance of SDMs for wild bee species depends on species traits, habitat type, and sampling technique. We used a species distribution modeling approach parametrized for the Netherlands, with presence records from 1990 to 2010 for 193 Dutch wild bees. For each species, we built a Maxent model based on 13 climate and landscape variables. We tested the predictive performance of the SDMs with independent datasets collected from orchards and arable fields across the Netherlands from 2010 to 2013, using transect surveys or pan traps. Model predictive performance depended on species traits and habitat type. Occurrence of bee species specialized in habitat and diet was better predicted than generalist bees. Predictions of habitat suitability were also more precise for habitats that are temporally more stable (orchards) than for habitats that suffer regular alterations (arable), particularly for small, solitary bees. As a conservation tool, SDMs are best suited to modeling rarer, specialist species than more generalist and will work best in long-term stable habitats. The variability of complex, short-term habitats is difficult to capture in such models and historical land use generally has low thematic resolution. To improve SDMs' usefulness, models require explanatory variables and collection data that include detailed landscape characteristics, for example, variability of crops and

Structural damage detection-oriented multi-type sensor placement with multi-objective optimization

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Lin, Jian-Fu; Xu, You-Lin; Law, Siu-Seong

2018-05-01

A structural damage detection-oriented multi-type sensor placement method with multi-objective optimization is developed in this study. The multi-type response covariance sensitivity-based damage detection method is first introduced. Two objective functions for optimal sensor placement are then introduced in terms of the response covariance sensitivity and the response independence. The multi-objective optimization problem is formed by using the two objective functions, and the non-dominated sorting genetic algorithm (NSGA)-II is adopted to find the solution for the optimal multi-type sensor placement to achieve the best structural damage detection. The proposed method is finally applied to a nine-bay three-dimensional frame structure. Numerical results show that the optimal multi-type sensor placement determined by the proposed method can avoid redundant sensors and provide satisfactory results for structural damage detection. The restriction on the number of each type of sensors in the optimization can reduce the searching space in the optimization to make the proposed method more effective. Moreover, how to select a most optimal sensor placement from the Pareto solutions via the utility function and the knee point method is demonstrated in the case study.

The dimer interface of the membrane type 1 matrix metalloproteinase hemopexin domain: crystal structure and biological functions.

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Tochowicz, Anna; Goettig, Peter; Evans, Richard; Visse, Robert; Shitomi, Yasuyuki; Palmisano, Ralf; Ito, Noriko; Richter, Klaus; Maskos, Klaus; Franke, Daniel; Svergun, Dmitri; Nagase, Hideaki; Bode, Wolfram; Itoh, Yoshifumi

2011-03-04

Homodimerization is an essential step for membrane type 1 matrix metalloproteinase (MT1-MMP) to activate proMMP-2 and to degrade collagen on the cell surface. To uncover the molecular basis of the hemopexin (Hpx) domain-driven dimerization of MT1-MMP, a crystal structure of the Hpx domain was solved at 1.7 Å resolution. Two interactions were identified as potential biological dimer interfaces in the crystal structure, and mutagenesis studies revealed that the biological dimer possesses a symmetrical interaction where blades II and III of molecule A interact with blades III and II of molecule B. The mutations of amino acids involved in the interaction weakened the dimer interaction of Hpx domains in solution, and incorporation of these mutations into the full-length enzyme significantly inhibited dimer-dependent functions on the cell surface, including proMMP-2 activation, collagen degradation, and invasion into the three-dimensional collagen matrix, whereas dimer-independent functions, including gelatin film degradation and two-dimensional cell migration, were not affected. These results shed light on the structural basis of MT1-MMP dimerization that is crucial to promote cellular invasion.

Nuclear inclusion bodies of mutant and wild-type p53 in cancer: a hallmark of p53 inactivation and proteostasis remodelling by p53 aggregation.

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De Smet, Frederik; Saiz Rubio, Mirian; Hompes, Daphne; Naus, Evelyne; De Baets, Greet; Langenberg, Tobias; Hipp, Mark S; Houben, Bert; Claes, Filip; Charbonneau, Sarah; Delgado Blanco, Javier; Plaisance, Stephane; Ramkissoon, Shakti; Ramkissoon, Lori; Simons, Colinda; van den Brandt, Piet; Weijenberg, Matty; Van England, Manon; Lambrechts, Sandrina; Amant, Frederic; D'Hoore, André; Ligon, Keith L; Sagaert, Xavier; Schymkowitz, Joost; Rousseau, Frederic

2017-05-01

Although p53 protein aggregates have been observed in cancer cell lines and tumour tissue, their impact in cancer remains largely unknown. Here, we extensively screened for p53 aggregation phenotypes in tumour biopsies, and identified nuclear inclusion bodies (nIBs) of transcriptionally inactive mutant or wild-type p53 as the most frequent aggregation-like phenotype across six different cancer types. p53-positive nIBs co-stained with nuclear aggregation markers, and shared molecular hallmarks of nIBs commonly found in neurodegenerative disorders. In cell culture, tumour-associated stress was a strong inducer of p53 aggregation and nIB formation. This was most prominent for mutant p53, but could also be observed in wild-type p53 cell lines, for which nIB formation correlated with the loss of p53's transcriptional activity. Importantly, protein aggregation also fuelled the dysregulation of the proteostasis network in the tumour cell by inducing a hyperactivated, oncogenic heat-shock response, to which tumours are commonly addicted, and by overloading the proteasomal degradation system, an observation that was most pronounced for structurally destabilized mutant p53. Patients showing tumours with p53-positive nIBs suffered from a poor clinical outcome, similar to those with loss of p53 expression, and tumour biopsies showed a differential proteostatic expression profile associated with p53-positive nIBs. p53-positive nIBs therefore highlight a malignant state of the tumour that results from the interplay between (1) the functional inactivation of p53 through mutation and/or aggregation, and (2) microenvironmental stress, a combination that catalyses proteostatic dysregulation. This study highlights several unexpected clinical, biological and therapeutically unexplored parallels between cancer and neurodegeneration. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great

Effects of temperature on the p53-DNA binding interactions and their dynamical behavior: comparing the wild type to the R248Q mutant.

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Khaled Barakat

Full Text Available BACKGROUND: The protein p53 plays an active role in the regulation of cell cycle. In about half of human cancers, the protein is inactivated by mutations located primarily in its DNA-binding domain. Interestingly, a number of these mutations possess temperature-induced DNA-binding characteristics. A striking example is the mutation of Arg248 into glutamine or tryptophan. These mutants are defective for binding to DNA at 310 K although they have been shown to bind specifically to several p53 response elements at sub-physiological temperatures (298-306 K. METHODOLOGY/PRINCIPAL FINDINGS: This important experimental finding motivated us to examine the effects of temperature on the structure and configuration of R248Q mutant and compare it to the wild type protein. Our aim is to determine how and where structural changes of mutant variants take place due to temperature changes. To answer these questions, we compared the mutant to the wild-type proteins from two different aspects. First, we investigated the systems at the atomistic level through their DNA-binding affinity, hydrogen bond networks and spatial distribution of water molecules. Next, we assessed changes in their long-lived conformational motions at the coarse-grained level through the collective dynamics of their side-chain and backbone atoms separately. CONCLUSIONS: The experimentally observed effect of temperature on the DNA-binding properties of p53 is reproduced. Analysis of atomistic and coarse-grained data reveal that changes in binding are determined by a few key residues and provide a rationale for the mutant-loss of binding at physiological temperatures. The findings can potentially enable a rescue strategy for the mutant structure.

BACHD rats expressing full-length mutant huntingtin exhibit differences in social behavior compared to wild-type littermates.

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Giuseppe Manfré

Full Text Available Huntington disease (HD is a devastating inherited neurodegenerative disorder characterized by progressive motor, cognitive, and psychiatric symptoms without any cure to slow down or stop the progress of the disease. The BACHD rat model for HD carrying the human full-length mutant huntingtin protein (mHTT with 97 polyQ repeats has been recently established as a promising model which reproduces several HD-like features. While motor and cognitive functions have been characterized in BACHD rats, little is known about their social phenotype.This study focuses especially on social behavior since evidence for social disturbances exists in human patients. Our objective was to compare social behavior in BACHD and wild-type (WT rats at different ages, using two different measures of sociability.Animals were tested longitudinally at the age of 2, 4 and 8 months in the social interaction test to examine different parameters of sociability. A separate cohort of 7 month old rats was tested in the three chamber social test to measure both sociability and social novelty. Gene expression analyses in 8 months old animals were performed by real time qRT-PCR to evaluate a potential involvement of D1 and D2 dopaminergic receptors and the contribution of Brain-derived neurotrophic factor (BDNF to the observed behavioral alterations.In the social interaction test, BACHD rats showed age-dependent changes in behaviour when they were-re introduced to their cagemate after a 24 hours-period of individual housing. The time spent on nape attacks increased with aging. Furthermore, a significant higher level of pinning at 2 months of age was shown in the BACHD rats compared to wild-types, followed by a reduction at 4 and 8 months. On the other hand, BACHD rats exhibited a decreased active social behaviour compared to wild-types, reflected by genotype-effects on approaching, following and social nose contact. In the three chamber social test, BACHD rats seemed to show a mild

BACHD rats expressing full-length mutant huntingtin exhibit differences in social behavior compared to wild-type littermates.

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Manfré, Giuseppe; Novati, Arianna; Faccini, Ilaria; Rossetti, Andrea C; Bosch, Kari; Molteni, Raffaella; Riva, Marco A; Van der Harst, Johanneke E; Nguyen, Huu Phuc; Homberg, Judith R

2018-01-01

Huntington disease (HD) is a devastating inherited neurodegenerative disorder characterized by progressive motor, cognitive, and psychiatric symptoms without any cure to slow down or stop the progress of the disease. The BACHD rat model for HD carrying the human full-length mutant huntingtin protein (mHTT) with 97 polyQ repeats has been recently established as a promising model which reproduces several HD-like features. While motor and cognitive functions have been characterized in BACHD rats, little is known about their social phenotype. This study focuses especially on social behavior since evidence for social disturbances exists in human patients. Our objective was to compare social behavior in BACHD and wild-type (WT) rats at different ages, using two different measures of sociability. Animals were tested longitudinally at the age of 2, 4 and 8 months in the social interaction test to examine different parameters of sociability. A separate cohort of 7 month old rats was tested in the three chamber social test to measure both sociability and social novelty. Gene expression analyses in 8 months old animals were performed by real time qRT-PCR to evaluate a potential involvement of D1 and D2 dopaminergic receptors and the contribution of Brain-derived neurotrophic factor (BDNF) to the observed behavioral alterations. In the social interaction test, BACHD rats showed age-dependent changes in behaviour when they were-re introduced to their cagemate after a 24 hours-period of individual housing. The time spent on nape attacks increased with aging. Furthermore, a significant higher level of pinning at 2 months of age was shown in the BACHD rats compared to wild-types, followed by a reduction at 4 and 8 months. On the other hand, BACHD rats exhibited a decreased active social behaviour compared to wild-types, reflected by genotype-effects on approaching, following and social nose contact. In the three chamber social test, BACHD rats seemed to show a mild deficit in

BACHD rats expressing full-length mutant huntingtin exhibit differences in social behavior compared to wild-type littermates

Science.gov (United States)

Manfré, Giuseppe; Novati, Arianna; Faccini, Ilaria; Rossetti, Andrea C.; Bosch, Kari; Molteni, Raffaella; Riva, Marco A.; Van der Harst, Johanneke E.; Homberg, Judith R.

2018-01-01

Background Huntington disease (HD) is a devastating inherited neurodegenerative disorder characterized by progressive motor, cognitive, and psychiatric symptoms without any cure to slow down or stop the progress of the disease. The BACHD rat model for HD carrying the human full-length mutant huntingtin protein (mHTT) with 97 polyQ repeats has been recently established as a promising model which reproduces several HD-like features. While motor and cognitive functions have been characterized in BACHD rats, little is known about their social phenotype. Objective This study focuses especially on social behavior since evidence for social disturbances exists in human patients. Our objective was to compare social behavior in BACHD and wild-type (WT) rats at different ages, using two different measures of sociability. Methods Animals were tested longitudinally at the age of 2, 4 and 8 months in the social interaction test to examine different parameters of sociability. A separate cohort of 7 month old rats was tested in the three chamber social test to measure both sociability and social novelty. Gene expression analyses in 8 months old animals were performed by real time qRT-PCR to evaluate a potential involvement of D1 and D2 dopaminergic receptors and the contribution of Brain-derived neurotrophic factor (BDNF) to the observed behavioral alterations. Results In the social interaction test, BACHD rats showed age-dependent changes in behaviour when they were-re introduced to their cagemate after a 24 hours-period of individual housing. The time spent on nape attacks increased with aging. Furthermore, a significant higher level of pinning at 2 months of age was shown in the BACHD rats compared to wild-types, followed by a reduction at 4 and 8 months. On the other hand, BACHD rats exhibited a decreased active social behaviour compared to wild-types, reflected by genotype-effects on approaching, following and social nose contact. In the three chamber social test, BACHD rats

The dead, hardened floral bracts of dispersal units of wild wheat function as storage for active hydrolases and in enhancing seedling vigor.

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Buzi Raviv

Full Text Available It is commonly assumed that the dead, hardened floral bracts of the dispersal unit of grasses have been evolved to protect seeds from predation and / or assist in fruit/caryopsis dispersal. While these structures have important agronomical and economical implications, their adaptive value has not been fully explored. We investigated the hypothesis that the maternally derived hardened floral bracts have been evolved not just as a means for caryopsis protection and dispersal, but also as storage for substances that might affect seed germination and seedling vigor. Dead glumes as well as lemmas and paleas of wild emmer wheat (Triticum turgidum var dicoccoides were found to store and release upon hydration active hydrolases including nucleases and chitinases. High nuclease activity was released upon hydration from glumes derived from wild strains of wheat including Triticum urartu and wild emmer wheat, while very low nuclease activity was detected in glumes derived from domesticated, free-threshing wheat cultivars (e.g., durum wheat. Germination from the intact dispersal unit of wild emmer wheat was delayed, but post germination growth was better than those of separated caryopses. Most notable was a significant increase in lateral root production on seedlings germinated from the intact dispersal unit. Proteome analysis of wild emmer wheat glumes revealed many proteins stored and released upon hydration including S1-type nucleases, peptidases, antifungal hydrolases such as chitinases and β-1,3-glucanase as well as pectin acetylesterase, a protein involved in cell wall degradation and remodeling. Also, reactive oxygen species (ROS-detoxifying enzymes such as superoxide dismutase and ascorbate peroxidase were overrepresented in dead glumes of wild emmer wheat. Thus our study highlighted previously unknown features of the dispersal unit in wild wheat in which the dead, hardened floral bracts enclosing the caryopsis store active hydrolases and

A Caenorhabditis elegans wild type defies the temperature-size rule owing to a single nucleotide polymorphism in tra-3.

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Jan E Kammenga

2007-03-01

Full Text Available Ectotherms rely for their body heat on surrounding temperatures. A key question in biology is why most ectotherms mature at a larger size at lower temperatures, a phenomenon known as the temperature-size rule. Since temperature affects virtually all processes in a living organism, current theories to explain this phenomenon are diverse and complex and assert often from opposing assumptions. Although widely studied, the molecular genetic control of the temperature-size rule is unknown. We found that the Caenorhabditis elegans wild-type N2 complied with the temperature-size rule, whereas wild-type CB4856 defied it. Using a candidate gene approach based on an N2 x CB4856 recombinant inbred panel in combination with mutant analysis, complementation, and transgenic studies, we show that a single nucleotide polymorphism in tra-3 leads to mutation F96L in the encoded calpain-like protease. This mutation attenuates the ability of CB4856 to grow larger at low temperature. Homology modelling predicts that F96L reduces TRA-3 activity by destabilizing the DII-A domain. The data show that size adaptation of ectotherms to temperature changes may be less complex than previously thought because a subtle wild-type polymorphism modulates the temperature responsiveness of body size. These findings provide a novel step toward the molecular understanding of the temperature-size rule, which has puzzled biologists for decades.

Suitable Methods in Spatial Pattern Analysis of Heterogeneous Wild Pistachio (Pistacia atlantica Desf. Woodlands in Zagros, Iran

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Y. Erfanifard

2014-12-01

Full Text Available Spatial pattern of trees in forests reveals how trees interact with each other and their environment. Spatial structure of trees in forest ecosystems is affected by environmental heterogeneity that leads to their heterogeneous distribution. This study was aimed to investigate the appropriate methods to analyze spatial pattern of heterogeneous wild pistachio woodlands in Zagros, Iran. A 40-ha pure stand of wild pistachio trees (Pistacia atlantica Desf. was selected in Wild Pistachio Research Forest in Fars Province for this purpose. The Kolmogrov-Smirnov test of goodness-of-fit of inhomogeneous Poisson point process showed that the distribution of wild pistachio trees was significantly heterogeneous (α=0.05. Inhomogeneous Ripley's K-, L-, and G-functions were applied beside their homogeneous forms. Inhomogeneous Ripley's K- and L-functions showed that wild pistachio trees were primarily clumped and dispersedly distributed thereafter, while g(r not only showed these results but also well expressed the detailed changes in spatial scale. The results of inappropriate homogeneous functions in the study area showed that all three functions expressed the primary clumping of the trees more than it was and their dispersed pattern as clumped. In general, it was concluded that inhomogeneous functions should be applied to analyze the spatial pattern of heterogeneous wild pistachio trees in the study area and it is recommended to develop g(r applications due to its more detailed information

Impairment of Vision in a Mouse Model of Usher Syndrome Type III.

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Tian, Guilian; Lee, Richard; Ropelewski, Philip; Imanishi, Yoshikazu

2016-03-01

The purpose of this study was to obtain an Usher syndrome type III mouse model with retinal phenotype. Speed congenic method was used to obtain Clrn1 exon 1 knockout (Clrn1-/-) and Clrn1N48K knockin (Clrn1N48K/N48K) mice under A/J background. To study the retinal functions of these mice, we measured scotopic and photopic ERG responses. To observe if there are any structural abnormalities, we conducted light and transmission electron microscopy of fixed retinal specimens. In 3-month-old Clrn1-/- mice, scotopic b-wave amplitude was reduced by more than 25% at the light intensities from -2.2 to 0.38 log cd·s/m2, but scotopic a-wave amplitudes were comparable to those of age-matched wild type mice at all the light intensities tested. In 9-month-old Clrn1-/- mice, scotopic b-wave amplitudes were further reduced by more than 35%, and scotopic a-wave amplitude also showed a small decline as compared with wild type mice. Photopic ERG responses were comparable between Clrn1-/- and wild type mice. Those electrophysiological defects were not associated with a loss of rods. In Clrn1N48K/N48K mice, both a- and b-wave amplitudes were not discernable from those of wild type mice aged up to 10 months. Mutations that are Clrn1-/- biallelic cause visual defects when placed under A/J background. The absence of apparent rod degeneration suggests that the observed phenotype is due to functional defects, and not due to loss of rods. Biallelic Clrn1N48K/N48K mutations did not cause discernible visual defects, suggesting that Clrn1- allele is more severely dysfunctional than ClrnN48K allele.

Peramivir analogues bearing hydrophilic side chains exhibit higher activities against H275Y mutant than wild-type influenza virus.

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Chiu, Din-Chi; Lin, Tzu-Chen; Huang, Wen-I; Cheng, Ting-Jen; Tsai, Keng-Chang; Fang, Jim-Min

2017-11-29

Peramivir is an effective anti-influenza drug in the clinical treatment of influenza, but its efficacy toward the H275Y mutant is reduced. The previously reported cocrystal structures of inhibitors in the mutant neuraminidase (NA) suggest that the hydrophobic side chain should be at the origin of reduced binding affinity. In contrast, zanamivir having a hydrophilic glycerol side chain still possesses high affinity toward the H275Y NA. We thus designed five peramivir analogues (5-9) carrying hydrophilic glycol or glycerol side chains, and evaluated their roles in anti-influenza activity, especially for the H275Y mutant. The synthetic sequence involves a key step of (3 + 2) cycloaddition reactions between alkenes and nitrile oxides to construct the scaffold of peramivir carrying the desired hydrophilic side chains and other appropriate functional groups. The molecular docking experiments reveal that the hydrophilic side chain can provide extra hydrogen bonding with the translocated Glu-276 residue in the H275Y NA active site. Thus, the H275Y mutant may be even more sensitive than wild-type virus toward the peramivir analogues bearing hydrophilic side chains. Notably, the peramivir analogue bearing a glycerol side chain inhibits the H275Y mutant with an IC 50 value of 35 nM, which is better than the WSN virus by 9 fold.

Control of acute, chronic, and constitutive hyperammonemia by wild-type and genetically engineered Lactobacillus plantarum in rodents.

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Nicaise, Charles; Prozzi, Deborah; Viaene, Eric; Moreno, Christophe; Gustot, Thierry; Quertinmont, Eric; Demetter, Pieter; Suain, Valérie; Goffin, Philippe; Devière, Jacques; Hols, Pascal

2008-10-01

Hyperammonemia is a common complication of acute and chronic liver diseases. Often accompanied with side effects, therapeutic interventions such as antibiotics or lactulose are generally targeted to decrease the intestinal production and absorption of ammonia. In this study, we aimed to modulate hyperammonemia in three rodent models by administration of wild-type Lactobacillus plantarum, a genetically engineered ammonia hyperconsuming strain, and a strain deficient for the ammonia transporter. Wild-type and metabolically engineered L. plantarum strains were administered in ornithine transcarbamoylase-deficient Sparse-fur mice, a model of constitutive hyperammonemia, in a carbon tetrachloride rat model of chronic liver insufficiency and in a thioacetamide-induced acute liver failure mice model. Constitutive hyperammonemia in Sparse-fur mice and hyperammonemia in a rat model of chronic hepatic insufficiency were efficiently decreased by Lactobacillus administration. In a murine thioacetamide-induced model of acute liver failure, administration of probiotics significantly increased survival and decreased blood and fecal ammonia. The ammonia hyperconsuming strain exhibited a beneficial effect at a lower dose than its wild-type counterpart. Improved survival in the acute liver failure mice model was associated with lower blood ammonia levels but also with a decrease of astrocyte swelling in the brain cortex. Modulation of ammonia was abolished after administration of the strain deficient in the ammonium transporter. Intestinal pH was clearly lowered for all strains and no changes in gut flora were observed. Hyperammonemia in constitutive model or after acute or chronic induced liver failure can be controlled by the administration of L. plantarum with a significant effect on survival. The mechanism involved in this ammonia decrease implicates direct ammonia consumption in the gut.

General anesthetic octanol and related compounds activate wild-type and delF508 cystic fibrosis chloride channels.

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Marcet, Brice; Becq, Frédéric; Norez, Caroline; Delmas, Patrick; Verrier, Bernard

2004-03-01

1. Cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel is defective during cystic fibrosis (CF). Activators of the CFTR Cl(-) channel may be useful for therapy of CF. Here, we demonstrate that a range of general anesthetics like normal-alkanols (n-alkanols) and related compounds can stimulate the Cl(-) channel activity of wild-type CFTR and delF508-CFTR mutant. 2. The effects of n-alkanols like octanol on CFTR activity were measured by iodide ((125)I) efflux and patch-clamp techniques on three distinct cellular models: (1). CFTR-expressing Chinese hamster ovary cells, (2). human airway Calu-3 epithelial cells and (3). human airway JME/CF15 epithelial cells which express the delF508-CFTR mutant. 3. Our data show for the first time that n-alkanols activate both wild-type CFTR and delF508-CFTR mutant. Octanol stimulated (125)I efflux in a dose-dependent manner in CFTR-expressing cells (wild-type and delF508) but not in cell lines lacking CFTR. (125)I efflux and Cl(-) currents induced by octanol were blocked by glibenclamide but insensitive to 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid, as expected for a CFTR Cl(-) current. 4. CFTR activation by octanol was neither due to cell-to-cell uncoupling properties of octanol nor to an intracellular cAMP increase. CFTR activation by octanol requires phosphorylation by protein kinase-A (PKA) since it was prevented by H-89, a PKA inhibitor. 5. n-Alkanols chain length was an important determinant for channel activation, with rank order of potencies: 1-heptanoloctanoloctanol<1-decanol. Our findings may be of valuable interest for developing novel therapeutic strategies for CF.

Systematic strain construction and process development: Xylitol production by Saccharomyces cerevisiae expressing Candida tenuis xylose reductase in wild-type or mutant form.

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Pratter, S M; Eixelsberger, T; Nidetzky, B

2015-12-01

A novel Saccharomyces cerevisiae whole-cell biocatalyst for xylitol production based on Candida tenuis xylose reductase (CtXR) is presented. Six recombinant strains expressing wild-type CtXR or an NADH-specific mutant were constructed and evaluated regarding effects of expression mode, promoter strength, biocatalyst concentration and medium composition. Intracellular XR activities ranged from 0.09 U mgProt(-1) to 1.05 U mgProt(-1) but did not correlate with the strains' xylitol productivities, indicating that other factors limited xylose conversion in the high-activity strains. The CtXR mutant decreased the biocatalyst's performance, suggesting use of the NADPH-preferring wild-type enzyme when (semi-)aerobic conditions are applied. In a bioreactor process, the best-performing strain converted 40 g L(-1) xylose with an initial productivity of 1.16 g L(-1)h(-1) and a xylitol yield of 100%. The obtained results underline the potential of CtXR wild-type for xylose reduction and point out parameters to improve "green" xylitol production. Copyright © 2015 Elsevier Ltd. All rights reserved.

Dietary Calcium and Dairy Modulation of Oxidative Stress and Mortality in aP2-Agouti and Wild-type Mice

Directory of Open Access Journals (Sweden)

Antje Bruckbauer

2009-08-01

Full Text Available Oxidative and inflammatory stress have been implicated as major contributors to the aging process. Dietary Ca reduced both factors in short-term interventions, while milk exerted a greater effect than supplemental Ca. In this work, we examined the effects of life-long supplemental and dairy calcium on lifespan and life-span related biomarkers in aP2-agouti transgenic (model of diet-induced obesity and wild-type mice fed obesigenic diets until their death. These data demonstrate that dairy Ca exerts sustained effects resulting in attenuated adiposity, protection against age-related muscle loss and reduction of oxidative and inflammatory stress in both mouse strains. Although these effects did not alter maximum lifespan, they did suppress early mortality in wild-type mice, but not in aP2-agouti transgenic mice.

Bioactivities and Health Benefits of Wild Fruits

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Ya Li

2016-08-01

Full Text Available Wild fruits are exotic or underutilized. Wild fruits contain many bioactive compounds, such as anthocyanins and flavonoids. Many studies have shown that wild fruits possess various bioactivities and health benefits, such as free radical scavenging, antioxidant, anti-inflammatory, antimicrobial, and anticancer activity. Therefore, wild fruits have the potential to be developed into functional foods or pharmaceuticals to prevent and treat several chronic diseases. In the present article, we review current knowledge about the bioactivities and health benefits of wild fruits, which is valuable for the exploitation and utilization of wild fruits.

Lymphotropism and host responses during acute wild-type canine distemper virus infections in a highly susceptible natural host

DEFF Research Database (Denmark)

Nielsen, Line; Søgaard, Mette; Jensen, Trine Hammer

2009-01-01

The mechanisms behind the in vivo virulence of immunosuppressive wild-type Morbillivirus infections are still not fully understood. To investigate lymphotropism and host responses we have selected the natural host model of canine distemper virus (CDV) infection in mink. This model displays...

Effect of deuterium on the circadian period and metabolism in wild-type and tau mutant Syrian hamsters

NARCIS (Netherlands)

Oklejewicz, M; Hut, RA; Daan, S

2000-01-01

Homozygous tau mutant Syrian hamsters (tau-/-) have a free-running circadian period (tau) around 20 h and a proportionally higher metabolic rate compared with wild-type hamsters (tau+/+) with a period of circa 24 h. In this study, we applied deuterium oxide (D2O) to hamsters to test whether

Cadmium tolerance, cysteine and thiol peptide levels in wild type and chromium-tolerant strains of Scenedesmus acutus (Chlorophyceae)

Energy Technology Data Exchange (ETDEWEB)

Torricelli, Elena; Gorbi, Gessica; Pawlik-Skowronska, Barbara; Di Toppi, Luigi Sanita; Corradi, Maria Grazia

2004-07-14

Two strains of the unicellular green alga Scenedesmus acutus with different sensitivity to hexavalent chromium were compared for their tolerance of cadmium, by means of growth and recovery tests, and determination of cysteine, reduced glutathione and phytochelatin content, after short-term exposure to various cadmium concentrations (from 1.125 to 27 {mu}M). Growth experiments showed that, after 7-day treatments with cadmium, the chromium-tolerant strain reached a significantly higher cell density and, after 24-h exposure to Cd, was able to resume growth significantly better than the wild type. Constitutive level of cysteine was higher in the chromium-tolerant strain, while glutathione levels were similar in the two strains. The higher content of cysteine and the maintenance of both reduced glutathione and phytochelatin high levels in the presence of cadmium, support the higher cadmium co-tolerance of the chromium-tolerant strain in comparison with the wild type one.

Cadmium tolerance, cysteine and thiol peptide levels in wild type and chromium-tolerant strains of Scenedesmus acutus (Chlorophyceae)

International Nuclear Information System (INIS)

Torricelli, Elena; Gorbi, Gessica; Pawlik-Skowronska, Barbara; Di Toppi, Luigi Sanita; Corradi, Maria Grazia

2004-01-01

Two strains of the unicellular green alga Scenedesmus acutus with different sensitivity to hexavalent chromium were compared for their tolerance of cadmium, by means of growth and recovery tests, and determination of cysteine, reduced glutathione and phytochelatin content, after short-term exposure to various cadmium concentrations (from 1.125 to 27 μM). Growth experiments showed that, after 7-day treatments with cadmium, the chromium-tolerant strain reached a significantly higher cell density and, after 24-h exposure to Cd, was able to resume growth significantly better than the wild type. Constitutive level of cysteine was higher in the chromium-tolerant strain, while glutathione levels were similar in the two strains. The higher content of cysteine and the maintenance of both reduced glutathione and phytochelatin high levels in the presence of cadmium, support the higher cadmium co-tolerance of the chromium-tolerant strain in comparison with the wild type one

Crystal structure of the second fibronectin type III (FN3) domain from human collagen α1 type XX.

Science.gov (United States)

Zhao, Jingfeng; Ren, Jixia; Wang, Nan; Cheng, Zhong; Yang, Runmei; Lin, Gen; Guo, Yi; Cai, Dayong; Xie, Yong; Zhao, Xiaohong

2017-12-01

Collagen α1 type XX, which contains fibronectin type III (FN3) repeats involving six FN3 domains (referred to as the FN#1-FN#6 domains), is an unusual member of the fibril-associated collagens with interrupted triple helices (FACIT) subfamily of collagens. The results of standard protein BLAST suggest that the FN3 repeats might contribute to collagen α1 type XX acting as a cytokine receptor. To date, solution NMR structures of the FN#3, FN#4 and FN#6 domains have been determined. To obtain further structural evidence to understand the relationship between the structure and function of the FN3 repeats from collagen α1 type XX, the crystal structure of the FN#2 domain from human collagen α1 type XX (residues Pro386-Pro466; referred to as FN2-HCXX) was solved at 2.5 Å resolution. The crystal structure of FN2-HCXX shows an immunoglobulin-like fold containing a β-sandwich structure, which is formed by a three-stranded β-sheet (β1, β2 and β5) packed onto a four-stranded β-sheet (β3, β4, β6 and β7). Two consensus domains, tencon and fibcon, are structural analogues of FN2-HCXX. Fn8, an FN3 domain from human oncofoetal fibronectin, is the closest structural analogue of FN2-HCXX derived from a naturally occurring sequence. Based solely on the structural similarity of FN2-HCXX to other FN3 domains, the detailed functions of FN2-HCXX and the FN3 repeats in collagen α1 type XX cannot be identified.

Reverse engineering of wörner type drilling machine structure.

Science.gov (United States)

Wibowo, A.; Belly, I.; llhamsyah, R.; Indrawanto; Yuwana, Y.

2018-03-01

A product design needs to be modified based on the conditions of production facilities and existing resource capabilities without reducing the functional aspects of the product itself. This paper describes the reverse engineering process of the main structure of the wörner type drilling machine to obtain a machine structure design that can be made by resources with limited ability by using simple processes. Some structural, functional and the work mechanism analyzes have been performed to understand the function and role of each basic components. The process of dismantling of the drilling machine and measuring each of the basic components was performed to obtain sets of the geometry and size data of each component. The geometric model of each structure components and the machine assembly were built to facilitate the simulation process and machine performance analysis that refers to ISO standard of drilling machine. The tolerance stackup analysis also performed to determine the type and value of geometrical and dimensional tolerances, which could affect the ease of the components to be manufactured and assembled

Role of wild-type p53 in apoptotic and non-apoptotic cell death induced by X-irradiation and heat treatment in p53-mutated mouse M10 cells

International Nuclear Information System (INIS)

Ito, Atsushi; Nakano, Hisako; Shinohara, Kunio

2010-01-01

The sensitizing effects of wild-type p53 on X-ray-induced cell death and on heat-induced apoptosis in M10, a radiosensitive and Trp53 (mouse p53 gene)-mutated lymphoma cell line which dies through necrosis by X-irradiation, were investigated using three M10 derived transfectants with wild-type TP53 (human p53 gene). Cell death was determined by colony formation and/or dye exclusion test, and apoptosis was detected as the changes in nuclear morphology by Giemsa staining. Expression of wild-type p53 protein increased radiosensitivity of cell death as determined by both clonogenic and dye exclusion assays. This increase in radiosensitivity was attributable largely to apoptosis induction in addition to a small enhancement of necrosis. Interestingly neither pathway to cell death was accompanied by caspase-3 activation. On the other hand, heat-induced caspase-3 dependent apoptotic cell death without transfection was further increased by the transfection of wild-type p53. In conclusion, the introduction of wild-type p53 enhanced apoptotic cell death by X-rays or heat via different mechanisms that do or do not activate caspase-3, respectively. In addition, p53 also enhanced the X-ray-induced necrosis in M10 cells. (author)

Learning Discloses Abnormal Structural and Functional Plasticity at Hippocampal Synapses in the APP23 Mouse Model of Alzheimer's Disease

Science.gov (United States)

Middei, Silvia; Roberto, Anna; Berretta, Nicola; Panico, Maria Beatrice; Lista, Simone; Bernardi, Giorgio; Mercuri, Nicola B.; Ammassari-Teule, Martine; Nistico, Robert

2010-01-01

B6-Tg/Thy1APP23Sdz (APP23) mutant mice exhibit neurohistological hallmarks of Alzheimer's disease but show intact basal hippocampal neurotransmission and synaptic plasticity. Here, we examine whether spatial learning differently modifies the structural and electrophysiological properties of hippocampal synapses in APP23 and wild-type mice. While…

Wave function analysis of type-II self-assembled quantum dot structures using magneto-optics

International Nuclear Information System (INIS)

Godoy, Marcio Peron Franco de; Nakaema, Marcelo K.K.; Gomes, Paulo F.; Iikawa, Fernando; Brasil, Maria Jose S.P.; Bortoleto, Jose Roberto R.; Cotta, Monica A.; Ribeiro, Evaldo; Medeiros-Ribeiro, Gilberto; Marques, Gilmar E.; Bittencourt, A.C.R.

2004-01-01

Full text: Recently, self-assembled quantum dots have attracted considerable attention for their potential for device applications. Type II interface, in particular, present interesting properties due to the space separation of the carriers. One of the carriers is confined at the lower band gap layer and the other remains at the barrier layers and is only localized by the Coulomb attraction. An essential information for using type II quantum wells and quantum dots on technological applications is the localization of the carrier wave function, which is an experimentally difficult parameter to be measured. Some techniques have been proposed to map the wave functions in quantum dots such as magneto-tunneling spectroscopy and near- field scanning optical microscopy. These techniques involve however a very complex experimental apparatus and sample processing. The magneto-exciton transition can be used as an alternative tool to investigate the exciton wave function distribution, since this distribution has a strong influence on the diamagnetic shift and Zeeman splitting. In this work, we present magneto-optical studies of In P/GaAs type II self-assembled quantum dots, where the electron is strongly confined at the In P, while the hole is weakly localized at the GaAs barrier due to the Coulombic attraction from the electrons. This scenery is very distinct from type I systems. The weaker hole confinement should alter the valence band mixing resulting in a different valence band contribution on the Zeeman splitting as compared to type I systems. Based on the results of the magneto-exciton emission from the wetting layer and from the individual dots, we obtained interesting results concerning the wave function distribution in our system. We discuss the localization of the hole wave function along the growth direction based on the measured Zeeman splitting and the in-plane wave function distribution, based on the observed diamagnetic shift. A remarkable result is that the

A speculated ribozyme site in the herpes simplex virus type 1 latency-associated transcript gene is not essential for a wild-type reactivation phenotype

Science.gov (United States)

Carpenter, Dale; Singh, Sukhpreet; Osorio, Nelson; Hsiang, Chinhui; Jiang, Xianzhi; Jin, Ling; Jones, Clinton; Wechsler, Steven L

2010-01-01

During herpes simplex virus-1 (HSV-1) latency in sensory neurons, LAT (latency-associated transcript) is the only abundantly expressed viral gene. LAT plays an important role in the HSV-1 latency-reactivation cycle, because LAT deletion mutants have a significantly decreased reactivation phenotype. Based solely on sequence analysis, it was speculated that LAT encodes a ribozyme that plays an important role in how LAT enhances the virus’ reactivation phenotype. Because LAT ribozyme activity has never been reported, we decided to test the converse hypothesis, namely, that this region of LAT does not encode a ribozyme function important for LAT’s ability to enhance the reactivation phenotype. We constructed a viral mutant (LAT-Rz) in which the speculated ribozyme consensus sequence was altered such that no ribozyme was encoded. We report here that LAT-Rz had a wild-type reactivation phenotype in mice, confirming the hypothesis that the speculated LAT ribozyme is not a dominant factor in stimulating the latency-reactivation cycle in mice. PMID:18982533

On the molecular basis of D-bifunctional protein deficiency type III.

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Maija L Mehtälä

Full Text Available Molecular basis of D-bifunctional protein (D-BP deficiency was studied with wild type and five disease-causing variants of 3R-hydroxyacyl-CoA dehydrogenase fragment of the human MFE-2 (multifunctional enzyme type 2 protein. Complementation analysis in vivo in yeast and in vitro enzyme kinetic and stability determinants as well as in silico stability and structural fluctuation calculations were correlated with clinical data of known patients. Despite variations not affecting the catalytic residues, enzyme kinetic performance (K(m, V(max and k(cat of the recombinant protein variants were compromised to a varying extent and this can be judged as the direct molecular cause for D-BP deficiency. Protein stability plays an additional role in producing non-functionality of MFE-2 in case structural variations affect cofactor or substrate binding sites. Structure-function considerations of the variant proteins matched well with the available data of the patients.

The oxidation of alkylaryl sulfides and benzo[b]thiophenes by Escherichia coli cells expressing wild-type and engineered styrene monooxygenase from Pseudomonas putida CA-3.

Science.gov (United States)

Nikodinovic-Runic, Jasmina; Coulombel, Lydie; Francuski, Djordje; Sharma, Narain D; Boyd, Derek R; Ferrall, Rory Moore O; O'Connor, Kevin E

2013-06-01

Nine different sulfur-containing compounds were biotransformed to the corresponding sulfoxides by Escherichia coli Bl21(DE3) cells expressing styrene monooxygenase (SMO) from Pseudomonas putida CA-3. Thioanisole was consumed at 83.3 μmoles min(-1) g cell dry weight(-1) resulting mainly in the formation of R-thioanisole sulfoxide with an enantiomeric excess (ee) value of 45 %. The rate of 2-methyl-, 2-chloro- and 2-bromo-thioanisole consumption was 2-fold lower than that of thioanisole. Surprisingly, the 2-methylthioanisole sulfoxide product had the opposite (S) configuration to that of the other 2-substituted thioanisole derivatives and had a higher ee value (84 %). The rate of oxidation of 4-substituted thioanisoles was higher than the corresponding 2-substituted substrates but the ee values of the products were consistently lower (10-23 %). The rate of benzo[b]thiophene and 2-methylbenzo[b]thiophene sulfoxidation was approximately 10-fold lower than that of thioanisole. The ee value of the benzo[b]thiophene sulfoxide could not be determined as the product racemized rapidly. E. coli cells expressing an engineered SMO (SMOeng R3-11) oxidised 2-substituted thioanisoles between 1.8- and 2.8-fold faster compared to cells expressing the wild-type enzyme. SMOeng R3-11 oxidised benzo[b]thiophene and 2-methylbenzo[b]thiophene 10.1 and 5.6 times faster that the wild-type enzyme. The stereospecificity of the reaction catalysed by SMOeng was unchanged from that of the wild type. Using the X-ray crystal structure of the P. putida S12 SMO, it was evident that the entrance of substrates into the SMO active site is limited by the binding pocket bottleneck formed by the side chains of Val-211 and Asn-46 carboxyamide group.

Extracellular enzyme activities during lignocellulose degradation by Streptomyces spp.: a comparative study of wild-type and genetically manipulated strains

International Nuclear Information System (INIS)

Ramachandra, M.; Crawford, D.L.; Pometto, A.L. III

1987-01-01

The wild-type ligninolytic actinomycete Streptomyces viridosporus T7A and two genetically manipulated strains with enhanced abilities to produce a water-soluble lignin degradation intermediate, an acid-precipitable polymeric lignin (APPL), were grown on lignocellulose in solid-state fermentation cultures. Culture filtrates were periodically collected, analyzed for APPL, and assayed for extracellular lignocellulose-catabolizing enzyme activities. Two APPL-overproducing strains, UV irradiation mutant T7A-81 and protoplast fusion recombinant SR-10, had higher and longer persisting peroxidase, esterase, and endoglucanase activities than did the wild-type strain T7A. Results implicated one or more of these enzymes in lignin solubilization. Only mutant T7A-81 had higher xylanase activity than the wild type. The peroxidase was induced by both lignocellulose and APPL. This extracellular enzyme has some similarities to previously described ligninases in fungi. This is the first report of such an enzyme in Streptomyces spp. Four peroxidase isozymes were present, and all catalyzed the oxidation of 3,4-dihydroxyphenylalanine, while one also catalyzed hydrogen peroxide-dependent oxidation of homoprotocatechuic acid and caffeic acid. Three constitutive esterase isozymes were produced which differed in substrate specificity toward α-naphthyl acetate and α-naphthyl butyrate. Three endoglucanase bands, which also exhibited a low level of xylanase activity, were identified on polyacrylamide gels as was one xylanase-specific band. There were no major differences in the isoenzymes produced by the different strains. The probable role of each enzyme in lignocellulose degradation is discussed

Neurodegenerative disease mutations in TREM2 reveal a functional surface and distinct loss-of-function mechanisms

Energy Technology Data Exchange (ETDEWEB)

Kober, Daniel L.; Alexander-Brett, Jennifer M.; Karch, Celeste M.; Cruchaga, Carlos; Colonna, Marco; Holtzman, Michael J.; Brett, Thomas J. (WU-MED)

2016-12-20

Genetic variations in the myeloid immune receptor TREM2 are linked to several neurodegenerative diseases. To determine how TREM2 variants contribute to these diseases, we performed structural and functional studies of wild-type and variant proteins. Our 3.1 Å TREM2 crystal structure revealed that mutations found in Nasu-Hakola disease are buried whereas Alzheimer’s disease risk variants are found on the surface, suggesting that these mutations have distinct effects on TREM2 function. Biophysical and cellular methods indicate that Nasu-Hakola mutations impact protein stability and decrease folded TREM2 surface expression, whereas Alzheimer’s risk variants impact binding to a TREM2 ligand. Additionally, the Alzheimer’s risk variants appear to epitope map a functional surface on TREM2 that is unique within the larger TREM family. These findings provide a guide to structural and functional differences among genetic variants of TREM2, indicating that therapies targeting the TREM2 pathway should be tailored to these genetic and functional differences with patient-specific medicine approaches for neurodegenerative disorders.

Characteristics of alpha/beta interferon induction after infection of murine fibroblasts with wild-type and mutant alphaviruses

International Nuclear Information System (INIS)

Burke, Crystal W.; Gardner, Christina L.; Steffan, Joshua J.; Ryman, Kate D.; Klimstra, William B.

2009-01-01

We examined the characteristics of interferon alpha/beta (IFN-α/β) induction after alphavirus or control Sendai virus (SeV) infection of murine fibroblasts (MEFs). As expected, SeV infection of wild-type (wt) MEFs resulted in strong dimerization of IRF3 and the production of high levels of IFN-α/β. In contrast, infection of MEFs with multiple alphaviruses failed to elicit detectable IFN-α/β. In more detailed studies, Sindbis virus (SINV) infection caused dimerization and nuclear migration of IRF3, but minimal IFN-β promoter activity, although surprisingly, the infected cells were competent for IFN production by other stimuli early after infection. A SINV mutant defective in host macromolecular synthesis shutoff induced IFN-α/β in the MEF cultures dependent upon the activities of the TBK1 IRF3 activating kinase and host pattern recognition receptors (PRRs) PKR and MDA5 but not RIG-I. These results suggest that wild-type alphaviruses antagonize IFN induction after IRF3 activation but also may avoid detection by host PRRs early after infection.

Structure and function of wood

Science.gov (United States)

Alex Wiedenhoeft

2010-01-01

Wood is a complex biological structure, a composite of many chemistries and cell types acting together to serve the needs of a living plant. Attempting to understand wood in the context of wood technology, we have often overlooked the key and basic fact that wood evolved over the course of millions of years to serve three main functions in plants― conduction of water...

GPER Mediates Functional Endothelial Aging in Renal Arteries.

Science.gov (United States)

Meyer, Matthias R; Rosemann, Thomas; Barton, Matthias; Prossnitz, Eric R

2017-01-01

Aging is associated with impaired renal artery function, which is partly characterized by arterial stiffening and a reduced vasodilatory capacity due to excessive generation of reactive oxygen species by NADPH oxidases (Nox). The abundance and activity of Nox depends on basal activity of the heptahelical transmembrane receptor GPER; however, whether GPER contributes to age-dependent functional changes in renal arteries is unknown. This study investigated the effect of aging and Nox activity on renal artery tone in wild-type and GPER-deficient (Gper-/-) mice (4 and 24 months old). In wild-type mice, aging markedly impaired endothelium-dependent, nitric oxide (NO)-mediated relaxations to acetylcholine, which were largely preserved in renal arteries of aged Gper-/- mice. The Nox inhibitor gp91ds-tat abolished this difference by greatly enhancing relaxations in wild-type mice, while having no effect in Gper-/- mice. Contractions to angiotensin II and phenylephrine in wild-type mice were partly sensitive to gp91ds-tat but unaffected by aging. Again, deletion of GPER abolished effects of Nox inhibition on contractile responses. In conclusion, basal activity of GPER is required for the age-dependent impairment of endothelium-dependent, NO-mediated relaxation in the renal artery. Restoration of relaxation by a Nox inhibitor in aged wild-type but not Gper-/- mice strongly supports a role for Nox-derived reactive oxygen species as the underlying cause. Pharmacological blockers of GPER signaling may thus be suitable to inhibit functional endothelial aging of renal arteries by reducing Nox-derived oxidative stress and, possibly, the associated age-dependent deterioration of kidney function. © 2017 S. Karger AG, Basel.

EvoWild: a demosimulator about wild life

Directory of Open Access Journals (Sweden)

Eduardo Palacio Gayoso

2008-12-01

Full Text Available During the last years we can see how AI (Artificial Intelligence is reappearing because of technological improvements. These improvements make possible the management of large groups of information with acceptable reply times.On the other hand, cost reductions in technology make possible that an investigation field like AI becomes to an inversion field closer to scale economies, that’s why it’ll be economically profitable to invert in this type of applications.One of the fastest consequences is the AI implantation in a big amount of devices of our environment, cell telephones, palms and of course, in the video game industry.This is the reason that took us to develop EvoWild, a simulation about wild life that has video game format and tools but at the same time implements AI algorithms like genetic algorithms and reasoning based in cases.

Comparative metabolic profiling of mce1 operon mutant vs wild-type Mycobacterium tuberculosis strains.

Science.gov (United States)

Queiroz, Adriano; Medina-Cleghorn, Daniel; Marjanovic, Olivera; Nomura, Daniel K; Riley, Lee W

2015-11-01

Mycobacterium tuberculosis disrupted in a 13-gene operon (mce1) accumulates free mycolic acids (FM) in its cell wall and causes accelerated death in mice. Here, to more comprehensively analyze differences in their cell wall lipid composition, we used an untargeted metabolomics approach to compare the lipid profiles of wild-type and mce1 operon mutant strains. By liquid chromatography-mass spectrometry, we identified >400 distinct lipids significantly altered in the mce1 mutant compared to wild type. These lipids included decreased levels of saccharolipids and glycerophospholipids, and increased levels of alpha-, methoxy- and keto mycolic acids (MA), and hydroxyphthioceranic acid. The mutant showed reduced expression of mmpL8, mmpL10, stf0, pks2 and papA2 genes involved in transport and metabolism of lipids recognized to induce proinflammatory response; these lipids were found to be decreased in the mutant. In contrast, the transcripts of mmpL3, fasI, kasA, kasB, acpM and RV3451 involved in MA transport and metabolism increased; MA inhibits inflammatory response in macrophages. Since the mce1 operon is known to be regulated in intracellular M. tuberculosis, we speculate that the differences we observed in cell wall lipid metabolism and composition may affect host response to M. tuberculosis infection and determine the clinical outcome of such an infection. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Beyond T and DHT - novel steroid derivatives capable of wild type androgen receptor activation.

Science.gov (United States)

Mostaghel, Elahe A

2014-01-01

While androgen deprivation therapy (ADT) remains the primary treatment for metastatic prostate cancer (PCa), castration does not eliminate androgens from the prostate tumor microenvironment, and residual intratumoral androgens are implicated in nearly every mechanism by which androgen receptor (AR)-mediated signaling promotes castration-resistant disease. The uptake and intratumoral (intracrine) conversion of circulating adrenal androgens such as dehydroepiandrosterone sulfate (DHEA-S) to steroids capable of activating the wild type AR is a recognized driver of castration resistant prostate cancer (CRPC). However, less well-characterized adrenal steroids, including 11-deoxcorticosterone (DOC) and 11beta-hydroxyandrostenedione (11OH-AED) may also play a previously unrecognized role in promoting AR activation. In particular, recent data demonstrate that the 5α-reduced metabolites of DOC and 11OH-AED are activators of the wild type AR. Given the well-recognized presence of SRD5A activity in CRPC tissue, these observations suggest that in the low androgen environment of CRPC, alternative sources of 5α-reduced ligands may supplement AR activation normally mediated by the canonical 5α-reduced agonist, 5α-DHT. Herein we review the emerging data that suggests a role for these alternative steroids of adrenal origin in activating the AR, and discuss the enzymatic pathways and novel downstream metabolites mediating these effects. We conclude by discussing the potential implications of these findings for CRPC progression, particularly in context of new agents such as abiraterone and enzalutamide which target the AR-axis for prostate cancer therapy.

Action potentials and ion conductances in wild-type and CALHM1-knockout type II taste cells

Science.gov (United States)

Saung, Wint Thu; Foskett, J. Kevin

2017-01-01

Taste bud type II cells fire action potentials in response to tastants, triggering nonvesicular ATP release to gustatory neurons via voltage-gated CALHM1-associated ion channels. Whereas CALHM1 regulates mouse cortical neuron excitability, its roles in regulating type II cell excitability are unknown. In this study, we compared membrane conductances and action potentials in single identified TRPM5-GFP-expressing circumvallate papillae type II cells acutely isolated from wild-type (WT) and Calhm1 knockout (KO) mice. The activation kinetics of large voltage-gated outward currents were accelerated in cells from Calhm1 KO mice, and their associated nonselective tail currents, previously shown to be highly correlated with ATP release, were completely absent in Calhm1 KO cells, suggesting that CALHM1 contributes to all of these currents. Calhm1 deletion did not significantly alter resting membrane potential or input resistance, the amplitudes and kinetics of Na+ currents either estimated from action potentials or recorded from steady-state voltage pulses, or action potential threshold, overshoot peak, afterhyperpolarization, and firing frequency. However, Calhm1 deletion reduced the half-widths of action potentials and accelerated the deactivation kinetics of transient outward currents, suggesting that the CALHM1-associated conductance becomes activated during the repolarization phase of action potentials. NEW & NOTEWORTHY CALHM1 is an essential ion channel component of the ATP neurotransmitter release mechanism in type II taste bud cells. Its contribution to type II cell resting membrane properties and excitability is unknown. Nonselective voltage-gated currents, previously associated with ATP release, were absent in cells lacking CALHM1. Calhm1 deletion was without effects on resting membrane properties or voltage-gated Na+ and K+ channels but contributed modestly to the kinetics of action potentials. PMID:28202574

Identification of enzymes and quantification of metabolic fluxes in the wild type and in a recombinant Aspergillus oryzae strain

DEFF Research Database (Denmark)

Pedersen, Henrik; Carlsen, Morten; Nielsen, Jens Bredal

1999-01-01

Two alpha-amylase-producing strains of Aspergillus oryzae, a wild-type strain and a recombinant containing additional copies of the alpha-amylase gene, were characterized,vith respect to enzyme activities, localization of enzymes to the mitochondria or cytosol, macromolecular composition...

Paternal spatial training enhances offspring's cognitive performance and synaptic plasticity in wild-type but not improve memory deficit in Alzheimer's mice.

Science.gov (United States)

Zhang, Shujuan; Li, Xiaoguang; Wang, Zhouyi; Liu, Yanchao; Gao, Yuan; Tan, Lu; Liu, Enjie; Zhou, Qiuzhi; Xu, Cheng; Wang, Xin; Liu, Gongping; Chen, Haote; Wang, Jian-Zhi

2017-05-08

Recent studies suggest that spatial training can maintain associative memory capacity in Tg2576 mice, but it is not known whether the beneficial effects can be inherited from the trained fathers to their offspring. Here, we exposed male wild-type and male 3XTg Alzheimer disease (AD) mice (3-m old) respectively to spatial training for one week and assessed the transgenerational effects in the F1 offspring when they were grown to 7-m old. We found that the paternal spatial training significantly enhanced progeny's spatial cognitive performance and synaptic transmission in wild-type mice. Among several synapse- or memory-associated proteins, we observed that the expression level of synaptotagmin 1 (SYT1) was significantly increased in the hippocampus of the paternally trained-offspring. Paternal training increased histone acetylation at the promoter of SYT1 in both fathers' and the offspring's hippocampus, and as well as in the fathers' sperm. Finally, paternal spatial training for one week did not improve memory and synaptic plasticity in 3XTg AD F1 offspring. Our findings suggest paternal spatial training for one week benefits the offspring's cognitive performance in wild-type mice with the mechanisms involving an enhanced transgenerational histone acetylation at SYT1 promoter.

Differential inhibition of ex-vivo tumor kinase activity by vemurafenib in BRAF(V600E and BRAF wild-type metastatic malignant melanoma.

Directory of Open Access Journals (Sweden)

Andliena Tahiri

Full Text Available Treatment of metastatic malignant melanoma patients harboring BRAF(V600E has improved drastically after the discovery of the BRAF inhibitor, vemurafenib. However, drug resistance is a recurring problem, and prognoses are still very bad for patients harboring BRAF wild-type. Better markers for targeted therapy are therefore urgently needed.In this study, we assessed the individual kinase activity profiles in 26 tumor samples obtained from patients with metastatic malignant melanoma using peptide arrays with 144 kinase substrates. In addition, we studied the overall ex-vivo inhibitory effects of vemurafenib and sunitinib on kinase activity status.Overall kinase activity was significantly higher in lysates from melanoma tumors compared to normal skin tissue. Furthermore, ex-vivo incubation with both vemurafenib and sunitinib caused significant decrease in phosphorylation of kinase substrates, i.e kinase activity. While basal phosphorylation profiles were similar in BRAF wild-type and BRAF(V600E tumors, analysis with ex-vivo vemurafenib treatment identified a subset of 40 kinase substrates showing stronger inhibition in BRAF(V600E tumor lysates, distinguishing the BRAF wild-type and BRAF(V600E tumors. Interestingly, a few BRAF wild-type tumors showed inhibition profiles similar to BRAF(V600E tumors. The kinase inhibitory effect of vemurafenib was subsequently analyzed in cell lines harboring different BRAF mutational status with various vemurafenib sensitivity in-vitro.Our findings suggest that multiplex kinase substrate array analysis give valuable information about overall tumor kinase activity. Furthermore, intra-assay exposure to kinase inhibiting drugs may provide a useful tool to study mechanisms of resistance, as well as to identify predictive markers.

Use of tissue-specific microRNA to control pathology of wild-type adenovirus without attenuation of its ability to kill cancer cells.

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Cawood, Ryan; Chen, Hannah H; Carroll, Fionnadh; Bazan-Peregrino, Miriam; van Rooijen, Nico; Seymour, Leonard W

2009-05-01

Replicating viruses have broad applications in biomedicine, notably in cancer virotherapy and in the design of attenuated vaccines; however, uncontrolled virus replication in vulnerable tissues can give pathology and often restricts the use of potent strains. Increased knowledge of tissue-selective microRNA expression now affords the possibility of engineering replicating viruses that are attenuated at the RNA level in sites of potential pathology, but retain wild-type replication activity at sites not expressing the relevant microRNA. To assess the usefulness of this approach for the DNA virus adenovirus, we have engineered a hepatocyte-safe wild-type adenovirus 5 (Ad5), which normally mediates significant toxicity and is potentially lethal in mice. To do this, we have included binding sites for hepatocyte-selective microRNA mir-122 within the 3' UTR of the E1A transcription cassette. Imaging versions of these viruses, produced by fusing E1A with luciferase, showed that inclusion of mir-122 binding sites caused up to 80-fold decreased hepatic expression of E1A following intravenous delivery to mice. Animals administered a ten-times lethal dose of wild-type Ad5 (5x10(10) viral particles/mouse) showed substantial hepatic genome replication and extensive liver pathology, while inclusion of 4 microRNA binding sites decreased replication 50-fold and virtually abrogated liver toxicity. This modified wild-type virus retained full activity within cancer cells and provided a potent, liver-safe oncolytic virus. In addition to providing many potent new viruses for cancer virotherapy, microRNA control of virus replication should provide a new strategy for designing safe attenuated vaccines applied across a broad range of viral diseases.

Assembly and stoichiometry of the core structure of the bacterial flagellar type III export gate complex.

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Fukumura, Takuma; Makino, Fumiaki; Dietsche, Tobias; Kinoshita, Miki; Kato, Takayuki; Wagner, Samuel; Namba, Keiichi; Imada, Katsumi; Minamino, Tohru

2017-08-01

The bacterial flagellar type III export apparatus, which is required for flagellar assembly beyond the cell membranes, consists of a transmembrane export gate complex and a cytoplasmic ATPase complex. FlhA, FlhB, FliP, FliQ, and FliR form the gate complex inside the basal body MS ring, although FliO is required for efficient export gate formation in Salmonella enterica. However, it remains unknown how they form the gate complex. Here we report that FliP forms a homohexameric ring with a diameter of 10 nm. Alanine substitutions of conserved Phe-137, Phe-150, and Glu-178 residues in the periplasmic domain of FliP (FliPP) inhibited FliP6 ring formation, suppressing flagellar protein export. FliO formed a 5-nm ring structure with 3 clamp-like structures that bind to the FliP6 ring. The crystal structure of FliPP derived from Thermotoga maritia, and structure-based photo-crosslinking experiments revealed that Phe-150 and Ser-156 of FliPP are involved in the FliP-FliP interactions and that Phe-150, Arg-152, Ser-156, and Pro-158 are responsible for the FliP-FliO interactions. Overexpression of FliP restored motility of a ∆fliO mutant to the wild-type level, suggesting that the FliP6 ring is a functional unit in the export gate complex and that FliO is not part of the final gate structure. Copurification assays revealed that FlhA, FlhB, FliQ, and FliR are associated with the FliO/FliP complex. We propose that the assembly of the export gate complex begins with FliP6 ring formation with the help of the FliO scaffold, followed by FliQ, FliR, and FlhB and finally FlhA during MS ring formation.

A Herpes Simplex Virus Type 1 Mutant Expressing a Baculovirus Inhibitor of Apoptosis Gene in Place of Latency-Associated Transcript Has a Wild-Type Reactivation Phenotype in the Mouse

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Jin, Ling; Perng, Guey-Chuen; Mott, Kevin R.; Osorio, Nelson; Naito, Julia; Brick, David J.; Carpenter, Dale; Jones, Clinton; Wechsler, Steven L.

2005-01-01

The latency-associated transcript (LAT) is essential for the wild-type herpes simplex virus type 1 (HSV-1) high-reactivation phenotype since LAT− mutants have a low-reactivation phenotype. We previously reported that LAT can decrease apoptosis and proposed that this activity is involved in LAT's ability to enhance the HSV-1 reactivation phenotype. The first 20% of the primary 8.3-kb LAT transcript is sufficient for enhancing the reactivation phenotype and for decreasing apoptosis, supporting this proposal. For this study, we constructed an HSV-1 LAT− mutant that expresses the baculovirus antiapoptosis gene product cpIAP under control of the LAT promoter and in place of the LAT region mentioned above. Mice were ocularly infected with this mutant, designated dLAT-cpIAP, and the reactivation phenotype was determined using the trigeminal ganglion explant model. dLAT-cpIAP had a reactivation phenotype similar to that of wild-type virus and significantly higher than that of (i) the LAT− mutant dLAT2903; (ii) dLAT1.5, a control virus containing the same LAT deletion as dLAT-cpIAP, but with no insertion of foreign DNA, thereby controlling for potential readthrough transcription past the cpIAP insert; and (iii) dLAT-EGFP, a control virus identical to dLAT-cpIAP except that it contained the enhanced green fluorescent protein open reading frame (ORF) in place of the cpIAP ORF, thereby controlling for expression of a random foreign gene instead of the cpIAP gene. These results show that an antiapoptosis gene with no sequence similarity to LAT can efficiently substitute for the LAT function involved in enhancing the in vitro-induced HSV-1 reactivation phenotype in the mouse. PMID:16160155

Discovering functions of unannotated genes from a transcriptome survey of wild fungal isolates.

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Ellison, Christopher E; Kowbel, David; Glass, N Louise; Taylor, John W; Brem, Rachel B

2014-04-01

Most fungal genomes are poorly annotated, and many fungal traits of industrial and biomedical relevance are not well suited to classical genetic screens. Assigning genes to phenotypes on a genomic scale thus remains an urgent need in the field. We developed an approach to infer gene function from expression profiles of wild fungal isolates, and we applied our strategy to the filamentous fungus Neurospora crassa. Using transcriptome measurements in 70 strains from two well-defined clades of this microbe, we first identified 2,247 cases in which the expression of an unannotated gene rose and fell across N. crassa strains in parallel with the expression of well-characterized genes. We then used image analysis of hyphal morphologies, quantitative growth assays, and expression profiling to test the functions of four genes predicted from our population analyses. The results revealed two factors that influenced regulation of metabolism of nonpreferred carbon and nitrogen sources, a gene that governed hyphal architecture, and a gene that mediated amino acid starvation resistance. These findings validate the power of our population-transcriptomic approach for inference of novel gene function, and we suggest that this strategy will be of broad utility for genome-scale annotation in many fungal systems. IMPORTANCE Some fungal species cause deadly infections in humans or crop plants, and other fungi are workhorses of industrial chemistry, including the production of biofuels. Advances in medical and industrial mycology require an understanding of the genes that control fungal traits. We developed a method to infer functions of uncharacterized genes by observing correlated expression of their mRNAs with those of known genes across wild fungal isolates. We applied this strategy to a filamentous fungus and predicted functions for thousands of unknown genes. In four cases, we experimentally validated the predictions from our method, discovering novel genes involved in the

TNF Lectin-Like Domain Restores Epithelial Sodium Channel Function in Frameshift Mutants Associated with Pseudohypoaldosteronism Type 1B

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Anita Willam

2017-05-01

Full Text Available Previous in vitro studies have indicated that tumor necrosis factor (TNF activates amiloride-sensitive epithelial sodium channel (ENaC current through its lectin-like (TIP domain, since cyclic peptides mimicking the TIP domain (e.g., solnatide, showed ENaC-activating properties. In the current study, the effects of TNF and solnatide on individual ENaC subunits or ENaC carrying mutated glycosylation sites in the α-ENaC subunit were compared, revealing a similar mode of action for TNF and solnatide and corroborating the previous assumption that the lectin-like domain of TNF is the relevant molecular structure for ENaC activation. Accordingly, TNF enhanced ENaC current by increasing open probability of the glycosylated channel, position N511 in the α-ENaC subunit being identified as the most important glycosylation site. TNF significantly increased Na+ current through ENaC comprising only the pore forming subunits α or δ, was less active in ENaC comprising only β-subunits, and showed no effect on ENaC comprising γ-subunits. TNF did not increase the membrane abundance of ENaC subunits to the extent observed with solnatide. Since the α-subunit is believed to play a prominent role in the ENaC current activating effect of TNF and TIP, we investigated whether TNF and solnatide can enhance αβγ-ENaC current in α-ENaC loss-of-function frameshift mutants. The efficacy of solnatide has been already proven in pathological conditions involving ENaC in phase II clinical trials. The frameshift mutations αI68fs, αT169fs, αP197fs, αE272fs, αF435fs, αR438fs, αY447fs, αR448fs, αS452fs, and αT482fs have been reported to cause pseudohypoaldosteronism type 1B (PHA1B, a rare, life-threatening, salt-wasting disease, which hitherto has been treated only symptomatically. In a heterologous expression system, all frameshift mutants showed significantly reduced amiloride-sensitive whole-cell current compared to wild type αβγ-ENaC, whereas membrane

Genomic Knockout of Endogenous Canine P-Glycoprotein in Wild-Type, Human P-Glycoprotein and Human BCRP Transfected MDCKII Cell Lines by Zinc Finger Nucleases.

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Gartzke, Dominik; Delzer, Jürgen; Laplanche, Loic; Uchida, Yasuo; Hoshi, Yutaro; Tachikawa, Masanori; Terasaki, Tetsuya; Sydor, Jens; Fricker, Gert

2015-06-01

To investigate whether it is possible to specifically suppress the expression and function of endogenous canine P-glycoprotein (cPgp) in Madin-Darby canine kidney type II cells (MDCKII) transfected with hPGP and breast cancer resistance protein (hBCRP) by zinc finger nuclease (ZFN) producing sequence specific DNA double strand breaks. Wild-type, hPGP-transfected, and hBCRP-transfected MDCKII cells were transfected with ZFN targeting for cPgp. Net efflux ratios (NER) of Pgp and Bcrp substrates were determined by dividing efflux ratios (basal-to-apical / apical-to-basal) in over-expressing cell monolayers by those in wild-type ones. From ZFN-transfected cells, cell populations (ko-cells) showing knockout of cPgp were selected based on genotyping by PCR. qRT-PCR analysis showed the significant knock-downs of cPgp and interestingly also cMrp2 expressions. Specific knock-downs of protein expression for cPgp were shown by western blotting and quantitative targeted absolute proteomics. Endogenous canine Bcrp proteins were not detected. For PGP-transfected cells, NERs of 5 Pgp substrates in ko-cells were significantly greater than those in parental cells not transfected with ZFN. Similar result was obtained for BCRP-transfected cells with a dual Pgp and Bcrp substrate. Specific efflux mediated by hPGP or hBCRP can be determined with MDCKII cells where cPgp has been knocked out by ZFN.

Linezolid-Dependent Function and Structure Adaptation of Ribosomes in a Staphylococcus epidermidis Strain Exhibiting Linezolid Dependence

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Kokkori, Sofia; Apostolidi, Maria; Tsakris, Athanassios; Pournaras, Spyros; Stathopoulos, Constantinos; Dinos, George

2014-01-01

Linezolid-dependent growth was recently reported in Staphylococcus epidermidis clinical strains carrying mutations associated with linezolid resistance. To investigate this unexpected behavior at the molecular level, we isolated active ribosomes from one of the linezolid-dependent strains and we compared them with ribosomes isolated from a wild-type strain. Both strains were grown in the absence and presence of linezolid. Detailed biochemical and structural analyses revealed essential differe...

Structure to function: Spider silk and human collagen

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Rabotyagova, Olena S.

, morphological features and assembly. Aside from fundamental perspectives, we anticipate that these results will provide a blueprint for the design of precise materials for a range of potential applications such as controlled release devices, functional coatings, components of tissue regeneration materials and environmentally friendly polymers in future studies. In the second part of this work, human collagen type I was studied as another representative of the family of fibrous proteins. Collagen type I is the most abundant extracellular matrix protein in the human body, providing the basis for tissue structure and directing cellular functions. Collagen has a complex structural hierarchy, organized at different length scales, including the characteristic triple helical feature. In the present study we assessed the relationship between collagen structure (native vs. denatured) and sensitivity to UV radiation with a focus on changes in the primary structure, conformation, microstructure and material properties. Free radical reactions are involved in collagen degradation and a mechanism for UV-induced collagen degradation related to structure was proposed. The results from this study demonstrated the role of collagen supramolecular organization (triple helix) in the context of the effects of electromagnetic radiation on extracellular matrices. Owing to the fact that both silks and collagens are proteins that have found widespread interest for biomaterial related needs, we anticipate that the current studies will serve as a foundation for future biomaterial designs with controlled properties. Furthermore, fundamental insight into self-assembly and environmentally-2mediated degradation, will build a foundation for fundamental understanding of the remodeling and functions of these types of fibrous proteins in vivo and in vitro. This type of insight is essential for many areas of scientific inquiry, from drug delivery, to scaffolds for tissue engineering, and to the stability of

Occurrence and distribution of Giardia species in wild rodents in Germany.

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Helmy, Yosra A; Spierling, Nastasja G; Schmidt, Sabrina; Rosenfeld, Ulrike M; Reil, Daniela; Imholt, Christian; Jacob, Jens; Ulrich, Rainer G; Aebischer, Toni; Klotz, Christian

2018-03-27

Giardiasis is an important gastrointestinal parasitic disease in humans and other mammals caused by the protozoan Giardia duodenalis. This species complex is represented by genetically distinct groups (assemblages A-H) with varying zoonotic potential and host preferences. Wild rodents can harbor potentially zoonotic assemblages A and B, and the rodent-specific assemblage G. Other Giardia spp. found in these animals are Giardia muris and Giardia microti. For the latter, only limited information on genetic typing is available. It has been speculated that wild rodents might represent an important reservoir for parasites causing human giardiasis. The aim of this study was to investigate the occurrence and distribution of Giardia spp. and assemblage types in wild rodents from different study sites in Germany. Screening of 577 wild rodents of the genera Apodemus, Microtus and Myodes, sampled at eleven study sites in Germany, revealed a high overall Giardia prevalence. Giardia species determination at the SSU rDNA gene locus revealed that Apodemus mice, depending on species, were predominantly infected with one of two distinct G. muris sequence types. Giardia microti was the predominant parasite species found in voles of the genera Microtus and Myodes. Only a few animals were positive for potentially zoonotic G. duodenalis. Subtyping at the beta-giardin (bg) and glutamine dehydrogenase (gdh) genes strongly supported the existence of different phylogenetic subgroups of G. microti that are preferentially harbored by distinct host species. The present study highlights the preference of G. muris for Apodemus, and G. microti for Microtus and Myodes hosts and argues for a very low prevalence of zoonotic G. duodenalis assemblages in wild rodents in Germany. It also provides evidence that G. muris and G. microti subdivide into several phylogenetically distinguishable subgroups, each of which appears to be preferentially harbored by species of a particular rodent host genus

The role of cysteine residues in the sulphate transporter, SHST1: construction of a functional cysteine-less transporter.

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Howitt, Susan M

2005-05-20

We investigated the role of cysteine residues in the sulphate transporter, SHST1, with the aim of generating a functional cysteine-less variant. SHST1 contains five cysteine residues and none was essential for function. However, replacement of C421 resulted in a reduction in transport activity. Sulphate transport by C205 mutants was dependent on the size of the residue at this position. Alanine at position 205 resulted in a complete loss of function whereas leucine resulted in a 3-fold increase in sulphate transport relative to wild type SHST1. C205 is located in a putative intracellular loop and our results suggest that this loop may be important for sulphate transport. By replacing C205 with leucine and the other four cysteine residues with alanine, we constructed a cysteine-less variant of SHST1 that has transport characteristics indistinguishable from wild type. This construct will be useful for further structure and function studies of SHST1.

Structural and functional analysis of perforin mutations in association with clinical data of familial hemophagocytic lymphohistiocytosis type 2 (FHL2) patients

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An, Omer; Gursoy, Attila; Gurgey, Aytemiz; Keskin, Ozlem

2013-01-01

Perforin plays a key role in the immune system via pore formation at the target cell membrane in the elimination of virus-infected and transformed cells. A vast number of observed mutations in perforin impair this mechanism resulting in a rare but fatal disease, familial hemophagocytic lymphohistiocytosis type 2 (FHL2). Here we report a comprehensive in silico structural analysis of a collection of 76 missense perforin mutations based on a proposed pore model. In our model, perforin monomers oligomerize having cyclic symmetry in consistent with previously found experimental constraints yet having flexibility in the size of the pore and the number of monomers involved. Clusters of the mutations on the model map to three distinct functional regions of the perforin. Calculated stability (free energy) changes show that the mutations mainly destabilize the protein structure, interestingly however, A91V polymorphism, leads to a more stable one. Structural characteristics of mutations help explain the severe functional consequences on perforin deficient patients. Our study provides a structural approach to the mutation effects on the perforin oligomerization and impaired cytotoxic function in FHL2 patients. PMID:23592409

Intrinsic functional defects of type 2 innate lymphoid cells impair innate allergic inflammation in promyelocytic leukemia zinc finger (PLZF)-deficient mice.

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Verhoef, Philip A; Constantinides, Michael G; McDonald, Benjamin D; Urban, Joseph F; Sperling, Anne I; Bendelac, Albert

2016-02-01

The transcription factor promyelocytic leukemia zinc finger (PLZF) is transiently expressed during development of type 2 innate lymphoid cells (ILC2s) but is not present at the mature stage. We hypothesized that PLZF-deficient ILC2s have functional defects in the innate allergic response and represent a tool for studying innate immunity in a mouse with a functional adaptive immune response. We determined the consequences of PLZF deficiency on ILC2 function in response to innate and adaptive immune stimuli by using PLZF(-/-) mice and mixed wild-type:PLZF(-/-) bone marrow chimeras. PLZF(-/-) mice, wild-type littermates, or mixed bone marrow chimeras were treated with the protease allergen papain or the cytokines IL-25 and IL-33 or infected with the helminth Nippostrongylus brasiliensis to induce innate type 2 allergic responses. Mice were sensitized with intraperitoneal ovalbumin-alum, followed by intranasal challenge with ovalbumin alone, to induce adaptive TH2 responses. Lungs were analyzed for immune cell subsets, and alveolar lavage fluid was analyzed for ILC2-derived cytokines. In addition, ILC2s were stimulated ex vivo for their capacity to release type 2 cytokines. PLZF-deficient lung ILC2s exhibit a cell-intrinsic defect in the secretion of IL-5 and IL-13 in response to innate stimuli, resulting in defective recruitment of eosinophils and goblet cell hyperplasia. In contrast, the adaptive allergic inflammatory response to ovalbumin and alum was unimpaired. PLZF expression at the innate lymphoid cell precursor stage has a long-range effect on the functional properties of mature ILC2s and highlights the importance of these cells for innate allergic responses in otherwise immunocompetent mice. Copyright © 2015 American Academy of Allergy, Asthma & Immunology. All rights reserved.

P2X2 Dominant Deafness Mutations Have No Negative Effect on Wild-Type Isoform: Implications for Functional Rescue and in Deafness Mechanism

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Yan Zhu

2017-11-01

Full Text Available The P2X2 receptor is an ATP-gated ion channel, assembled by three subunits. Recently, it has been found that heterozygous mutations of P2X2 V60L and G353R can cause autosomal dominant nonsyndromic hearing loss. However, the underlying mechanism remains unclear. The fact that heterozygous mutations cause deafness suggests that the mutations may have dominant-negative effect (DNE on wild-type (WT P2X2 isoforms and/or other partners leading to hearing loss. In this study, the effect of these dominant deafness P2X2 mutations on WT P2X2 was investigated. We found that sole transfection of both V60L and G353R deafness mutants could efficiently target to the plasma membrane, like WT P2X2, but exhibit a significantly reduced response to ATP stimulation. Both mutants reduced the channel conductance, but G353R mutation also altered the voltage dependency. Co-expression with WT P2X2 could restore the response to ATP. As the ratio of WT P2X2 vs. mutants increased, the response to ATP was also increased. Computer modeling confirmed that both V60L and G353R dominant-deafness mutant subunits do not have any negative effect on WT P2X2 subunit, when assembled as a heterotrimer. Improper docking or defective gating is the more likely mechanism for impaired channel function by these P2X2 deafness mutations. These results suggest that P2X2 dominant deafness mutations do not have negative effects on WT P2X2 isoforms, and that adding additional WT P2X2 could rescue the lost channel function caused by the deafness mutations. These P2X2 dominant deafness mutations may have negative-effects on other partners leading to hearing loss.

Forming a complex with MHC class I molecules interferes with mouse CD1d functional expression.

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Renukaradhya J Gourapura

Full Text Available CD1d molecules are structurally similar to MHC class I, but present lipid antigens as opposed to peptides. Here, we show that MHC class I molecules physically associate with (and regulate the functional expression of mouse CD1d on the surface of cells. Low pH (3.0 acid stripping of MHC class I molecules resulted in increased surface expression of murine CD1d on antigen presenting cells as well as augmented CD1d-mediated antigen presentation to NKT cells. Consistent with the above results, TAP1-/- mice were found to have a higher percentage of type I NKT cells as compared to wild type mice. Moreover, bone marrow-derived dendritic cells from TAP1-/- mice showed increased antigen presentation by CD1d compared to wild type mice. Together, these results suggest that MHC class I molecules can regulate NKT cell function, in part, by masking CD1d.

Structure-based engineering to restore high affinity binding of an isoform-selective anti-TGFβ1 antibody

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Honey, Denise M.; Best, Annie; Qiu, Huawei

2018-01-01

ABSTRACT Metelimumab (CAT192) is a human IgG4 monoclonal antibody developed as a TGFβ1-specific antagonist. It was tested in clinical trials for the treatment of scleroderma but later terminated due to lack of efficacy. Subsequent characterization of CAT192 indicated that its TGFβ1 binding affinity was reduced by ∼50-fold upon conversion from the parental single-chain variable fragment (scFv) to IgG4. We hypothesized this result was due to decreased conformational flexibility of the IgG that could be altered via engineering. Therefore, we designed insertion mutants in the elbow region and screened for binding and potency. Our results indicated that increasing the elbow region linker length in each chain successfully restored the isoform-specific and high affinity binding of CAT192 to TGFβ1. The crystal structure of the high binding affinity mutant displays large conformational rearrangements of the variable domains compared to the wild-type antigen-binding fragment (Fab) and the low binding affinity mutants. Insertion of two glycines in both the heavy and light chain elbow regions provided sufficient flexibility for the variable domains to extend further apart than the wild-type Fab, and allow the CDR3s to make additional interactions not seen in the wild-type Fab structure. These interactions coupled with the dramatic conformational changes provide a possible explanation of how the scFv and elbow-engineered Fabs bind TGFβ1 with high affinity. This study demonstrates the benefits of re-examining both structure and function when converting scFv to IgG molecules, and highlights the potential of structure-based engineering to produce fully functional antibodies. PMID:29333938

Functional nanometer-scale structures

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Chan, Tsz On Mario

cases, which led to desired chemical phase formation. The residue of polymer thermal decomposition was also controlled and utilized for certain functionality in some nanostructures. Throughout this study, we successfully fabricated several novel functional structures and revealed a new formation mechanism of metal/metal oxide nanotubes. The magnetic and electrical properties of these nanostructures were studied and optimized for applications in soft magnetic materials and spintronics devices. In the second part, (Chapter 7) a study on memristive switching devices with magnetron-sputtered metal-semiconductor-metal thin film structures based on ZnO is presented. Resistive random access memory (RRAM) is a new, non-volatile memory based on the memristor effect theoretically predicted by Leon Chua in 1971 and first experimentally demonstrated by Hewlett Packard in 2008. The unit cell of a RRAM (a memristor) is a two-terminal device in which the switching medium is sandwiched between the top and bottom electrodes and the resistance of the switching medium can be modulated by applying an electrical signal (current or voltage) to the electrodes. On the other hand, the significance of a memristor, as the fourth element of circuit elements besides resistor, capacitor and inductor, is not limited to just being a candidate for next-generation memory. Owing to the unique i-v characteristics of non-linear memristors that cannot be duplicated with any combinations of the other three basic elements in a passive circuitry, many new electrical functions are being developed based on the memristors. In our study, various contact electrode combinations and semiconductor doping profiles were utilized to achieve different functional resistive switching behaviors and to help fundamentally understand the underlying switching mechanisms in ZnO-based thin film structures. Two distinctive switching mechanisms (ferroelectric charge-induced resistive switching and dopant-induced filament-type

Rheological studies of creams. I. Rheological functions and structure of creams.

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Erös, I; Thaleb, A

1994-05-01

Large number of washable (o/w type) creams were prepared for rheological investigation. The rheological functions known from the literature were determined in our studies. Rheological constants were determined by measurements and calculations. From these, we selected those ones which were applicable to characterize the energy status of the coherent structure and which gave the most information for practical work, elaboration of composition and evaluation of stability. These functions and parameters are the following: flow curves, viscosity vs shear time and viscosity vs temperature functions, Bingham-type yield value, plastic viscosity, structure breakdown rate constant, activation energy.

MODY - calculation of ordered structures by symmetry-adapted functions

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Białas, Franciszek; Pytlik, Lucjan; Sikora, Wiesława

2016-01-01

In this paper we focus on the new version of computer program MODY for calculations of symmetryadapted functions based on the theory of groups and representations. The choice of such a functional frame of coordinates for description of ordered structures leads to a minimal number of parameters which must be used for presentation of such structures and investigations of their properties. The aim of this work is to find those parameters, which are coefficients of a linear combination of calculated functions, leading to construction of different types of structure ordering with a given symmetry. A spreadsheet script for simplification of this work has been created and attached to the program.

MT-PCR panel detection of canine parvovirus (CPV-2): Vaccine and wild-type CPV-2 can be difficult to differentiate in canine diagnostic fecal samples.

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Meggiolaro, Maira N; Ly, Anna; Rysnik-Steck, Benjamin; Silva, Carolina; Zhang, Joshua; Higgins, Damien P; Muscatello, Gary; Norris, Jacqueline M; Krockenberger, Mark; Šlapeta, Jan

2017-06-01

Canine parvovirus (CPV-2) remains an important cause of devastating enteritis in young dogs. It can be successfully prevented with live attenuated CPV-2 vaccines when given at the appropriate age and in the absence of maternal antibody interference. Rapid diagnosis of parvoviral enteritis in young dogs is essential to ensuring suitable barrier nursing protocols within veterinary hospitals. The current diagnostic trend is to use multiplexed PCR panels to detect an array of pathogens commonly responsible for diarrhea in dogs. The multiplexed PCR assays do not distinguish wild from vaccine CPV-2. They are highly sensitive and detect even a low level of virus shedding, such as those caused by the CPV-2 vaccine. The aim of this study was to identify the CPV-2 subtypes detected in diagnostic specimens and rule out occult shedding of CPV-2 vaccine strains. For a total of 21 samples that tested positive for CPV-2 in a small animal fecal pathogens diagnostic multiplexed tandem PCR (MT-PCR) panel during 2014-2016 we partially characterized the VP2 gene of CPV-2. Vaccine CPV-2 strain, wild type CPV-2a subtypes and vaccine-like CPV-2b subtypes were detected. High copy number was indicative of wild-type CPV-2a presence, but presence of vaccine-like CPV-2b had a variable copy number in fecal samples. A yardstick approach to a copy number or C t -value to discriminate vaccine strain from a wild type virus of CPV-2 can be, in some cases, potentially misleading. Therefore, discriminating vaccine strain from a wild type subtype of CPV-2 remains ambitious. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effects of single nucleotide polymorphisms on human N-acetyltransferase 2 structure and dynamics by molecular dynamics simulation.

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M Rajasekaran

Full Text Available BACKGROUND: Arylamine N-acetyltransferase 2 (NAT2 is an important catalytic enzyme that metabolizes the carcinogenic arylamines, hydrazine drugs and chemicals. This enzyme is highly polymorphic in different human populations. Several polymorphisms of NAT2, including the single amino acid substitutions R64Q, I114T, D122N, L137F, Q145P, R197Q, and G286E, are classified as slow acetylators, whereas the wild-type NAT2 is classified as a fast acetylator. The slow acetylators are often associated with drug toxicity and efficacy as well as cancer susceptibility. The biological functions of these 7 mutations have previously been characterized, but the structural basis behind the reduced catalytic activity and reduced protein level is not clear. METHODOLOGY/PRINCIPAL FINDINGS: We performed multiple molecular dynamics simulations of these mutants as well as NAT2 to investigate the structural and dynamical effects throughout the protein structure, specifically the catalytic triad, cofactor binding site, and the substrate binding pocket. None of these mutations induced unfolding; instead, their effects were confined to the inter-domain, domain 3 and 17-residue insert region, where the flexibility was significantly reduced relative to the wild-type. Structural effects of these mutations propagate through space and cause a change in catalytic triad conformation, cofactor binding site, substrate binding pocket size/shape and electrostatic potential. CONCLUSIONS/SIGNIFICANCE: Our results showed that the dynamical properties of all the mutant structures, especially in inter-domain, domain 3 and 17-residue insert region were affected in the same manner. Similarly, the electrostatic potential of all the mutants were altered and also the functionally important regions such as catalytic triad, cofactor binding site, and substrate binding pocket adopted different orientation and/or conformation relative to the wild-type that may affect the functions of the mutants

Towards discovering dual functional inhibitors against both wild type and K103N mutant HIV-1 reverse transcriptases: molecular docking and QSAR studies on 4,1-benzoxazepinone analogues

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Zhang, Zhenshan; Zheng, Mingyue; Du, Li; Shen, Jianhua; Luo, Xiaomin; Zhu, Weiliang; Jiang, Hualiang

2006-05-01

To find useful information for discovering dual functional inhibitors against both wild type (WT) and K103N mutant reverse transcriptases (RTs) of HIV-1, molecular docking and 3D-QSAR approaches were applied to a set of twenty-five 4,1-benzoxazepinone analogues of efavirenz (SUSTIVA®), some of them are active against the two RTs. 3D-QSAR models were constructed, based on their binding conformations determined by molecular docking, with r 2 cv values ranging from 0.656 to 0.834 for CoMFA and CoMSIA, respectively. The models were then validated to be highly predictive and extrapolative by inhibitors in two test sets with different molecular skeletons. Furthermore, CoMFA models were found to be well matched with the binding sites of both WT and K103N RTs. Finally, a reasonable pharmacophore model of 4,1-benzoxazepinones were established. The application of the model not only successfully differentiated the experimentally determined inhibitors from non-inhibitors, but also discovered two potent inhibitors from the compound database SPECS. On the basis of both the 3D-QSAR and pharmacophore models, new clues for discovering and designing potent dual functional drug leads against HIV-1 were proposed: (i) adopting positively charged aliphatic group at the cis-substituent of C3; (ii) reducing the electronic density at the position of O4; (iii) positioning a small branched aliphatic group at position of C5; (iv) using the negatively charged bulky substituents at position of C7.

Multi-functionality of the few: current and past uses of wild plants for food and healing in Liubań region, Belarus.

Science.gov (United States)

Sõukand, Renata; Hrynevich, Yanina; Vasilyeva, Iryna; Prakofjewa, Julia; Vnukovich, Yuriy; Paciupa, Jury; Hlushko, Aliaksei; Knureva, Yana; Litvinava, Yulia; Vyskvarka, Siarhei; Silivonchyk, Hanna; Paulava, Alena; Kõiva, Mare; Kalle, Raivo

2017-02-08

This study examined the use of wild plants in the food, medicinal and veterinary areas within a small territory limited to one village council in the Liubań district of Belarus. The objectives of the research were to document the current and past uses of wild plants in this region for food and human/animal medication; to analyse the food, medicinal and veterinary areas in the context of wild plants; and to qualitatively compare the results with relevant publications concerning the wild food plants of Belarus. Fieldwork was carried out as a practical part of a development cooperation project in May 2016 in 11 villages of the Liubań district. One hundred thirty-four respondents were selected randomly. Information about local uses of wild plants was obtained via semi-structured interviews and the folk-history method. Interview records were digitalized and the data structured in Detailed Use Records (DUR), which were divided into food, medicinal and veterinary areas and then analysed to ascertain local perceptions. A total of 2252 DUR of wild plants were recorded. Eighty-eight wild plant taxa belonging to 45 plant families were used across all three areas. Of these, 58 taxa were used in the food, 74 in the medicinal and 23 in the veterinary areas. A relatively high percentage of the taxa were used in both the food and medicinal areas (55%) and an even greater percentage in both the medicinal and veterinary areas (87%). Comparison with earlier research on wild food plants shows the considerable difference among seldom-mentioned taxa or uses, showing possible regional differences despite the homogenization of the population during the Soviet era. As the majority of taxa with overlapping uses belonged to the most utilized plants, there appears to be clear a tendency to use plants in several different areas once they are brought into the home. This may be due to the need to maximize the versatility of limited resources. While the number of wild taxa used is relatively

Wnt signaling loss accelerates the appearance of neuropathological hallmarks of Alzheimer's disease in J20-APP transgenic and wild-type mice.

Science.gov (United States)

Tapia-Rojas, Cheril; Inestrosa, Nibaldo C

2018-02-01

Alzheimer's disease (AD) is a neurodegenerative pathology characterized by aggregates of amyloid-β (Aβ) and phosphorylated tau protein, synaptic dysfunction, and spatial memory impairment. The Wnt signaling pathway has several key functions in the adult brain and has been associated with AD, mainly as a neuroprotective factor against Aβ toxicity and tau phosphorylation. However, dysfunction of Wnt/β-catenin signaling might also play a role in the onset and development of the disease. J20 APPswInd transgenic (Tg) mouse model of AD was treated i.p. with various Wnt signaling inhibitors for 10 weeks during pre-symptomatic stages. Then, cognitive, biochemical and histochemical analyses were performed. Wnt signaling inhibitors induced severe changes in the hippocampus, including alterations in Wnt pathway components and loss of Wnt signaling function, severe cognitive deficits, increased tau phosphorylation and Aβ 1-42 peptide levels, decreased Aβ42/Aβ40 ratio and Aβ 1-42 concentration in the cerebral spinal fluid, and high levels of soluble Aβ species and synaptotoxic oligomers in the hippocampus, together with changes in the amount and size of senile plaques. More important, we also observed severe alterations in treated wild-type (WT) mice, including behavioral impairment, tau phosphorylation, increased Aβ 1-42 in the hippocampus, decreased Aβ 1-42 in the cerebral spinal fluid, and hippocampal dysfunction. Wnt inhibition accelerated the development of the pathology in a Tg AD mouse model and contributed to the development of Alzheimer's-like changes in WT mice. These results indicate that Wnt signaling plays important roles in the structure and function of the adult hippocampus and suggest that inhibition of the Wnt signaling pathway is an important factor in the pathogenesis of AD. Read the Editorial Highlight for this article on page 356. © 2017 International Society for Neurochemistry.

Functional analysis of the cloverleaf-like structure in the 3' untranslated region of bamboo mosaic potexvirus RNA revealed dual roles in viral RNA replication and long distance movement

International Nuclear Information System (INIS)

Chen, I-H.; Meng Hsiao; Hsu, Y.-H.; Tsai, C.-H.

2003-01-01

The 3' untranslated region (UTR) of bamboo mosaic potexvirus (BaMV) RNA was identified to fold into a tertiary structure comprising a cloverleaf-like structure designated ABC domain followed by a major stem-loop D, which in turn is followed by a pseudoknot E and a poly(A) tail. The coat protein accumulation level of the mutant, BaMV40A/ΔABC, lacking ABC domain was just 15% that of wild-type when inoculated into protoplasts of Nicotiana benthamiana. This suggested that ABC domain might play an important role in BaMV RNA replication. To define the precise role of each of the three stem-loops of ABC domain in RNA replication, three mutants BaMV40B and C each lacking stem-loop A, B, and C, respectively, were created. Our results showed that accumulation of viral products of mutants BaMV40B and C were not as efficient as wild-type. On the contrary, level of accumulation of viral products of BaMVA was similar to that of wild-type in protoplasts and inoculated leaves. Interestingly, the accumulation of viral products was not as efficient as that of wild-type in systemic leaves, implying that stem-loop A is dispensable for replication, but signifies a role in systemic accumulation. Using UV cross-linking and competition experiments, it was demonstrated that the E. coli expressed helicase domain of BaMV ORF1 can preferentially interact with the ABC domain

Lysogenic Streptococcus suis isolate SS2-4 containing prophage SMP showed increased mortality in zebra fish compared to the wild-type isolate.

Directory of Open Access Journals (Sweden)

Fang Tang

Full Text Available Streptococcus suis (S. suis infection is considered to be a major problem in the swine industry worldwide. Based on the capsular type, 33 serotypes of S. suis have been described, with serotype 2 (SS2 being the most frequently isolated from diseased piglets. Little is known, however, about the pathogenesis and virulence factors of S. suis. Research on bacteriophages highlights a new area in S. suis research. A S. suis serotype 2 bacteriophage, designated SMP, has been previously isolated in our laboratory. Here, we selected a lysogenic isolate in which the SMP phage was integrated into the chromosome of strain SS2-4. Compared to the wild-type isolate, the lysogenic strain showed increased mortality in zebra fish. Moreover the sensitivity of the lysogenic strain to lysozyme was seven times higher than that of the wild-type.

Environmental Surveillance System To Track Wild Poliovirus Transmission

Science.gov (United States)

Deshpande, Jagadish M.; Shetty, Sushmitha J.; Siddiqui, Zaeem A.

2003-01-01

Eradication of poliomyelitis from large metropolis cities in India has been difficult due to high population density and the presence of large urban slums. Three paralytic poliomyelitis cases were reported in Mumbai, India, in 1999 and 2000 in spite of high immunization coverage and good-quality supplementary immunization activities. We therefore established a systematic environmental surveillance study by weekly screening of sewage samples from three high-risk slum areas to detect the silent transmission of wild poliovirus. In 2001, from among the 137 sewage samples tested, wild poliovirus type 1 was isolated from 35 and wild poliovirus type 3 was isolated from 1. Acute flaccid paralysis (AFP) surveillance indicated one case of paralytic poliomyelitis from the city. Phylogenetic analysis with complete VP1 sequences revealed that the isolates from environmental samples belonged to four lineages of wild polioviruses recently isolated from poliomyelitis cases in Uttar Pradesh and not to those previously isolated from AFP cases in Mumbai. Wild poliovirus thus introduced caused one case of paralytic poliomyelitis. The virus was detected in environmental samples 3 months before. It was found that wild polioviruses introduced several times during the year circulated in Mumbai for a limited period before being eliminated. Environmental surveillance was found to be sensitive for the detection of wild poliovirus silent transmission. Nucleotide sequence analysis helped identify wild poliovirus reservoir areas. PMID:12732567

Electronic structure and physical properties of the spinel-type phase of BeP2N4 from all-electron density functional calculations

International Nuclear Information System (INIS)

Ching, W. Y.; Aryal, Sitram; Rulis, Paul; Schnick, Wolfgang

2011-01-01

Using density-functional-theory-based ab initio methods, the electronic structure and physical properties of the newly synthesized nitride BeP 2 N 4 with a phenakite-type structure and the predicted high-pressure spinel phase of BeP 2 N 4 are studied in detail. It is shown that both polymorphs are wide band-gap semiconductors with relatively small electron effective masses at the conduction-band minima. The spinel-type phase is more covalently bonded due to the increased number of P-N bonds for P at the octahedral sites. Calculations of mechanical properties indicate that the spinel-type polymorph is a promising superhard material with notably large bulk, shear, and Young's moduli. Also calculated are the Be K, P K, P L 3 , and N K edges of the electron energy-loss near-edge structure for both phases. They show marked differences because of the different local environments of the atoms in the two crystalline polymorphs. These differences will be very useful for the experimental identification of the products of high-pressure syntheses targeting the predicted spinel-type phase of BeP 2 N 4 .

Experimental adaptation of wild-type canine distemper virus (CDV to the human entry receptor CD150.

Directory of Open Access Journals (Sweden)

Maria Bieringer

Full Text Available Canine distemper virus (CDV, a close relative of measles virus (MV, is widespread and well known for its broad host range. When the goal of measles eradication may be achieved, and when measles vaccination will be stopped, CDV might eventually cross the species barrier to humans and emerge as a new human pathogen. In order to get an impression how fast such alterations may occur, we characterized required adaptive mutations to the human entry receptors CD150 (SLAM and nectin-4 as first step to infect human target cells. Recombinant wild-type CDV-A75/17(red adapted quickly to growth in human H358 epithelial cells expressing human nectin-4. Sequencing of the viral attachment proteins (hemagglutinin, H, and fusion protein, F genes revealed that no adaptive alteration was required to utilize human nectin-4. In contrast, the virus replicated only to low titres (10(2 pfu/ml in Vero cells expressing human CD150 (Vero-hSLAM. After three passages using these cells virus was adapted to human CD150 and replicated to high titres (10(5 pfu/ml. Sequence analyses revealed that only one amino acid exchange in the H-protein at position 540 Asp→Gly (D540G was required for functional adaptation to human CD150. Structural modelling suggests that the adaptive mutation D540G in H reflects the sequence alteration from canine to human CD150 at position 70 and 71 from Pro to Leu (P70L and Gly to Glu (G71E, and compensates for the gain of a negative charge in the human CD150 molecule. Using this model system our data indicate that only a minimal alteration, in this case one adaptive mutation, is required for adaptation of CDV to the human entry receptors, and help to understand the molecular basis why this adaptive mutation occurs.

Wild and domesticated mushroom consumption in Nigeria ...

African Journals Online (AJOL)

African Crop Science Journal ... On the other hand, if nutrition analysis reveals different nutrition parameters for both types of mushrooms, 43.3% opted for cultivated mushroom, 42.2%, wild; 12.2% both; while 2.2% would eat ... Keywords: Consumption pattern, Lentinus squarrosulus, nutrition, perception, wild mushroom ...

The Microbiome of Seriola lalandi of Wild and Aquaculture Origin Reveals Differences in Composition and Potential Function

Directory of Open Access Journals (Sweden)

Carolina Ramírez

2017-09-01

Full Text Available Seriola lalandi is an economically important species that is globally distributed in temperate and subtropical marine waters. Aquaculture production of this species has had problems associated with intensive fish farming, such as disease outbreaks or nutritional deficiencies causing high mortalities. Intestinal microbiota has been involved in many processes that benefit the host, such as disease control, stimulation of the immune response, and the promotion of nutrient metabolism, among others. However, little is known about the potential functionality of the microbiota and the differences in the composition between wild and aquacultured fish. Here, we assayed the V4-region of the 16S rRNA gene using high-throughput sequencing. Our results showed that there are significant differences between S. lalandi of wild and aquaculture origin (ANOSIM and PERMANOVA, P < 0.05. At the genus level, a total of 13 genera were differentially represented between the two groups, all of which have been described as beneficial microorganisms that have an antagonistic effect against pathogenic bacteria, improve immunological parameters and growth performance, and contribute to nutrition. Additionally, the changes in the presumptive functions of the intestinal microbiota of yellowtail were examined by predicting the metagenomes using PICRUSt. The most abundant functional categories were those corresponding to the metabolism of cofactors and vitamins, amino acid metabolism and carbohydrate metabolism, revealing differences in the contribution of the microbiota depending on the origin of the animals. To our knowledge, this is the first study to characterize and compare the intestinal microbiota of S. lalandi of wild and aquaculture origin using high-throughput sequencing.

The Microbiome of Seriola lalandi of Wild and Aquaculture Origin Reveals Differences in Composition and Potential Function.

Science.gov (United States)

Ramírez, Carolina; Romero, Jaime

2017-01-01

Seriola lalandi is an economically important species that is globally distributed in temperate and subtropical marine waters. Aquaculture production of this species has had problems associated with intensive fish farming, such as disease outbreaks or nutritional deficiencies causing high mortalities. Intestinal microbiota has been involved in many processes that benefit the host, such as disease control, stimulation of the immune response, and the promotion of nutrient metabolism, among others. However, little is known about the potential functionality of the microbiota and the differences in the composition between wild and aquacultured fish. Here, we assayed the V4-region of the 16S rRNA gene using high-throughput sequencing. Our results showed that there are significant differences between S. lalandi of wild and aquaculture origin (ANOSIM and PERMANOVA, P < 0.05). At the genus level, a total of 13 genera were differentially represented between the two groups, all of which have been described as beneficial microorganisms that have an antagonistic effect against pathogenic bacteria, improve immunological parameters and growth performance, and contribute to nutrition. Additionally, the changes in the presumptive functions of the intestinal microbiota of yellowtail were examined by predicting the metagenomes using PICRUSt. The most abundant functional categories were those corresponding to the metabolism of cofactors and vitamins, amino acid metabolism and carbohydrate metabolism, revealing differences in the contribution of the microbiota depending on the origin of the animals. To our knowledge, this is the first study to characterize and compare the intestinal microbiota of S. lalandi of wild and aquaculture origin using high-throughput sequencing.

Gain-of-function mutations in the ALS8 causative gene VAPB have detrimental effects on neurons and muscles

Directory of Open Access Journals (Sweden)

Mario Sanhueza

2013-12-01

Amyotrophic Lateral Sclerosis (ALS is a motor neuron degenerative disease characterized by a progressive, and ultimately fatal, muscle paralysis. The human VAMP-Associated Protein B (hVAPB is the causative gene of ALS type 8. Previous studies have shown that a loss-of-function mechanism is responsible for VAPB-induced ALS. Recently, a novel mutation in hVAPB (V234I has been identified but its pathogenic potential has not been assessed. We found that neuronal expression of the V234I mutant allele in Drosophila (DVAP-V260I induces defects in synaptic structure and microtubule architecture that are opposite to those associated with DVAP mutants and transgenic expression of other ALS-linked alleles. Expression of DVAP-V260I also induces aggregate formation, reduced viability, wing postural defects, abnormal locomotion behavior, nuclear abnormalities, neurodegeneration and upregulation of the heat-shock-mediated stress response. Similar, albeit milder, phenotypes are associated with the overexpression of the wild-type protein. These data show that overexpressing the wild-type DVAP is sufficient to induce the disease and that DVAP-V260I is a pathogenic allele with increased wild-type activity. We propose that a combination of gain- and loss-of-function mechanisms is responsible for VAPB-induced ALS.

High spin structure functions

International Nuclear Information System (INIS)

Khan, H.

1990-01-01

This thesis explores deep inelastic scattering of a lepton beam from a polarized nuclear target with spin J=1. After reviewing the formation for spin-1/2, the structure functions for a spin-1 target are defined in terms of the helicity amplitudes for forward compton scattering. A version of the convolution model, which incorporates relativistic and binding energy corrections is used to calculate the structure functions of a neutron target. A simple parameterization of these structure functions is given in terms of a few neutron wave function parameters and the free nucleon structure functions. This allows for an easy comparison of structure functions calculated using different neutron models. (author)

Decreasing abundance, increasing diversity and changing structure of the wild bee community (Hymenoptera: Anthophila along an urbanization gradient.

Directory of Open Access Journals (Sweden)

Laura Fortel

Full Text Available Wild bees are important pollinators that have declined in diversity and abundance during the last decades. Habitat destruction and fragmentation associated with urbanization are reported as part of the main causes of this decline. Urbanization involves dramatic changes of the landscape, increasing the proportion of impervious surface while decreasing that of green areas. Few studies have investigated the effects of urbanization on bee communities. We assessed changes in the abundance, species richness, and composition of wild bee community along an urbanization gradient.Over two years and on a monthly basis, bees were sampled with colored pan traps and insect nets at 24 sites located along an urbanization gradient. Landscape structure within three different radii was measured at each study site. We captured 291 wild bee species. The abundance of wild bees was negatively correlated with the proportion of impervious surface, while species richness reached a maximum at an intermediate (50% proportion of impervious surface. The structure of the community changed along the urbanization gradient with more parasitic species in sites with an intermediate proportion of impervious surface. There were also greater numbers of cavity-nesting species and long-tongued species in sites with intermediate or higher proportion of impervious surface. However, urbanization had no effect on the occurrence of species depending on their social behavior or body size.We found nearly a third of the wild bee fauna known from France in our study sites. Indeed, urban areas supported a diverse bee community, but sites with an intermediate level of urbanization were the most speciose ones, including greater proportion of parasitic species. The presence of a diverse array of bee species even in the most urbanized area makes these pollinators worthy of being a flagship group to raise the awareness of urban citizens about biodiversity.

Decreasing abundance, increasing diversity and changing structure of the wild bee community (Hymenoptera: Anthophila) along an urbanization gradient.

Science.gov (United States)

Fortel, Laura; Henry, Mickaël; Guilbaud, Laurent; Guirao, Anne Laure; Kuhlmann, Michael; Mouret, Hugues; Rollin, Orianne; Vaissière, Bernard E

2014-01-01

Wild bees are important pollinators that have declined in diversity and abundance during the last decades. Habitat destruction and fragmentation associated with urbanization are reported as part of the main causes of this decline. Urbanization involves dramatic changes of the landscape, increasing the proportion of impervious surface while decreasing that of green areas. Few studies have investigated the effects of urbanization on bee communities. We assessed changes in the abundance, species richness, and composition of wild bee community along an urbanization gradient. Over two years and on a monthly basis, bees were sampled with colored pan traps and insect nets at 24 sites located along an urbanization gradient. Landscape structure within three different radii was measured at each study site. We captured 291 wild bee species. The abundance of wild bees was negatively correlated with the proportion of impervious surface, while species richness reached a maximum at an intermediate (50%) proportion of impervious surface. The structure of the community changed along the urbanization gradient with more parasitic species in sites with an intermediate proportion of impervious surface. There were also greater numbers of cavity-nesting species and long-tongued species in sites with intermediate or higher proportion of impervious surface. However, urbanization had no effect on the occurrence of species depending on their social behavior or body size. We found nearly a third of the wild bee fauna known from France in our study sites. Indeed, urban areas supported a diverse bee community, but sites with an intermediate level of urbanization were the most speciose ones, including greater proportion of parasitic species. The presence of a diverse array of bee species even in the most urbanized area makes these pollinators worthy of being a flagship group to raise the awareness of urban citizens about biodiversity.

Analyzing the 3D Structure of Human Carbonic Anhydrase II and Its Mutants Using Deep View and the Protein Data Bank

Science.gov (United States)

Ship, Noam J.; Zamble, Deborah B.

2005-01-01

The self directed study of a 3D image of a biomolecule stresses the complex nature of the intra- and intermolecular interactions that come together to define its structure. This is made up of a series of in vitro experiments with a wild-type and mutants forms of human carbonic anhydrase II (hCAII) that examine the structure function relationship…

Panitumumab and pegylated liposomal doxorubicin in platinum-resistant epithelial ovarian cancer with KRAS wild-type

DEFF Research Database (Denmark)

Dahl Steffensen, Karina; Waldstrøm, Marianne; Lund, Bente

, and head and neck cancer. No previous studies have evaluated the effect of panitumumab in OC based on KRAS mutation status. Methods: Eligibility criteria are confirmed stage I-IV primary epithelial ovarian/fallopian/peritoneal cancer patients with progression either during or within 6 months after end...... to a total of 33 patients. At present, 15 patients have been enrolled. The primary endpoint is to investigate the response rate in platinum-resistant, KRAS wild- type OC patients treated with PLD supplemented with panitumumab. Translational research is included as a secondary endpoint and tumor tissue...

The structure of Plasmodium falciparum serine hydroxymethyltransferase reveals a novel redox switch that regulates its activities

Energy Technology Data Exchange (ETDEWEB)

Chitnumsub, Penchit; Ittarat, Wanwipa; Jaruwat, Aritsara; Noytanom, Krittikar [National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Paholyothin Road, Klong 1, Klong Luang, Pathumthani 12120 (Thailand); Amornwatcharapong, Watcharee [Mahidol University, Bangkok (Thailand); Pornthanakasem, Wichai [National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Paholyothin Road, Klong 1, Klong Luang, Pathumthani 12120 (Thailand); Chaiyen, Pimchai [Mahidol University, Bangkok (Thailand); Yuthavong, Yongyuth; Leartsakulpanich, Ubolsree [National Center for Genetic Engineering and Biotechnology, 113 Thailand Science Park, Paholyothin Road, Klong 1, Klong Luang, Pathumthani 12120 (Thailand)

2014-06-01

The crystal structure of P. falciparum SHMT revealed snapshots of an intriguing disulfide/sulfhydryl switch controlling the functional activity. Plasmodium falciparum serine hydroxymethyltransferase (PfSHMT), an enzyme in the dTMP synthesis cycle, is an antimalarial target because inhibition of its expression or function has been shown to be lethal to the parasite. As the wild-type enzyme could not be crystallized, protein engineering of residues on the surface was carried out. The surface-engineered mutant PfSHMT-F292E was successfully crystallized and its structure was determined at 3 Å resolution. The PfSHMT-F292E structure is a good representation of PfSHMT as this variant revealed biochemical properties similar to those of the wild type. Although the overall structure of PfSHMT is similar to those of other SHMTs, unique features including the presence of two loops and a distinctive cysteine pair formed by Cys125 and Cys364 in the tetrahydrofolate (THF) substrate binding pocket were identified. These structural characteristics have never been reported in other SHMTs. Biochemical characterization and mutation analysis of these two residues confirm that they act as a disulfide/sulfhydryl switch to regulate the THF-dependent catalytic function of the enzyme. This redox switch is not present in the human enzyme, in which the cysteine pair is absent. The data reported here can be further exploited as a new strategy to specifically disrupt the activity of the parasite enzyme without interfering with the function of the human enzyme.

The structure of Plasmodium falciparum serine hydroxymethyltransferase reveals a novel redox switch that regulates its activities

International Nuclear Information System (INIS)

Chitnumsub, Penchit; Ittarat, Wanwipa; Jaruwat, Aritsara; Noytanom, Krittikar; Amornwatcharapong, Watcharee; Pornthanakasem, Wichai; Chaiyen, Pimchai; Yuthavong, Yongyuth; Leartsakulpanich, Ubolsree

2014-01-01

The crystal structure of P. falciparum SHMT revealed snapshots of an intriguing disulfide/sulfhydryl switch controlling the functional activity. Plasmodium falciparum serine hydroxymethyltransferase (PfSHMT), an enzyme in the dTMP synthesis cycle, is an antimalarial target because inhibition of its expression or function has been shown to be lethal to the parasite. As the wild-type enzyme could not be crystallized, protein engineering of residues on the surface was carried out. The surface-engineered mutant PfSHMT-F292E was successfully crystallized and its structure was determined at 3 Å resolution. The PfSHMT-F292E structure is a good representation of PfSHMT as this variant revealed biochemical properties similar to those of the wild type. Although the overall structure of PfSHMT is similar to those of other SHMTs, unique features including the presence of two loops and a distinctive cysteine pair formed by Cys125 and Cys364 in the tetrahydrofolate (THF) substrate binding pocket were identified. These structural characteristics have never been reported in other SHMTs. Biochemical characterization and mutation analysis of these two residues confirm that they act as a disulfide/sulfhydryl switch to regulate the THF-dependent catalytic function of the enzyme. This redox switch is not present in the human enzyme, in which the cysteine pair is absent. The data reported here can be further exploited as a new strategy to specifically disrupt the activity of the parasite enzyme without interfering with the function of the human enzyme

Detection of multiple cocirculating wild poliovirus type 1 lineages through environmental surveillance: impact and progress during 2011-2013 in Pakistan.

Science.gov (United States)

Alam, Muhammad Masroor; Shaukat, Shahzad; Sharif, Salmaan; Angez, Mehar; Khurshid, Adnan; Malik, Farzana; Rehman, Lubna; Zaidi, Syed Sohail Zahoor

2014-11-01

The environmental surveillance has proven to be a useful tool to identify poliovirus circulation in different countries and was started in Pakistan during July 2009 to support the acute flaccid paralysis (AFP) surveillance system. Sewage samples were collected from 27 environmental sampling (ENV) sites and processed for poliovirus isolation through 2-phase separation method. Poliovirus isolates were identified as Sabin-like or wild type through real-time polymerase chain reaction (PCR). Wild-type strains were subjected to VP1 gene sequencing and phylogenetic analysis performed using MEGA 5.0. During 2011-2013, a total of 668 samples were collected from 4 provinces that resulted in 40% of samples positive for wild poliovirus type-1 (WPV-1). None of the samples were positive for WPV-3. The areas with high frequency of WPV-1 detection were Karachi-Gadap (69%), Peshawar (82%), and Rawalpindi (65%), whereas the samples from Quetta and Sukkur remained negative for WPV during 2013. Phylogenetic analysis revealed 3 major clusters with multiple poliovirus lineages circulating across different country areas as well as in bordering areas of Afghanistan. Environmental surveillance in Pakistan has been proven to be a powerful tool to detect WPV circulation in the absence of poliomyelitis cases in many communities. Our findings emphasize the need to continue and expand such surveillance activities to other high-risk areas in the country. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Hadron structure functions

International Nuclear Information System (INIS)

Martin, F.

1981-03-01

The x dependence of hadron structure functions is investigated. If quarks can exist in very low mass states (10 MeV for d and u quarks) the pion structure function is predicted to behave like (1-x) and not (1-x) 2 in a x-region around 1. Relativistic and non-relativistic quark bound state pictures of hadrons are considered together with their relation with the Q 2 evolution of structure functions. Good agreement with data is in general obtained

Photon structure function

International Nuclear Information System (INIS)

Bardeen, W.A.

1980-11-01

Theoretical understanding of the photon structure function is reviewed. As an illustration of the pointlike component, the parton model is briefly discussed. However, the systematic study of the photon structure function is presented through the framework of the operator product expansion. Perturbative QCD is used as the theoretical basis for the calculation of leading contributions to the operator product expansion. The influence of higher order QCD effects on these results is discussed. Recent results for the polarized structure functions are discussed

Atomic Structure of Type VI Contractile Sheath from Pseudomonas aeruginosa.

Science.gov (United States)

Salih, Osman; He, Shaoda; Planamente, Sara; Stach, Lasse; MacDonald, James T; Manoli, Eleni; Scheres, Sjors H W; Filloux, Alain; Freemont, Paul S

2018-02-06

Pseudomonas aeruginosa has three type VI secretion systems (T6SSs), H1-, H2-, and H3-T6SS, each belonging to a distinct group. The two T6SS components, TssB/VipA and TssC/VipB, assemble to form tubules that conserve structural/functional homology with tail sheaths of contractile bacteriophages and pyocins. Here, we used cryoelectron microscopy to solve the structure of the H1-T6SS P. aeruginosa TssB1C1 sheath at 3.3 Å resolution. Our structure allowed us to resolve some features of the T6SS sheath that were not resolved in the Vibrio cholerae VipAB and Francisella tularensis IglAB structures. Comparison with sheath structures from other contractile machines, including T4 phage and R-type pyocins, provides a better understanding of how these systems have conserved similar functions/mechanisms despite evolution. We used the P. aeruginosa R2 pyocin as a structural template to build an atomic model of the TssB1C1 sheath in its extended conformation, allowing us to propose a coiled-spring-like mechanism for T6SS sheath contraction. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Remote Sensing of plant functional types: Relative importance of biochemical and structural plant traits

Science.gov (United States)

Kattenborn, Teja; Schmidtlein, Sebastian

2017-04-01

Monitoring ecosystems is a key priority in order to understand vegetation patterns, underlying resource cycles and changes their off. Driven by biotic and abiotic factors, plant species within an ecosystem are likely to share similar structural, physiological or phenological traits and can therefore be grouped into plant functional types (PFT). It can be assumed that plants which share similar traits also share similar optical characteristics. Therefore optical remote sensing was identified as a valuable tool for differentiating PFT. Although several authors list structural and biochemical plant traits which are important for differentiating PFT using hyperspectral remote sensing, there is no quantitative or qualitative information on the relative importance of these traits. Thus, little is known about the explicit role of plant traits for an optical discrimination of PFT. One of the main reasons for this is that various optical traits affect the same wavelength regions and it is therefore difficult to isolate the discriminative power of a single trait. A way to determine the effect of single plant traits on the optical reflectance of plant canopies is given by radiative transfer models. The most established radiative transfer model is PROSAIL, which incorporates biochemical and structural plant traits, such as pigment contents or leaf area index. In the present study 25 grassland species of different PFT were cultivated and traits relevant for PROSAIL were measured for the entire vegetation season of 2016. The information content of each trait for differentiating PFTs was determined by applying a Multi-response Permutation Procedure on the actual traits, as well as on simulated canopy spectra derived from PROSAIL. According to our results some traits, especially biochemical traits, show a weaker separability of PFT on a spectral level than compared to the actual trait measurements. Overall structural traits (leaf angle and leaf area index) are more important for

The five functions of psychological type.

Science.gov (United States)

Myers, Steve

2016-04-01

From the mid-1930s to the end of his life, Jung complained that most readers misunderstood the main point of his book Psychological Types. He viewed being a type as one-sided and problematic for a variety of reasons. His symbol-based solution to the 'type problem' involved developing a transcendent function to become the new dominant function of consciousness. However, this function has not featured in the popular use of his typology and Isabel Briggs Myers believed that the one-sidedness of Jung's eight types could be balanced by the auxiliary function. This has led to the transcendent function being widely ignored, and to a developmental philosophy that encourages a degree of one-sidedness. This divergence of popular type theory and analytical psychology is the result of various factors, such as Jung describing typology as containing four functions, and a letter in 1950 where Jung apparently supported Myers' version of type theory. This hinders the application of analytical psychology to normal psychology, and particularly individual and cultural development. If we refer to Jung's typology as containing five functions not four, this more accurately represents both the content of the book Psychological Types and the primary value Jung saw in typology. © 2016 John Wiley & Sons Ltd.

First report of wild boar susceptibility to Porcine circovirus type 3: High prevalence in the Colli Euganei Regional Park (Italy) in the absence of clinical signs.

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Franzo, Giovanni; Tucciarone, Claudia Maria; Drigo, Michele; Cecchinato, Mattia; Martini, Marco; Mondin, Alessandra; Menandro, Maria Luisa

2018-05-18

The genus Circovirus includes one of the most relevant infectious agents affecting domestic pigs, Porcine circovirus type 2 (PCV-2). The wild boar susceptibility to this pathogen has also been demonstrated although the actual epidemiological role of wild populations is still debated. In recent times, a new circovirus, Porcine circovirus type 3 (PCV-3), has been discovered and reported in the presence of several clinical conditions. However, no information is currently available about PCV-3 circulation and prevalence in wild boar. To fill this gap, 187 wild boar serum samples were collected in the Colli Euganei Regional Park (Northern Italy) and screened for PCV-3, demonstrating a high viral prevalence (approximately 30%). No gender differences were demonstrated while a lower infection prevalence was observed in animals younger than 12 months compared to older ones, differently from what described in commercial pigs. Almost all sampled animals were in good health conditions and no association was proven between PCV-3 status and clinical syndromes in wild animals. The genetic characterization of selected strains enlightened a relevant variability and the absence of closely related strains originating from domestic pigs. Therefore, the observed scenario is suggestive of multiple introductions from other wild or domestic swine populations followed by prolonged circulation and independent evolution. Worldwide, this study reports for the first time the high susceptibility of the wild boar to PCV-3 infection. The high prevalence and the absence of association with clinical signs support the marginal role of this virus in the wild boar population ecology. However, its epidemiological role as reservoir endangering commercial swine cannot be excluded and will require further investigations. © 2018 Blackwell Verlag GmbH.

Expression and Functional Analysis of WRKY Transcription Factors in Chinese Wild Hazel, Corylus heterophylla Fisch.

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Zhao, Tian-Tian; Zhang, Jin; Liang, Li-Song; Ma, Qing-Hua; Chen, Xin; Zong, Jian-Wei; Wang, Gui-Xi

2015-01-01

Plant WRKY transcription factors are known to regulate various biotic and abiotic stress responses. In this study we identified a total of 30 putative WRKY unigenes in a transcriptome dataset of the Chinese wild Hazel, Corylus heterophylla, a species that is noted for its cold tolerance. Thirteen full-length of these ChWRKY genes were cloned and found to encode complete protein sequences, and they were divided into three groups, based on the number of WRKY domains and the pattern of zinc finger structures. Representatives of each of the groups, Unigene25835 (group I), Unigene37641 (group II) and Unigene20441 (group III), were transiently expressed as fusion proteins with yellow fluorescent fusion protein in Nicotiana benthamiana, where they were observed to accumulate in the nucleus, in accordance with their predicted roles as transcriptional activators. An analysis of the expression patterns of all 30 WRKY genes revealed differences in transcript abundance profiles following exposure to cold, drought and high salinity conditions. Among the stress-inducible genes, 23 were up-regulated by all three abiotic stresses and the WRKY genes collectively exhibited four different patterns of expression in flower buds during the overwintering period from November to April. The organ/tissue related expression analysis showed that 18 WRKY genes were highly expressed in stem but only 2 (Unigene9262 and Unigene43101) were greatest in male anthotaxies. The expression of Unigene37641, a member of the group II WRKY genes, was substantially up-regulated by cold, drought and salinity treatments, and its overexpression in Arabidopsis thaliana resulted in better seedling growth, compared with wild type plants, under cold treatment conditions. The transgenic lines also had exhibited higher soluble protein content, superoxide dismutase and peroxidase activiety and lower levels of malondialdehyde, which collectively suggets that Unigene37641 expression promotes cold tolerance.

Influence of developmental stage and genotype on liver mRNA levels among wild, domesticated, and hybrid rainbow trout (Oncorhynchus mykiss).

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White, Samantha L; Sakhrani, Dionne; Danzmann, Roy G; Devlin, Robert H

2013-10-02

Release of domesticated strains of fish into nature may pose a threat to wild populations with respect to their evolved genetic structure and fitness. Understanding alterations that have occurred in both physiology and genetics as a consequence of domestication can assist in evaluating the risks posed by introgression of domesticated genomes into wild genetic backgrounds, however the molecular causes of these consequences are currently poorly defined. The present study has examined levels of mRNA in fast-growing pure domesticated (D), slow-growing age-matched pure wild (Wa), slow-growing size-matched pure wild (Ws), and first generation hybrid cross (W/D) rainbow trout (Oncorhynchus mykiss) to investigate the influence of genotype (domesticated vs. wild, and their interactions in hybrids) and developmental stage (age- or size-matched animals) on genetic responses (i.e. dominant vs. recessive) and specific physiological pathways. Highly significant differences in mRNA levels were found between domesticated and wild-type rainbow trout genotypes (321 mRNAs), with many mRNAs in the wild-domesticated hybrid progeny showing intermediate levels. Differences were also found between age-matched and size-matched wild-type trout groups (64 mRNAs), with unique mRNA differences for each of the wild-type groups when compared to domesticated trout (Wa: 114 mRNAs, Ws: 88 mRNAs), illustrating an influence of fish developmental stage affecting findings when used as comparator groups to other genotypes. Analysis of differentially expressed mRNAs (found for both wild-type trout to domesticated comparisons) among the genotypes indicates that 34.8% are regulated consistent with an additive genetic model, whereas 39.1% and 26.1% show a recessive or dominant mode of regulation, respectively. These molecular data are largely consistent with phenotypic data (growth and behavioural assessments) assessed in domesticated and wild trout strains. The present molecular data are concordant with

Evaluation of Electrical Impedance as a Biomarker of Myostatin Inhibition in Wild Type and Muscular Dystrophy Mice.

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Benjamin Sanchez

Full Text Available Non-invasive and effort independent biomarkers are needed to better assess the effects of drug therapy on healthy muscle and that affected by muscular dystrophy (mdx. Here we evaluated the use of multi-frequency electrical impedance for this purpose with comparison to force and histological parameters.Eight wild-type (wt and 10 mdx mice were treated weekly with RAP-031 activin type IIB receptor at a dose of 10 mg kg-1 twice weekly for 16 weeks; the investigators were blinded to treatment and disease status. At the completion of treatment, impedance measurements, in situ force measurements, and histology analyses were performed.As compared to untreated animals, RAP-031 wt and mdx treated mice had greater body mass (18% and 17%, p 70 Hz, but not in the mdx animals. In contrast, maximum force normalized by muscle mass was unchanged in the wt animals and lower in the mdx animals by 21% (p < 0.01. Similarly, myofiber size was only non-significantly higher in treated versus untreated animals (8% p = 0.44 and 12% p = 0.31 for wt and mdx animals, respectively.Our findings demonstrate electrical impedance of muscle reproduce the functional and histological changes associated with myostatin pathway inhibition and do not reflect differences in muscle size or volume. This technique deserves further study in both animal and human therapeutic trials.

Single-Agent Panitumumab in Frail Elderly Patients With Advanced RAS and BRAF Wild-Type Colorectal Cancer: Challenging Drug Label to Light Up New Hope

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Cremolini, Chiara; Aprile, Giuseppe; Lonardi, Sara; Orlandi, Armando; Mennitto, Alessia; Berenato, Rosa; Antoniotti, Carlotta; Casagrande, Mariaelena; Marsico, Valentina; Marmorino, Federica; Cardellino, Giovanni Gerardo; Bergamo, Francesca; Tomasello, Gianluca; Formica, Vincenzo; Longarini, Raffaella; Giommoni, Elisa; Caporale, Marta; Di Bartolomeo, Maria; Loupakis, Fotios; de Braud, Filippo

2015-01-01

Background. No prospective trials have specifically addressed the efficacy and safety of panitumumab in elderly patients with metastatic colorectal cancer (CRC). We aimed at assessing the efficacy and safety of single agent panitumumab in “frail” elderly patients diagnosed with metastatic RAS and BRAF wild-type CRC. Materials and Methods. Forty elderly patients (aged ≥75 years) with metastatic RAS-BRAF wild-type CRC received off-label prescriptions of single-agent panitumumab at seven Italian institutions. Treatment was administered as first line in patients with absolute contraindication to any chemotherapy or as second-line treatment after failure of a fluoropyrimidine-based treatment, in the presence of contraindication to irinotecan. The outcome measures included objective response rate (ORR), as well as progression-free survival (PFS), disease control rate (DCR), overall survival (OS), and safety. Results. The median PFS and OS were 6.4 months (95% confidence interval [CI]: 4.9–8 months) and 14.3 months (95% CI: 10.9–17.7 months), respectively. ORR was 32.5%, and DCR was 72.5%. Dose reductions related to adverse events (AEs) were reported in 9 (23%) patients, but no permanent treatment discontinuation caused by was reported. The most frequent grade 3 AE was skin rash, with an incidence of 20%. Conclusion. Panitumumab is effective and well-tolerated in frail elderly patients with RAS-BRAF wild-type metastatic CRC and deemed unfit for chemotherapy. A randomized study is needed to confirm these data. Implications for Practice: Treatment of elderly patients with metastatic colorectal cancer represents a difficult challenge in clinical practice. A significant proportion of frail elderly patients do not receive treatment, reflecting ongoing uncertainty of clinical benefit and toxicity of chemotherapy. Unfit condition in this cohort of patients further limits antineoplastic prescription and consequently patient survival. RAS and BRAF wild-type status could

Tenofovir alafenamide demonstrates broad cross-genotype activity against wild-type HBV clinical isolates and maintains susceptibility to drug-resistant HBV isolates in vitro.

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Liu, Yang; Miller, Michael D; Kitrinos, Kathryn M

2017-03-01

Tenofovir alafenamide (TAF) is a novel prodrug of tenofovir (TFV). This study evaluated the antiviral activity of TAF against wild-type genotype A-H HBV clinical isolates as well as adefovir-resistant, lamivudine-resistant, and entecavir-resistant HBV isolates. Full length HBV genomes or the polymerase/reverse transcriptase (pol/RT) region from treatment-naïve patients infected with HBV genotypes A-H were amplified and cloned into an expression vector under the control of a CMV promoter. In addition, 11 drug resistant HBV constructs were created by site-directed mutagenesis of a full length genotype D construct. Activity of TAF was measured by transfection of each construct into HepG2 cells and assessment of HBV DNA levels following treatment across a range of TAF concentrations. TAF activity in vitro was similar against wild-type genotype A-H HBV clinical isolates. All lamivudine- and entecavir-resistant isolates and 4/5 adefovir-resistant isolates were found to be sensitive to inhibition by TAF in vitro as compared to the wild-type isolate. The adefovir-resistant isolate rtA181V + rtN236T exhibited low-level reduced susceptibility to TAF. TAF is similarly active in vitro against wild-type genotype A-H HBV clinical isolates. The TAF sensitivity results for all drug-resistant isolates are consistent with what has been observed with the parent drug TFV. The in vitro cell-based HBV phenotyping assay results support the use of TAF in treatment of HBV infected subjects with diverse HBV genotypes, in both treatment-naive and treatment-experienced HBV infected patients. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of Reaction Temperature on Structure, Appearance and Bonding Type of Functionalized Graphene Oxide Modified P-Phenylene Diamine

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Hong-Juan Sun

2018-04-01

Full Text Available In this study, graphene oxides with different functionalization degrees were prepared by a facile one-step hydrothermal reflux method at various reaction temperatures using graphene oxide (GO as starting material and p-phenylenediamine (PPD as the modifier. The effects of reaction temperature on structure, appearance and bonding type of the obtained materials were investigated by X-ray diffraction (XRD, Fourier transform infrared spectroscopy (FT-IR, X-ray photoelectron spectroscopy (XPS, and scanning electron microscopy (SEM. The results showed that when the reaction temperature was 10–70 °C, the GO reacted with PPD through non-covalent ionic bonds (–COO−H3+N–R and hydrogen bonds (C–OH…H2N–X. When the reaction temperature reached 90 °C, the GO was functionalized with PPD through covalent bonds of C–N. The crystal structure of products became more ordered and regular, and the interlayer spacing (d value and surface roughness increased as the temperature increased. Furthermore, the results suggested that PPD was grafted on the surface of GO through covalent bonding by first attacking the carboxyl groups and then the epoxy groups of GO.

Phase II Study of the Dual EGFR/HER3 Inhibitor Duligotuzumab (MEHD7945A) versus Cetuximab in Combination with FOLFIRI in Second-Line RAS Wild-Type Metastatic Colorectal Cancer.

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Hill, Andrew G; Findlay, Michael P; Burge, Matthew E; Jackson, Christopher; Alfonso, Pilar Garcia; Samuel, Leslie; Ganju, Vinod; Karthaus, Meinolf; Amatu, Alessio; Jeffery, Mark; Bartolomeo, Maria Di; Bridgewater, John; Coveler, Andrew L; Hidalgo, Manuel; Kapp, Amy V; Sufan, Roxana I; McCall, Bruce B; Hanley, William D; Penuel, Elicia M; Pirzkall, Andrea; Tabernero, Josep

2018-05-15

Purpose: Duligotuzumab is a dual-action antibody directed against EGFR and HER3. Experimental Design: Metastatic colorectal cancer (mCRC) patients with KRAS ex2 wild-type received duligotuzumab or cetuximab and FOLFIRI until progression or intolerable toxicity. Mandatory tumor samples underwent mutation and biomarker analysis. Efficacy analysis was conducted in patients with RAS exon 2/3 wild-type tumors. Results: Of 134 randomly assigned patients, 98 had RAS ex2/3 wild-type. Duligotuzumab provided no progression-free survival (PFS) or overall survival (OS) benefit compared with cetuximab, although there was a trend for a lower objective response rate (ORR) in the duligotuzumab arm. No relationship was seen between PFS or ORR and ERBB3, NRG1, or AREG expression. There were fewer skin rash events for duligotuzumab but more diarrhea. Although the incidence of grade ≥3 AEs was similar, the frequency of serious AEs was higher for duligotuzumab. Conclusions: Duligotuzumab plus FOLFIRI did not appear to improve the outcomes in patients with RAS exon 2/3 wild-type mCRC compared with cetuximab + FOLFIRI. Clin Cancer Res; 24(10); 2276-84. ©2018 AACR . ©2018 American Association for Cancer Research.

Molecular dynamics simulations revealed structural differences among WRKY domain-DNA interaction in barley (Hordeum vulgare).

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Pandey, Bharati; Grover, Abhinav; Sharma, Pradeep

2018-02-12

The WRKY transcription factors are a class of DNA-binding proteins involved in diverse plant processes play critical roles in response to abiotic and biotic stresses. Genome-wide divergence analysis of WRKY gene family in Hordeum vulgare provided a framework for molecular evolution and functional roles. So far, the crystal structure of WRKY from barley has not been resolved; moreover, knowledge of the three-dimensional structure of WRKY domain is pre-requisites for exploring the protein-DNA recognition mechanisms. Homology modelling based approach was used to generate structures for WRKY DNA binding domain (DBD) and its variants using AtWRKY1 as a template. Finally, the stability and conformational changes of the generated model in unbound and bound form was examined through atomistic molecular dynamics (MD) simulations for 100 ns time period. In this study, we investigated the comparative binding pattern of WRKY domain and its variants with W-box cis-regulatory element using molecular docking and dynamics (MD) simulations assays. The atomic insight into WRKY domain exhibited significant variation in the intermolecular hydrogen bonding pattern, leading to the structural anomalies in the variant type and differences in the DNA-binding specificities. Based on the MD analysis, residual contribution and interaction contour, wild-type WRKY (HvWRKY46) were found to interact with DNA through highly conserved heptapeptide in the pre- and post-MD simulated complexes, whereas heptapeptide interaction with DNA was missing in variants (I and II) in post-MD complexes. Consequently, through principal component analysis, wild-type WRKY was also found to be more stable by obscuring a reduced conformational space than the variant I (HvWRKY34). Lastly, high binding free energy for wild-type and variant II allowed us to conclude that wild-type WRKY-DNA complex was more stable relative to variants I. The results of our study revealed complete dynamic and structural information

Refined algebraic quantisation in a system with nonconstant gauge invariant structure functions

International Nuclear Information System (INIS)

Martínez-Pascual, Eric

2013-01-01

In a previous work [J. Louko and E. Martínez-Pascual, “Constraint rescaling in refined algebraic quantisation: Momentum constraint,” J. Math. Phys. 52, 123504 (2011)], refined algebraic quantisation (RAQ) within a family of classically equivalent constrained Hamiltonian systems that are related to each other by rescaling one momentum-type constraint was investigated. In the present work, the first steps to generalise this analysis to cases where more constraints occur are developed. The system under consideration contains two momentum-type constraints, originally abelian, where rescalings of these constraints by a non-vanishing function of the coordinates are allowed. These rescalings induce structure functions at the level of the gauge algebra. Providing a specific parametrised family of real-valued scaling functions, the implementation of the corresponding rescaled quantum momentum-type constraints is performed using RAQ when the gauge algebra: (i) remains abelian and (ii) undergoes into an algebra of a nonunimodular group with nonconstant gauge invariant structure functions. Case (ii) becomes the first example known to the author where an open algebra is handled in refined algebraic quantisation. Challenging issues that arise in the presence of non-gauge invariant structure functions are also addressed

Structure of β-decay strength functions

International Nuclear Information System (INIS)

Naumov, Y.V.; Bykov, A.A.; Izosimov, I.N.

1983-01-01

The experimental and theoretical studies on the structure of the Gamow--Teller β-decay strength functions are reviewed. Also considered are processes such as M1 γ decay of analog states, the emission of delayed protons, neutrons, and α particles, delayed fission, and the (p, n) reaction at proton energies 100--200 MeV. The results of measurements of the strength functions by γ-ray total absorption are analyzed. It is shown that the β + decay of nuclei far from the stability band exhibits a new type of collective charge-exchange excitation: Gamow--Teller resonance with μ/sub tau/ = +1

Plant species and soil type cooperatively shape the structure and function of microbial communities in the rhizosphere.

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Berg, Gabriele; Smalla, Kornelia

2009-04-01

The rhizosphere is of central importance not only for plant nutrition, health and quality but also for microorganism-driven carbon sequestration, ecosystem functioning and nutrient cycling in terrestrial ecosystems. A multitude of biotic and abiotic factors are assumed to influence the structural and functional diversity of microbial communities in the rhizosphere. In this review, recent studies on the influence of the two factors, plant species and soil type, on rhizosphere-associated microbial communities are discussed. Root exudates and the response of microorganisms to the latter as well as to root morphology were shown to shape rhizosphere microbial communities. All studies revealed that soil is the main reservoir for rhizosphere microorganisms. Many secrets of microbial life in the rhizosphere were recently uncovered due to the enormous progress in molecular and microscopic tools. Physiological and molecular data on the factors that drive selection processes in the rhizosphere are presented here. Furthermore, implications for agriculture, nature conservation and biotechnology will also be discussed.

Bound-state quark and gluon contributions to structure functions in QCD

International Nuclear Information System (INIS)

Brodsky, S.J.

1991-01-01

One can distinguish two types of contributions to the quark and gluon structure functions of hadrons in quantum chromodynamics: 'intrinsic' contributions, which are due to the direct scattering on the bound-state constituents, and 'extrinsic' contributions, which are derived from particles created in the collision. In this talk, I discuss several aspects of deep inealstic structure functions in which the bound-state structure of the proton plays a crucial role: (1) the properties of the intrinsic gluon distribution associated with the proton bound-state wavefunction; (2) the separation of the quark structure function of the proton into intrinsic 'bound-valence' and extrinsic 'non-valence' components which takes into account the Pauli principle; (3) the properties and identification of intrinsic heavy quark structure functions; and (4) a theory of shadowing and anti-shadowing of nuclear structure functions, directly related to quark-nucleon interactions and the gluon saturation phenomenon. (orig.)

Bound-state quark and gluon contributions to structure functions in QCD

International Nuclear Information System (INIS)

Brodsky, S.J.

1990-08-01

One can distinguish two types of contributions to the quark and gluon structure functions of hadrons in quantum chromodynamics: ''intrinsic'' contributions, which are due to the direct scattering on the bound-state constituents, and ''extrinsic'' contributions, which are derived from particles created in the collision. In this talk, I discussed several aspects of deep inelastic structure functions in which the bound-state structure of the proton plays a crucial role: the properties of the intrinsic gluon distribution associated with the proton bound-state wavefunction; the separation of the quark structure function of the proton onto intrinsic ''bound-valence'' and extrinsic ''non-valence'' components which takes into account the Pauli principle; the properties and identification of intrinsic heavy quark structure functions; and a theory of shadowing and anti-shadowing of nuclear structure functions, directly related to quark-nucleon interactions and the gluon saturation phenomenon. 49 refs., 5 figs

Revalorization of wild Asparagus stipularis Forssk. as a traditional vegetable with nutritional and functional properties.

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Adouni, Khaoula; Chahdoura, Hassiba; Mosbah, Habib; Santos-Buelga, Celestino; González-Paramás, Ana M; Ciudad-Mulero, María; Fernandes, Ângela; Calhelha, Ricardo C; Morales, Patricia; Flamini, Guido; Ferreira, Isabel C F R; Achour, Lotfi

2018-03-01

The nutritional values and phytochemical composition of young shoots of wild Asparagus stipularis Forssk. from Tunisia were determined in this study. The antioxidant (tested by four different in vitro assays) and antiproliferative properties (against human tumour cell lines) of aqueous and hydroalcoholic extracts of the young shoots were also investigated. The obtained results indicated that the young shoots are a potential source of valuable nutrients and dietary fiber, and their hydroalcoholic extract showed marked antioxidant and antiproliferative activities in all tested assays. The major phenolic compound, characterized by using a high-performance liquid chromatograph coupled with diode-array and mass spectrometer detectors, in both extracts was diferuloyl glycerol. Our results reinforce the interest in recovering the traditional culinary uses of this wild plant, with potential functional properties, which can be included in the modern diet as an alternative to the variety of vegetables normally used.

Types and Functions of Coastal Structures

DEFF Research Database (Denmark)

Burcharth, H. F.; A. Hughes, Steven

2003-01-01

Coastal structures are used in coastal defence schemes with the objective of preventing shoreline erosion and flooding of the hinterland. Other objectives include sheltering of harbour basins and harbour entrances against waves, stabilization of navigation channels at inlets, and protection...

Brucella abortus ΔrpoE1 confers protective immunity against wild type challenge in a mouse model of brucellosis.

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Willett, Jonathan W; Herrou, Julien; Czyz, Daniel M; Cheng, Jason X; Crosson, Sean

2016-09-30

The Brucella abortus general stress response (GSR) system regulates activity of the alternative sigma factor, σ(E1), which controls transcription of approximately 100 genes and is required for persistence in a BALB/c mouse chronic infection model. We evaluated the host response to infection by a B. abortus strain lacking σ(E1) (ΔrpoE1), and identified pathological and immunological features that distinguish ΔrpoE1-infected mice from wild-type (WT), and that correspond with clearance of ΔrpoE1 from the host. ΔrpoE1 infection was indistinguishable from WT in terms of splenic bacterial burden, inflammation and histopathology up to 6weeks post-infection. However, Brucella-specific serum IgG levels in ΔrpoE1-infected mice were 5 times higher than WT by 4weeks post-infection, and remained significantly higher throughout the course of a 12-week infection. Total IgG and Brucella-specific IgG levels peaked strongly in ΔrpoE1-infected mice at 6weeks, which correlated with reduced splenomegaly and bacterial burden relative to WT-infected mice. Given the difference in immune response to infection with wild-type and ΔrpoE1, we tested whether ΔrpoE1 confers protective immunity to wild-type challenge. Mice immunized with ΔrpoE1 completely resisted WT infection and had significantly higher serum titers of Brucella-specific IgG, IgG2a and IFN-γ after WT challenge relative to age-matched naïve mice. We conclude that immunization of BALB/c mice with the B. abortus GSR pathway mutant, ΔrpoE1, elicits an adaptive immune response that confers significant protective immunity against WT infection. Copyright © 2016 Elsevier Ltd. All rights reserved.

Preclinical efficacy of the MDM2 inhibitor RG7112 in MDM2 amplified and TP53 wild-type glioblastomas

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Verreault, Maite; Schmitt, Charlotte; Goldwirt, Lauriane; Pelton, Kristine; Haidar, Samer; Levasseur, Camille; Guehennec, Jeremy; Knoff, David; Labussiere, Marianne; Marie, Yannick; Ligon, Azra H.; Mokhtari, Karima; Hoang-Xuan, Khe; Sanson, Marc; Alexander, Brian M; Wen, Patrick Y.; Delattre, Jean-Yves; Ligon, Keith L.; Idbaih, Ahmed

2016-01-01

Rationale p53 pathway alterations are key molecular events in glioblastoma (GBM). MDM2 inhibitors increase expression and stability of p53 and are presumed to be most efficacious in patients with TP53 wild-type and MDM2-amplified cancers. However, this biomarker hypothesis has not been tested in patients or patient-derived models for GBM. Methods We performed a preclinical evaluation of RG7112 MDM2 inhibitor, across a panel of 36 patient-derived GBM cell lines (PDCLs), each genetically characterized according to their P53 pathway status. We then performed a pharmacokinetic (PK) profiling of RG7112 distribution in mice and evaluated the therapeutic activity of RG7112 in orthotopic and subcutaneous GBM models. Results MDM2-amplified PDCLs were 44 times more sensitive than TP53 mutated lines that showed complete resistance at therapeutically attainable concentrations (avg. IC50 of 0.52 μM vs 21.9 μM). MDM4 amplified PDCLs were highly sensitive but showed intermediate response (avg. IC50 of 1.2 μM), whereas response was heterogeneous in TP53 wild-type PDCLs with normal MDM2/4 levels (avg. IC50 of 7.7 μM). In MDM2-amplified lines, RG7112 restored p53 activity inducing robust p21 expression and apoptosis. PK profiling of RG7112-treated PDCL intracranial xenografts demonstrated that the compound significantly crosses the blood-brain and the blood-tumor barriers. Most importantly, treatment of MDM2-amplified/TP53 wild-type PDCL-derived model (subcutaneous and orthotopic) reduced tumor growth, was cytotoxic, and significantly increased survival. Conclusion These data strongly support development of MDM2 inhibitors for clinical testing in MDM2-amplified GBM patients. Moreover, significant efficacy in a subset of non-MDM2 amplified models suggests that additional markers of response to MDM2 inhibitors must be identified. PMID:26482041

Interspecific social networks promote information transmission in wild songbirds.

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Farine, Damien R; Aplin, Lucy M; Sheldon, Ben C; Hoppitt, William

2015-03-22

Understanding the functional links between social structure and population processes is a central aim of evolutionary ecology. Multiple types of interactions can be represented by networks drawn for the same population, such as kinship, dominance or affiliative networks, but the relative importance of alternative networks in modulating population processes may not be clear. We illustrate this problem, and a solution, by developing a framework for testing the importance of different types of association in facilitating the transmission of information. We apply this framework to experimental data from wild songbirds that form mixed-species flocks, recording the arrival (patch discovery) of individuals to novel foraging sites. We tested whether intraspecific and interspecific social networks predicted the spread of information about novel food sites, and found that both contributed to transmission. The likelihood of acquiring information per unit of connection to knowledgeable individuals increased 22-fold for conspecifics, and 12-fold for heterospecifics. We also found that species varied in how much information they produced, suggesting that some species play a keystone role in winter foraging flocks. More generally, these analyses demonstrate that this method provides a powerful approach, using social networks to quantify the relative transmission rates across different social relationships.

Structural properties and gelatinisation characteristics of potato and cassava starches and mutants thereof

NARCIS (Netherlands)

Gomand, S.V.; Lamberts, L.; Derde, L.J.; Groesaert, H.; Vandeputte, G.E.; Goderis, B.; Visser, R.G.F.; Delcour, J.A.

2010-01-01

The molecular size of amylopectin (AP) and amylose (AM), AP chain length distribution, crystallinity and granular structure (morphology and granule size distribution) of five wild type potato starches (wtps), five AM free potato starches (amfps), four high-AM potato starches (haps), one wild type

Differences in ultrasonic vocalizations between wild and laboratory California mice (Peromyscus californicus.

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Matina C Kalcounis-Rueppell

Full Text Available BACKGROUND: Ultrasonic vocalizations (USVs emitted by muroid rodents, including laboratory mice and rats, are used as phenotypic markers in behavioral assays and biomedical research. Interpretation of these USVs depends on understanding the significance of USV production by rodents in the wild. However, there has never been a study of muroid rodent ultrasound function in the wild and comparisons of USVs produced by wild and laboratory rodents are lacking to date. Here, we report the first comparison of wild and captive rodent USVs recorded from the same species, Peromyscus californicus. METHODOLOGY AND PRINCIPAL FINDINGS: We used standard ultrasound recording techniques to measure USVs from California mice in the laboratory (Peromyscus Genetic Stock Center, SC, USA and the wild (Hastings Natural History Reserve, CA, USA. To determine which California mouse in the wild was vocalizing, we used a remote sensing method that used a 12-microphone acoustic localization array coupled with automated radio telemetry of all resident Peromyscus californicus in the area of the acoustic localization array. California mice in the laboratory and the wild produced the same types of USV motifs. However, wild California mice produced USVs that were 2-8 kHz higher in median frequency and significantly more variable in frequency than laboratory California mice. SIGNIFICANCE: The similarity in overall form of USVs from wild and laboratory California mice demonstrates that production of USVs by captive Peromyscus is not an artifact of captivity. Our study validates the widespread use of USVs in laboratory rodents as behavioral indicators but highlights that particular characteristics of laboratory USVs may not reflect natural conditions.

Multi-functional composite structures

Science.gov (United States)

Mulligan, Anthony C.; Halloran, John; Popovich, Dragan; Rigali, Mark J.; Sutaria, Manish P.; Vaidyanathan, K. Ranji; Fulcher, Michael L.; Knittel, Kenneth L.

2004-10-19

Fibrous monolith processing techniques to fabricate multifunctional structures capable of performing more than one discrete function such as structures capable of bearing structural loads and mechanical stresses in service and also capable of performing at least one additional non-structural function.

Molecular Evolution of a Type 1 Wild-Vaccine Poliovirus Recombinant during Widespread Circulation in China

Science.gov (United States)

Liu, Hong-Mei; Zheng, Du-Ping; Zhang, Li-Bi; Oberste, M. Steven; Pallansch, Mark A.; Kew, Olen M.

2000-01-01

Type 1 wild-vaccine recombinant polioviruses were isolated from poliomyelitis patients in China from 1991 to 1993. We compared the sequences of 34 recombinant isolates over the 1,353-nucleotide (nt) genomic interval (nt 2480 to 3832) encoding the major capsid protein, VP1, and the protease, 2A. All recombinants had a 367-nt block of sequence (nt 3271 to 3637) derived from the Sabin 1 oral poliovirus vaccine strain spanning the 3′-terminal sequences of VP1 (115 nt) and the 5′ half of 2A (252 nt). The remaining VP1 sequences were closely (up to 99.5%) related to those of a major genotype of wild type 1 poliovirus endemic to China up to 1994. In contrast, the non-vaccine-derived sequences at the 3′ half of 2A were more distantly related (polioviruses from China. The vaccine-derived sequences of the earliest (April 1991) isolates completely matched those of Sabin 1. Later isolates diverged from the early isolates primarily by accumulation of synonymous base substitutions (at a rate of ∼3.7 × 10−2 substitutions per synonymous site per year) over the entire VP1-2A interval. Distinct evolutionary lineages were found in different Chinese provinces. From the combined epidemiologic and evolutionary analyses, we propose that the recombinant virus arose during mixed infection of a single individual in northern China in early 1991 and that its progeny spread by multiple independent chains of transmission into some of the most populous areas of China within a year of the initiating infection. PMID:11070012

Agrobacterium-mediated transformation of grapefruit with the wild-type and mutant RNA-dependent RNA polymerase genes of Citrus tristeza virus

Science.gov (United States)

Citrus paradisi Macf. cv. Duncan was transformed with constructs coding for the wild-type and mutant RNA-dependent RNA polymerase (RdRp) of Citrus tristeza virus (CTV) for exploring replicase-mediated pathogen-derived resistance (RM-PDR). The RdRp gene was amplified from CTV genome and used to gener...

Purification and cellular localization of wild type and mutated dihydrolipoyltransacetylases from Azotobacter vinelandii and Escherichia coli expressed in E. coli

NARCIS (Netherlands)

Schulze, Egbert; Westphal, Adrie H.; Veenhuis, Marten; Kok, Arie de

1992-01-01

Wild type dihydrolipoyltransacetylase(E2p)-components from the pyruvate dehydrogenase complex of A. vinelandii or E. coli, and mutants of A. vinelandii E2p with stepwise deletions of the lipoyl domains or the alanine- and proline-rich region between the binding and the catalytic domain have been

Differential functional readthrough over homozygous nonsense mutations contributes to the bleeding phenotype in coagulation factor VII deficiency.

Science.gov (United States)

Branchini, A; Ferrarese, M; Lombardi, S; Mari, R; Bernardi, F; Pinotti, M

2016-10-01

Essentials Potentially null homozygous Factor(F)7 nonsense mutations are associated to variable bleeding symptoms. Readthrough of p.Ser112X (life-threatening) and p.Cys132X (moderate) stop codons was investigated. Readthrough-mediated insertion of wild-type or tolerated residues produce functional proteins. Functional readthrough over homozygous F7 nonsense mutations contributes to the bleeding phenotype. Background Whereas the rare homozygous nonsense mutations causing factor (F)VII deficiency may predict null conditions that are almost completely incompatible with life, they are associated with appreciable differences in hemorrhagic symptoms. The misrecognition of premature stop codons (readthrough) may account for variable levels of functional full-length proteins. Objectives To experimentally evaluate the basal and drug-induced levels of FVII resulting from the homozygous p.Cys132X and p.Ser112X nonsense mutations that are associated with moderate (132X) or life-threatening (112X) symptoms, and that are predicted to undergo readthrough with (132X) or without (112X) production of wild-type FVII. Methods We transiently expressed recombinant FVII (rFVII) nonsense and missense variants in human embryonic kidney 293 cells, and evaluated secreted FVII protein and functional levels by ELISA, activated FX generation, and coagulation assays. Results The levels of functional FVII produced by p.Cys132X and p.Ser112X mutants (rFVII-132X, 1.1% ± 0.2% of wild-type rFVII; rFVII-112X, 0.5% ± 0.1% of wild-type rFVII) were compatible with the occurrence of spontaneous readthrough, which was magnified by the addition of G418 - up to 12% of the wild-type value for the rFVII-132X nonsense variant. The predicted missense variants arising from readthrough abolished (rFVII-132Trp/Arg) or reduced (rFVII-112Trp/Cys/Arg, 22-45% of wild-type levels) secretion and function. These data suggest that the appreciable rescue of p.Cys132X function was driven by reinsertion of the wild-type

Identification, structure, and function of a novel type VI secretion peptidoglycan glycoside hydrolase effector-immunity pair.

Science.gov (United States)

Whitney, John C; Chou, Seemay; Russell, Alistair B; Biboy, Jacob; Gardiner, Taylor E; Ferrin, Michael A; Brittnacher, Mitchell; Vollmer, Waldemar; Mougous, Joseph D

2013-09-13

Bacteria employ type VI secretion systems (T6SSs) to facilitate interactions with prokaryotic and eukaryotic cells. Despite the widespread identification of T6SSs among Gram-negative bacteria, the number of experimentally validated substrate effector proteins mediating these interactions remains small. Here, employing an informatics approach, we define novel families of T6S peptidoglycan glycoside hydrolase effectors. Consistent with the known intercellular self-intoxication exhibited by the T6S pathway, we observe that each effector gene is located adjacent to a hypothetical open reading frame encoding a putative periplasmically localized immunity determinant. To validate our sequence-based approach, we functionally investigate a representative family member from the soil-dwelling bacterium Pseudomonas protegens. We demonstrate that this protein is secreted in a T6SS-dependent manner and that it confers a fitness advantage in growth competition assays with Pseudomonas putida. In addition, we determined the 1.4 Å x-ray crystal structure of this effector in complex with its cognate immunity protein. The structure reveals the effector shares highest overall structural similarity to a glycoside hydrolase family associated with peptidoglycan N-acetylglucosaminidase activity, suggesting that T6S peptidoglycan glycoside hydrolase effector families may comprise significant enzymatic diversity. Our structural analyses also demonstrate that self-intoxication is prevented by the immunity protein through direct occlusion of the effector active site. This work significantly expands our current understanding of T6S effector diversity.

Identification, Structure, and Function of a Novel Type VI Secretion Peptidoglycan Glycoside Hydrolase Effector-Immunity Pair*

Science.gov (United States)

Whitney, John C.; Chou, Seemay; Russell, Alistair B.; Biboy, Jacob; Gardiner, Taylor E.; Ferrin, Michael A.; Brittnacher, Mitchell; Vollmer, Waldemar; Mougous, Joseph D.

2013-01-01

Bacteria employ type VI secretion systems (T6SSs) to facilitate interactions with prokaryotic and eukaryotic cells. Despite the widespread identification of T6SSs among Gram-negative bacteria, the number of experimentally validated substrate effector proteins mediating these interactions remains small. Here, employing an informatics approach, we define novel families of T6S peptidoglycan glycoside hydrolase effectors. Consistent with the known intercellular self-intoxication exhibited by the T6S pathway, we observe that each effector gene is located adjacent to a hypothetical open reading frame encoding a putative periplasmically localized immunity determinant. To validate our sequence-based approach, we functionally investigate a representative family member from the soil-dwelling bacterium Pseudomonas protegens. We demonstrate that this protein is secreted in a T6SS-dependent manner and that it confers a fitness advantage in growth competition assays with Pseudomonas putida. In addition, we determined the 1.4 Å x-ray crystal structure of this effector in complex with its cognate immunity protein. The structure reveals the effector shares highest overall structural similarity to a glycoside hydrolase family associated with peptidoglycan N-acetylglucosaminidase activity, suggesting that T6S peptidoglycan glycoside hydrolase effector families may comprise significant enzymatic diversity. Our structural analyses also demonstrate that self-intoxication is prevented by the immunity protein through direct occlusion of the effector active site. This work significantly expands our current understanding of T6S effector diversity. PMID:23878199

Genetic variability in Cynara cardunculus L. domestic and wild types for grain oil production and fatty acids composition

International Nuclear Information System (INIS)

Raccuia, Salvatore Antonino; Piscioneri, Ilario; Sharma, Neeta; Melilli, Maria Grazia

2011-01-01

This paper aimed to study the genetic variability within different types of Cynara cardunculus L., domestic and wild types, for their grain oil amount and oil fatty acid composition. The grain oils were extracted from 8 domestic cardoons and 4 wild cardoons, by Soxhlet method, and obtained oils were characterized for palmitic, stearic, oleic and linoleic acids by gas chromatography. The oil amount, resulted on average of accessions 216 g kg -1 DM with a good range of variability (CV = 11.7%). Unsaturated acids (oleic and linoleic) predominated over saturated ones (stearic and palmitic acids), the chemical characterization of extracted oil, showed the main compound (as % of analysed fatty acids), averaged for all populations, was linoleic acid (44.5%), followed by oleic acid (42.6%), palmitic acid (9.8%) and stearic acid (3.1%). In particular referring the oleic acid wild cardoon populations showed a mean value of 289 g kg -1 oil, against a mean value of 472 g kg -1 oil showed by domestic cardoon accessions. Three of the studied domestic cardoon ('DC1', 'DC3' and 'DC7') showed values higher than 795 g kg -1 oil, while all the other accessions had concentration lower than 370 g kg -1 oil. The three types of domestic cardoon 'DC1', 'DC3' and 'DC7' showed a fatty acids profile similar to genetic modified sunflower oil, representing new genetic material that potentially could be used for high quality biodiesel production, characterised by a low Iodine Number. -- Highlights: → The grain oils from 12 cardoons were characterized for fatty acids composition. → The oil amount, resulted on average of accessions 216 g kg -1 DM. → Oleic and linoleic acids predominated over stearic and palmitic acids. → Three domestic cardoons grain oil showed high oleic acid content (795 g kg -1 oil). → This oil could be used for high quality biodiesel production, with a low IN.

The use of impression management tactics in structured interviews: a function of question type?

Science.gov (United States)

Ellis, Aleksander P J; West, Bradley J; Ryan, Ann Marie; DeShon, Richard P

2002-12-01

This study investigated impression management tactic use during structured interviews containing both experience-based and situational questions. Specifically, the authors examined whether applicants' use of impression management tactics depended on question type. Results from 119 structured interviews indicated that almost all of the applicants used some form of impression management. Significantly more assertive than defensive impression management tactics were used, and among assertive tactics, applicants tended to use self-promotion rather than ingratiation. However, different question types prompted the use of different impression management tactics. Ingratiation tactics were used significantly more when applicants answered situational questions, whereas self-promotion tactics were used significantly more when applicants answered experience-based questions. Furthermore, the use of self-promotion and ingratiation tactics was positively related to interviewer evaluations.

Cetuximab Plus Various Chemotherapy Regimens for Patients with KRAS Wild-Type Metastatic Colorectal Cancer.

Science.gov (United States)

Azadeh, Payam; Mortazavi, Nafiseh; Tahmasebi, Arezoo; Hosseini Kamal, Farnaz; Novin, Kambiz

2016-01-01

The aim of this study was to compare the efficacy and hematologic toxicity of cetuximab combined with various types of chemotherapy regimens in patients with KRAS wild-type metastatic colorectal cancer (mCRC). The response rate, progression-free survival (PFS) and overall survival of the patients were analyzed. In total, 45 patients were included in the study. The overall response rate for the combination of cetuximab and FOLFOX, FOLFIRI and CAPOX was 20, 46 and 30%, respectively, but the differences were not statistically significant. The median PFS for the three groups were 8, 6 and 3.5 months, respectively, but again these differences were not significant. All-grade leukopenia and anemia for the cetuximab plus FOLFOX group were significantly higher than for the other chemotherapy regimens. Our findings suggest that the combination of cetuximab and the three standard chemotherapy regimens resulted in the same outcomes in our patient population of mCRC, with higher hematologic toxicities among the FOLFOX subgroup. © 2015 S. Karger AG, Basel.

Molecular dynamics simulation studies of the wild type and E92Q/N155H mutant of Elvitegravir-resistance HIV-1 integrase.

Science.gov (United States)

Chen, Qi; Cheng, Xiaolin; Wei, Dongqing; Xu, Qin

2015-03-01

Although Elvitegravir (EVG) is a newly developed antiretrovirals drug to treat the acquired immunodeficiency syndrome (AIDS), drug resistance has already been found in clinic, such as E92Q/N155H and Q148H/G140S. Several structural investigations have already been reported to reveal the molecular mechanism of the drug resistance. As full length crystal structure for HIV-1 integrase is still unsolved, we herein use the crystal structure of the full length prototype foamy virus (PFV) in complex with virus DNA and inhibitor Elvitegravir as a template to construct the wild type and E92Q/N155H mutant system of HIV-1 integrase. Molecular dynamic simulations was used to revel the binding mode and the drug resistance of the EVG ligand in E92Q/N155H. Several important interactions were discovered between the mutated residues and the residues in the active site of the E92Q/N155H double mutant pattern, and cross correlation and clustering methods were used for detailed analysis. The results from the MD simulation studies will be used to guide the experimental efforts of developing novel inhibitors against drug-resistant HIV integrase mutants.

Therapeutic monoclonal antibody N-glycosylation - Structure, function and therapeutic potential.

Science.gov (United States)

Cymer, Florian; Beck, Hermann; Rohde, Adelheid; Reusch, Dietmar

2018-03-01

Therapeutic antibodies (IgG-type) contain several post-translational modifications (PTMs) whereby introducing a large heterogeneity, both structural and functional, into this class of therapeutics. Of these modifications, glycosylation in the fragment crystallizable (Fc) region is the most heterogeneous PTM, which can affect the stability of the molecule and interactions with Fc-receptors in vivo. Hence, the glycoform distribution can affect the mode of action and have implications for bioactivity, safety and efficacy of the drug. Main topics of the manuscript include: What factors influence the (Fc) glycan pattern in therapeutic antibodies and how can these glycans be characterized? How does structure of the Fc-glycan relate to function and what methods are available to characterize those functions? Although heterogeneous in their scope, the different sections are intended to combine current knowledge on structure-function correlations of IgG glycan structures with regard to Fc (effector) functions, as well as basic aspects and methodologies for their assessment. Copyright © 2017. Published by Elsevier Ltd.

C-type lectins do not act as functional receptors for filovirus entry into cells

Energy Technology Data Exchange (ETDEWEB)

Matsuno, Keita; Nakayama, Eri; Noyori, Osamu [Department of Global Epidemiology, Hokkaido University Research Center for Zoonosis Control, Sapporo (Japan); Marzi, Andrea; Ebihara, Hideki [Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, Hamilton, MT (United States); Irimura, Tatsuro [Graduate School of Pharmaceutical Science, University of Tokyo, Tokyo (Japan); Feldmann, Heinz [Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, Hamilton, MT (United States); Takada, Ayato, E-mail: atakada@czc.hokudai.ac.jp [Department of Global Epidemiology, Hokkaido University Research Center for Zoonosis Control, Sapporo (Japan)

2010-12-03

Research highlights: {yields} Filovirus glycoprotein (GP) having a deficient receptor binding region were generated. {yields} Mutant GPs mediated virus entry less efficiently than wild-type GP. {yields} Mutant GPs bound to C-type lectins but not mediated entire steps of cellular entry. {yields} C-type lectins do not independently mediate filovirus entry into cells. {yields} Other molecule(s) are required for C-type lectin-mediated entry of filoviruses. -- Abstract: Cellular C-type lectins have been reported to facilitate filovirus infection by binding to glycans on filovirus glycoprotein (GP). However, it is not clearly known whether interaction between C-type lectins and GP mediates all the steps of virus entry (i.e., attachment, internalization, and membrane fusion). In this study, we generated vesicular stomatitis viruses pseudotyped with mutant GPs that have impaired structures of the putative receptor binding regions and thus reduced ability to infect the monkey kidney cells that are routinely used for virus propagation. We found that infectivities of viruses with the mutant GPs dropped in C-type lectin-expressing cells, parallel with those in the monkey kidney cells, whereas binding activities of these GPs to the C-type lectins were not correlated with the reduced infectivities. These results suggest that C-type lectin-mediated entry of filoviruses requires other cellular molecule(s) that may be involved in virion internalization or membrane fusion.

C-type lectins do not act as functional receptors for filovirus entry into cells

International Nuclear Information System (INIS)

Matsuno, Keita; Nakayama, Eri; Noyori, Osamu; Marzi, Andrea; Ebihara, Hideki; Irimura, Tatsuro; Feldmann, Heinz; Takada, Ayato

2010-01-01

Research highlights: → Filovirus glycoprotein (GP) having a deficient receptor binding region were generated. → Mutant GPs mediated virus entry less efficiently than wild-type GP. → Mutant GPs bound to C-type lectins but not mediated entire steps of cellular entry. → C-type lectins do not independently mediate filovirus entry into cells. → Other molecule(s) are required for C-type lectin-mediated entry of filoviruses. -- Abstract: Cellular C-type lectins have been reported to facilitate filovirus infection by binding to glycans on filovirus glycoprotein (GP). However, it is not clearly known whether interaction between C-type lectins and GP mediates all the steps of virus entry (i.e., attachment, internalization, and membrane fusion). In this study, we generated vesicular stomatitis viruses pseudotyped with mutant GPs that have impaired structures of the putative receptor binding regions and thus reduced ability to infect the monkey kidney cells that are routinely used for virus propagation. We found that infectivities of viruses with the mutant GPs dropped in C-type lectin-expressing cells, parallel with those in the monkey kidney cells, whereas binding activities of these GPs to the C-type lectins were not correlated with the reduced infectivities. These results suggest that C-type lectin-mediated entry of filoviruses requires other cellular molecule(s) that may be involved in virion internalization or membrane fusion.

Innate immune responses to obesity in cloned and wild-type domestic pig

DEFF Research Database (Denmark)

Højbøge, Tina Rødgaard; Skovgaard, Kerstin; Stagsted, Jan

as a refined pig model for obesity-induced innate host responses by reducing pig-to-pig biological variation compared to wild-type (WT) pigs (n=19). Pigs were fed ad libitum with a high fat/high sucrose diet to induce obesity or kept lean on a restricted diet (60% of ad libitum intake) beginning at three...... months of age. mRNA expression levels were determined for 39 innate immune factors on a high-throughput qPCR system in samples from liver, abdominal fat, mesenteric fat and subcutaneous fat. Previous findings have suggested that cloning may affect certain phenotypic traits of pigs including basic...... concentrations and responsiveness of components of the innate immune system. Terminal body weights at 7½ - 9½ months of age were significantly higher for both (WT and cloned) obese groups compared to the lean groups. However, obese WT pigs weighed significantly more than obese cloned pigs (P

Unpolarized Structure Functions

International Nuclear Information System (INIS)

Christy, M.E.; Melnitchouk, W.

2011-01-01

Over the past decade measurements of unpolarized structure functions with unprecedented precision have significantly advanced our knowledge of nucleon structure. These have for the first time allowed quantitative tests of the phenomenon of quark-hadron duality, and provided a deeper understanding of the transition from hadron to quark degrees of freedom in inclusive scattering. Dedicated Rosenbluth-separation experiments have yielded high-precision transverse and longitudinal structure functions in regions previously unexplored, and new techniques have enabled the first glimpses of the structure of the free neutron, without contamination from nuclear effects.

Identification of proteins that regulate radiation-induced apoptosis in murine tumors with wild type p53

Energy Technology Data Exchange (ETDEWEB)

Seong, Jinsil; Oh, Hae Jin; Kim, Jiyoung; An, Jeung Hee; Kim, Wonwoo [Dept. of Radiation Oncology, Yonsei Univ. Medical College, Seoul (Korea, Republic of)

2007-09-15

In this study, we investigated the molecular factors determining the induction of apoptosis by radiation. Two murine tumors syngeneic to C3H/HeJ mice were used: an ovarian carcinoma OCa-I, and a hepatocarcinoma HCa-I. Both have wild type p53, but display distinctly different radiosensitivity in terms of specific growth delay (12.7 d in OCa-I and 0.3 d in HCa-I) and tumor cure dose 50% (52.6 Gy in OCa-I and >80 Gy in HCa-I). Eight-mm tumors on the thighs of mice were irradiated with 25 Gy and tumor samples were collected at regular time intervals after irradiation. The peak levels of apoptosis were 16.1{+-}0.6% in OCa-I and 0.2{+-}0.0% in HCa-I at 4 h after radiation, and this time point was used for subsequent proteomics analysis. Protein spots were identified by peptide mass fingerprinting with a focus on those related to apoptosis. In OCa-I tumors, radiation increased the expression of cytochrome c oxidase and Bcl2/adenovirus E1B-interacting 2 (Nip 2) protein higher than 3-fold. However in HCa-I, these two proteins showed no significant change. The results suggest that radiosensitivity in tumors with wild type p53 is regulated by a complex mechanism. Furthermore, these proteins could be molecular targets for a novel therapeutic strategy involving the regulation of radiosensitivity. (author)

Identification of proteins that regulate radiation-induced apoptosis in murine tumors with wild type p53

International Nuclear Information System (INIS)

Seong, Jinsil; Oh, Hae Jin; Kim, Jiyoung; An, Jeung Hee; Kim, Wonwoo

2007-01-01

In this study, we investigated the molecular factors determining the induction of apoptosis by radiation. Two murine tumors syngeneic to C3H/HeJ mice were used: an ovarian carcinoma OCa-I, and a hepatocarcinoma HCa-I. Both have wild type p53, but display distinctly different radiosensitivity in terms of specific growth delay (12.7 d in OCa-I and 0.3 d in HCa-I) and tumor cure dose 50% (52.6 Gy in OCa-I and >80 Gy in HCa-I). Eight-mm tumors on the thighs of mice were irradiated with 25 Gy and tumor samples were collected at regular time intervals after irradiation. The peak levels of apoptosis were 16.1±0.6% in OCa-I and 0.2±0.0% in HCa-I at 4 h after radiation, and this time point was used for subsequent proteomics analysis. Protein spots were identified by peptide mass fingerprinting with a focus on those related to apoptosis. In OCa-I tumors, radiation increased the expression of cytochrome c oxidase and Bcl2/adenovirus E1B-interacting 2 (Nip 2) protein higher than 3-fold. However in HCa-I, these two proteins showed no significant change. The results suggest that radiosensitivity in tumors with wild type p53 is regulated by a complex mechanism. Furthermore, these proteins could be molecular targets for a novel therapeutic strategy involving the regulation of radiosensitivity. (author)

Structure-Function Network Mapping and Its Assessment via Persistent Homology

Science.gov (United States)

2017-01-01

Understanding the relationship between brain structure and function is a fundamental problem in network neuroscience. This work deals with the general method of structure-function mapping at the whole-brain level. We formulate the problem as a topological mapping of structure-function connectivity via matrix function, and find a stable solution by exploiting a regularization procedure to cope with large matrices. We introduce a novel measure of network similarity based on persistent homology for assessing the quality of the network mapping, which enables a detailed comparison of network topological changes across all possible thresholds, rather than just at a single, arbitrary threshold that may not be optimal. We demonstrate that our approach can uncover the direct and indirect structural paths for predicting functional connectivity, and our network similarity measure outperforms other currently available methods. We systematically validate our approach with (1) a comparison of regularized vs. non-regularized procedures, (2) a null model of the degree-preserving random rewired structural matrix, (3) different network types (binary vs. weighted matrices), and (4) different brain parcellation schemes (low vs. high resolutions). Finally, we evaluate the scalability of our method with relatively large matrices (2514x2514) of structural and functional connectivity obtained from 12 healthy human subjects measured non-invasively while at rest. Our results reveal a nonlinear structure-function relationship, suggesting that the resting-state functional connectivity depends on direct structural connections, as well as relatively parsimonious indirect connections via polysynaptic pathways. PMID:28046127

The effect of dietary folic acid deficiency on the cytotoxic and mutagenic responses to methyl methanesulfonate in wild-type and in 3-methyladenine DNA glycosylase-deficient Aag null mice.

Science.gov (United States)

Branda, Richard F; O'Neill, J Patrick; Brooks, Elice M; Powden, Cheryl; Naud, Shelly J; Nicklas, Janice A

2007-02-03

Folic acid deficiency (FA-) augments DNA damage caused by alkylating agents. The role of DNA repair in modulating this damage was investigated in mice. Weanling wild-type or 3-methyladenine glycosylase (Aag) null mice were maintained on a FA- diet or the same diet supplemented with folic acid (FA+) for 4 weeks. They were then treated with methyl methanesulfonate (MMS), 100mg/kg i.p. Six weeks later, spleen cells were collected for assays of non-selected and 6-thioguanine (TG) selected cloning efficiency to measure the mutant frequency at the Hprt locus. In wild-type mice, there was no significant effect of either MMS treatment or folate dietary content on splenocyte non-selected cloning efficiency. In contrast, non-selected cloning efficiency was significantly higher in MMS-treated Aag null mice than in saline treated controls (diet-gene interaction variable, p=0.04). The non-selected cloning efficiency was significantly higher in the FA+ diet than in the FA- diet group after MMS treatment of Aag null mice. Mutant frequency after MMS treatment was significantly higher in FA- wild-type and Aag null mice and in FA+ Aag null mice, but not in FA+ wild-type mice. For the Aag null mice, mutant frequency was higher in the FA+ mice than in the FA- mice after either saline or MMS treatment. These studies indicate that in wild-type mice treated with MMS, dietary folate content (FA+ or FA-) had no effect on cytotoxicity, but FA- diet increased DNA mutation frequency compared to FA+ diet. In Aag null mice, FA- diet increased the cytotoxic effects of alkylating agents but decreased the risk of DNA mutation.

Labeled Azospirillum brasilense wild type and excretion-ammonium strains in association with barley roots.

Science.gov (United States)

Santos, Adrian Richard Schenberger; Etto, Rafael Mazer; Furmam, Rafaela Wiegand; Freitas, Denis Leandro de; Santos, Karina Freire d'Eça Nogueira; Souza, Emanuel Maltempi de; Pedrosa, Fábio de Oliveira; Ayub, Ricardo Antônio; Steffens, Maria Berenice Reynaud; Galvão, Carolina Weigert

2017-09-01

Soil bacteria colonization in plants is a complex process, which involves interaction between many bacterial characters and plant responses. In this work, we labeled Azospirillum brasilense FP2 (wild type) and HM053 (excretion-ammonium) strains by insertion of the reporter gene gusA-kanamycin into the dinitrogenase reductase coding gene, nifH, and evaluated bacteria colonization in barley (Hordeum vulgare). In addition, we determined inoculation effect based on growth promotion parameters. We report an uncommon endophytic behavior of A. brasilense Sp7 derivative inside the root hair cells of barley and highlight the promising use of A. brasilense HM053 as plant growth-promoting bacterium. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Novel Escherichia coli umuD′ Mutants: Structure-Function Insights into SOS Mutagenesis

Science.gov (United States)

McLenigan, Mary; Peat, Thomas S.; Frank, Ekaterina G.; McDonald, John P.; Gonzalez, Martín; Levine, Arthur S.; Hendrickson, Wayne A.; Woodgate, Roger

1998-01-01

Although it has been 10 years since the discovery that the Escherichia coli UmuD protein undergoes a RecA-mediated cleavage reaction to generate mutagenically active UmuD′, the function of UmuD′ has yet to be determined. In an attempt to elucidate the role of UmuD′ in SOS mutagenesis, we have utilized a colorimetric papillation assay to screen for mutants of a hydroxylamine-treated, low-copy-number umuD′ plasmid that are unable to promote SOS-dependent spontaneous mutagenesis. Using such an approach, we have identified 14 independent umuD′ mutants. Analysis of these mutants revealed that two resulted from promoter changes which reduced the expression of wild-type UmuD′, three were nonsense mutations that resulted in a truncated UmuD′ protein, and the remaining nine were missense alterations. In addition to the hydroxylamine-generated mutants, we have subcloned the mutations found in three chromosomal umuD1, umuD44, and umuD77 alleles into umuD′. All 17 umuD′ mutants resulted in lower levels of SOS-dependent spontaneous mutagenesis but varied in the extent to which they promoted methyl methanesulfonate-induced mutagenesis. We have attempted to correlate these phenotypes with the potential effect of each mutation on the recently described structure of UmuD′. PMID:9721309

Attenuated stress response to acute restraint and forced swimming stress in arginine vasopressin 1b receptor subtype (Avpr1b) receptor knockout mice and wild-type mice treated with a novel Avpr1b receptor antagonist.

Science.gov (United States)

Roper, J A; Craighead, M; O'Carroll, A-M; Lolait, S J

2010-11-01

Arginine vasopressin (AVP) synthesised in the parvocellular region of the hypothalamic paraventricular nucleus and released into the pituitary portal vessels acts on the 1b receptor subtype (Avpr1b) present in anterior pituitary corticotrophs to modulate the release of adrenocorticotrophic hormone (ACTH). Corticotrophin-releasing hormone is considered the major drive behind ACTH release; however, its action is augmented synergistically by AVP. To determine the extent of vasopressinergic influence in the hypothalamic-pituitary-adrenal axis response to restraint and forced swimming stress, we compared the stress hormone levels [plasma ACTH in both stressors and corticosterone (CORT) in restraint stress only] following acute stress in mutant Avpr1b knockout (KO) mice compared to their wild-type controls following the administration of a novel Avpr1b antagonist. Restraint and forced swimming stress-induced increases in plasma ACTH were significantly diminished in mice lacking a functional Avpr1b and in wild-type mice that had been pre-treated with Avpr1b antagonist. A corresponding decrease in plasma CORT levels was also observed in acute restraint-stressed knockout male mice, and in Avpr1b-antagonist-treated male wild-type mice. By contrast, plasma CORT levels were not reduced in acutely restraint-stressed female knockout animals, or in female wild-type animals pre-treated with Avpr1b antagonist. These results demonstrate that pharmacological antagonism or inactivation of Avpr1b causes a reduction in the hypothalamic-pituitary-adrenal (HPA) axis response, particularly ACTH, to acute restraint and forced swimming stress, and show that Avpr1b knockout mice constitute a model by which to study the contribution of Avpr1b to the HPA axis response to acute stressors. © 2010 The Authors. Journal of Neuroendocrinology © 2010 Blackwell Publishing Ltd.

Hand functions in type 1 and type 2 diabetes mellitus

Directory of Open Access Journals (Sweden)

Akpinar Pinar

2017-01-01

Full Text Available Introduction/Objective. Hand functions have an enormous impact on activities of daily living in patients with diabetes mellitus (DM, such as self-care, administering insulin injections, and preparing and eating meals. The aim of the study was to evaluate hand functions and grip strength in patients with type 1 and type 2 DM. Methods. This was an observational case-control study investigating the hand functions and grip strength in patients with type 1 and type 2 DM. The study comprised 41 patients with type 1 DM aged 25–50 years sex- and age-matched, 40 non-diabetic controls, and 91 patients with type 2 DM aged 40–65 years sex- and age-matched 60 non-diabetic controls. Patients with documented history of diabetic sensorimotor neuropathy and adhesive capsulitis were excluded. The Duruoz Hand Index was used to assess the functional hand disability. Grip strength was tested with a calibrated Jamar dynamometer. Results. The Duruoz Hand Index scores in patients with type 2 DM were significantly higher than in persons in the control group (p 0.05. Grip strength values of patients with type 1 DM were significantly lower compared to those in the control group (p < 0.05, whereas there was no significant difference between patients with type 2 DM and their control group. There was a negatively significant correlation between grip strength and the Duruoz Hand Index scores in patients with both type 1 and type 2 DM (p < 0.05. Conclusion. Patients with type 1 DM and type 2 DM have different degrees of hand disability as compared to healthy control groups.

Mutations of Glucose-6-Phosphate Dehydrogenase Durham, Santa-Maria and A+ Variants Are Associated with Loss Functional and Structural Stability of the Protein

Science.gov (United States)

Gómez-Manzo, Saúl; Marcial-Quino, Jaime; Vanoye-Carlo, America; Enríquez-Flores, Sergio; De la Mora-De la Mora, Ignacio; González-Valdez, Abigail; García-Torres, Itzhel; Martínez-Rosas, Víctor; Sierra-Palacios, Edgar; Lazcano-Pérez, Fernando; Rodríguez-Bustamante, Eduardo; Arreguin-Espinosa, Roberto

2015-01-01

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy in the world. More than 160 mutations causing the disease have been identified, but only 10% of these variants have been studied at biochemical and biophysical levels. In this study we report on the functional and structural characterization of three naturally occurring variants corresponding to different classes of disease severity: Class I G6PD Durham, Class II G6PD Santa Maria, and Class III G6PD A+. The results showed that the G6PD Durham (severe deficiency), and the G6PD Santa Maria and A+ (less severe deficiency) (Class I, II and III, respectively) affect the catalytic efficiency of these enzymes, are more sensitive to temperature denaturing, and affect the stability of the overall protein when compared to the wild type WT-G6PD. In the variants, the exposure of more and buried hydrophobic pockets was induced and monitored with 8-Anilinonaphthalene-1-sulfonic acid (ANS) fluorescence, directly affecting the compaction of structure at different levels and probably reducing the stability of the protein. The degree of functional and structural perturbation by each variant correlates with the clinical severity reported in different patients. PMID:26633385

Mutations of Glucose-6-Phosphate Dehydrogenase Durham, Santa-Maria and A+ Variants Are Associated with Loss Functional and Structural Stability of the Protein

Directory of Open Access Journals (Sweden)

Saúl Gómez-Manzo

2015-12-01

Full Text Available Glucose-6-phosphate dehydrogenase (G6PD deficiency is the most common enzymopathy in the world. More than 160 mutations causing the disease have been identified, but only 10% of these variants have been studied at biochemical and biophysical levels. In this study we report on the functional and structural characterization of three naturally occurring variants corresponding to different classes of disease severity: Class I G6PD Durham, Class II G6PD Santa Maria, and Class III G6PD A+. The results showed that the G6PD Durham (severe deficiency, and the G6PD Santa Maria and A+ (less severe deficiency (Class I, II and III, respectively affect the catalytic efficiency of these enzymes, are more sensitive to temperature denaturing, and affect the stability of the overall protein when compared to the wild type WT-G6PD. In the variants, the exposure of more and buried hydrophobic pockets was induced and monitored with 8-Anilinonaphthalene-1-sulfonic acid (ANS fluorescence, directly affecting the compaction of structure at different levels and probably reducing the stability of the protein. The degree of functional and structural perturbation by each variant correlates with the clinical severity reported in different patients.

Structure of a fibronectin type III-like module from Clostridium thermocellum

International Nuclear Information System (INIS)

Alahuhta, Markus; Xu, Qi; Brunecky, Roman; Adney, William S.; Ding, Shi-You; Himmel, Michael E.; Lunin, Vladimir V.

2010-01-01

The 1.6 Å resolution structure of a fibronectin type III-like module from Clostridium thermocellum with two molecules in the asymmetric unit is reported. The 1.6 Å resolution structure of a fibronectin type III-like module from Clostridium thermocellum with two molecules in the asymmetric unit is reported. The crystals used for data collection belonged to space group P2 1 2 1 2 1 , with unit-cell parameters a = 35.43, b = 45.73, c = 107.72 Å, and the structure was refined to an R factor of 0.166. Structural comparisons found over 800 similar structures in the Protein Data Bank. The broad range of different proteins or protein domains with high structural similarity makes it especially demanding to classify these proteins. Previous studies of fibronectin type III-like modules have indicated that they might function as ligand-binding modules, as a compact form of peptide linkers or spacers between other domains, as cellulose-disrupting modules or as proteins that help large enzyme complexes remain soluble

Differential tumor biology effects of double-initiation in a mouse skin chemical carcinogenesis model comparing wild type versus protein kinase Cepsilon overexpression mice.

Science.gov (United States)

Li, Yafan; Wheeler, Deric L; Ananthaswamy, Honnavara N; Verma, Ajit K; Oberley, Terry D

2007-12-01

Our previous studies showed that protein kinase Cepsilon (PKCepsilon) verexpression in mouse skin resulted in metastatic squamous cell carcinoma (SCC) elicited by single 7,12-dimethylbenz(a)anthracene (DMBA)-initiation and 12-O-tetradecanoylphorbol-13-acetate (TPA)-promotion in the absence of preceding papilloma formation as is typically observed in wild type mice. The present study demonstrates that double-DMBA initiation modulates tumor incidence, multiplicity, and latency period in both wild type and PKCepsilon overexpression transgenic (PKCepsilon-Tg) mice. After 17 weeks (wks) of tumor promotion, a reduction in papilloma multiplicity was observed in double- versus single-DMBA initiated wild type mice. Papilloma multiplicity was inversely correlated with cell death indices of interfollicular keratinocytes, indicating decreased papilloma formation was caused by increased cell death and suggesting the origin of papillomas is in interfollicular epidermis. Double-initiated PKCepsilon-Tg mice had accelerated carcinoma formation and cancer incidence in comparison to single-initiated PKCepsilon-Tg mice. Morphologic analysis of mouse skin following double initiation and tumor promotion showed a similar if not identical series of events to those previously observed following single initiation and tumor promotion: putative preneoplastic cells were observed arising from hyperplastic hair follicles (HFs) with subsequent cancer cell infiltration into the dermis. Single-initiated PKCepsilon-Tg mice exhibited increased mitosis in epidermal cells of HFs during tumor promotion.

Gastric Medullary Carcinoma with Sporadic Mismatch Repair Deficiency and a TP53 R273C Mutation: An Unusual Case with Wild-Type BRAF

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Brett M. Lowenthal

2017-01-01

Full Text Available Medullary carcinoma has long been recognized as a subtype of colorectal cancer associated with microsatellite instability and Lynch syndrome. Gastric medullary carcinoma is a very rare neoplasm. We report a 67-year-old male who presented with a solitary gastric mass. Total gastrectomy revealed a well-demarcated, poorly differentiated carcinoma with an organoid growth pattern, pushing borders, and abundant peritumoral lymphocytic response. The prior cytology was cellular with immunohistochemical panel consistent with upper gastrointestinal/pancreaticobiliary origin. Overall, the histopathologic findings were consistent with gastric medullary carcinoma. A mismatch repair panel revealed a mismatch repair protein deficient tumor with loss of MLH1 and PMS2 expression. BRAF V600E immunostain (VE1 and BRAF molecular testing were negative, indicating a wild-type gene. Tumor sequencing of MLH1 demonstrated a wild-type gene, while our molecular panel identified TP53 c.817C>T (p.R273C mutation. These findings were compatible with a sporadic tumor. Given that morphologically identical medullary tumors often occur in Lynch syndrome, it is possible that mismatch repair loss is an early event in sporadic tumors with p53 mutation being a late event. Despite having wild-type BRAF, this tumor is sporadic and unrelated to Lynch syndrome. This case report demonstrates that coordinate ancillary studies are needed to resolve sporadic versus hereditary rare tumors.

Using ICP and micro-PIXE to investigate possible differences in the mineral composition of genetically modified versus wild-type sorghum grain

Science.gov (United States)

Ndimba, R.; Cloete, K.; Mehlo, L.; Kossmann, J.; Mtshali, C.; Pineda-Vargas, C.

2017-08-01

In the present study, possible differences in the mineral composition of transgenic versus non-transgenic sorghum grains were investigated using inductively coupled atomic emission spectroscopy (ICP-AES); and, in-tissue elemental mapping by micro Proton-Induced X-ray Emission (micro-PIXE) analysis. ICP AES was used to analyse the bulk mineral content of the wholegrain flour derived from each genotype; whilst micro-PIXE was used to interrogate localised differences in mineral composition specific to certain areas of the grain, such as the bran layer and the central endosperm tissue. According to the results obtained, no significant difference in the average Fe, Zn or Ca content was found to differentiate the transgenic from the wild-type grain using ICP-AES. However, using micro-PIXE, a significant reduction in zinc could be detected in the bran layer of the transgenic grains relative to wild-type. Although it is difficult to draw firm conclusions, as a result of the small sample size used in this study, micro-PIXE has nonetheless proven itself as a useful technique for highlighting the possibility that there may be reduced levels of zinc accumulation in the bran layer of the transgenic grains. Given that the genetic modification targets proteins that are highly concentrated in certain parts of the bran tissue, it seems plausible that the reduced levels of zinc may be an unintended consequence of the silencing of kafirin proteins. Although no immediate health or nutritional concerns emerge from this preliminary finding, it is noted that zinc plays an important biological role within this part of the grain as a structural stabiliser and antioxidant factor. Further study is therefore needed to assess more definitively the extent of the apparent localised reduction in zinc in the transgenic grains and how this may affect other important grain quality characteristics.

IFN-{gamma} enhances neurogenesis in wild-type mice and in a mouse model of Alzheimer's disease

DEFF Research Database (Denmark)

Baron, Rona; Nemirovsky, Anna; Harpaz, Idan

2008-01-01

the spatial learning and memory performance of the animals. In older mice, the effect of IFN-gamma is more pronounced in both wild-type mice and mice with Alzheimer's-like disease and is associated with neuroprotection. In addition, IFN-gamma reverses the increase in oligodendrogenesis observed in a mouse...... mechanisms can generate immunity to such deficits in neuronal repair. We demonstrate that in contrast to primarily innate immunity cytokines, such as interleukin-6 and tumor necrosis factor-alpha, the adaptive immunity cytokine IFN-gamma enhances neurogenesis in the dentate gyrus of adult mice and improves...

OryzaGenome: Genome Diversity Database of Wild Oryza Species

KAUST Repository

Ohyanagi, Hajime

2015-11-18

The species in the genus Oryza, encompassing nine genome types and 23 species, are a rich genetic resource and may have applications in deeper genomic analyses aiming to understand the evolution of plant genomes. With the advancement of next-generation sequencing (NGS) technology, a flood of Oryza species reference genomes and genomic variation information has become available in recent years. This genomic information, combined with the comprehensive phenotypic information that we are accumulating in our Oryzabase, can serve as an excellent genotype-phenotype association resource for analyzing rice functional and structural evolution, and the associated diversity of the Oryza genus. Here we integrate our previous and future phenotypic/habitat information and newly determined genotype information into a united repository, named OryzaGenome, providing the variant information with hyperlinks to Oryzabase. The current version of OryzaGenome includes genotype information of 446 O. rufipogon accessions derived by imputation and of 17 accessions derived by imputation-free deep sequencing. Two variant viewers are implemented: SNP Viewer as a conventional genome browser interface and Variant Table as a textbased browser for precise inspection of each variant one by one. Portable VCF (variant call format) file or tabdelimited file download is also available. Following these SNP (single nucleotide polymorphism) data, reference pseudomolecules/ scaffolds/contigs and genome-wide variation information for almost all of the closely and distantly related wild Oryza species from the NIG Wild Rice Collection will be available in future releases. All of the resources can be accessed through http://viewer.shigen.info/oryzagenome/.

Wild-type phosphoribosylpyrophosphate synthase (PRS from Mycobacterium tuberculosis: a bacterial class II PRS?

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Ardala Breda

Full Text Available The 5-phospho-α-D-ribose 1-diphosphate (PRPP metabolite plays essential roles in several biosynthetic pathways, including histidine, tryptophan, nucleotides, and, in mycobacteria, cell wall precursors. PRPP is synthesized from α-D-ribose 5-phosphate (R5P and ATP by the Mycobacterium tuberculosis prsA gene product, phosphoribosylpyrophosphate synthase (MtPRS. Here, we report amplification, cloning, expression and purification of wild-type MtPRS. Glutaraldehyde cross-linking results suggest that MtPRS predominates as a hexamer, presenting varied oligomeric states due to distinct ligand binding. MtPRS activity measurements were carried out by a novel coupled continuous spectrophotometric assay. MtPRS enzyme activity could be detected in the absence of P(i. ADP, GDP and UMP inhibit MtPRS activity. Steady-state kinetics results indicate that MtPRS has broad substrate specificity, being able to accept ATP, GTP, CTP, and UTP as diphosphoryl group donors. Fluorescence spectroscopy data suggest that the enzyme mechanism for purine diphosphoryl donors follows a random order of substrate addition, and for pyrimidine diphosphoryl donors follows an ordered mechanism of substrate addition in which R5P binds first to free enzyme. An ordered mechanism for product dissociation is followed by MtPRS, in which PRPP is the first product to be released followed by the nucleoside monophosphate products to yield free enzyme for the next round of catalysis. The broad specificity for diphosphoryl group donors and detection of enzyme activity in the absence of P(i would suggest that MtPRS belongs to Class II PRS proteins. On the other hand, the hexameric quaternary structure and allosteric ADP inhibition would place MtPRS in Class I PRSs. Further data are needed to classify MtPRS as belonging to a particular family of PRS proteins. The data here presented should help augment our understanding of MtPRS mode of action. Current efforts are toward experimental structure

Wild-Type Phosphoribosylpyrophosphate Synthase (PRS) from Mycobacterium tuberculosis: A Bacterial Class II PRS?

Science.gov (United States)

Breda, Ardala; Martinelli, Leonardo K. B.; Bizarro, Cristiano V.; Rosado, Leonardo A.; Borges, Caroline B.; Santos, Diógenes S.; Basso, Luiz A.

2012-01-01

The 5-phospho-α-D-ribose 1-diphosphate (PRPP) metabolite plays essential roles in several biosynthetic pathways, including histidine, tryptophan, nucleotides, and, in mycobacteria, cell wall precursors. PRPP is synthesized from α-D-ribose 5-phosphate (R5P) and ATP by the Mycobacterium tuberculosis prsA gene product, phosphoribosylpyrophosphate synthase (MtPRS). Here, we report amplification, cloning, expression and purification of wild-type MtPRS. Glutaraldehyde cross-linking results suggest that MtPRS predominates as a hexamer, presenting varied oligomeric states due to distinct ligand binding. MtPRS activity measurements were carried out by a novel coupled continuous spectrophotometric assay. MtPRS enzyme activity could be detected in the absence of Pi. ADP, GDP and UMP inhibit MtPRS activity. Steady-state kinetics results indicate that MtPRS has broad substrate specificity, being able to accept ATP, GTP, CTP, and UTP as diphosphoryl group donors. Fluorescence spectroscopy data suggest that the enzyme mechanism for purine diphosphoryl donors follows a random order of substrate addition, and for pyrimidine diphosphoryl donors follows an ordered mechanism of substrate addition in which R5P binds first to free enzyme. An ordered mechanism for product dissociation is followed by MtPRS, in which PRPP is the first product to be released followed by the nucleoside monophosphate products to yield free enzyme for the next round of catalysis. The broad specificity for diphosphoryl group donors and detection of enzyme activity in the absence of Pi would suggest that MtPRS belongs to Class II PRS proteins. On the other hand, the hexameric quaternary structure and allosteric ADP inhibition would place MtPRS in Class I PRSs. Further data are needed to classify MtPRS as belonging to a particular family of PRS proteins. The data here presented should help augment our understanding of MtPRS mode of action. Current efforts are toward experimental structure determination of Mt

Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5Δ32 mutation confers resistance to HIV infection.

Science.gov (United States)

Ye, Lin; Wang, Jiaming; Beyer, Ashley I; Teque, Fernando; Cradick, Thomas J; Qi, Zhongxia; Chang, Judy C; Bao, Gang; Muench, Marcus O; Yu, Jingwei; Levy, Jay A; Kan, Yuet Wai

2014-07-01

Individuals homozygous for the C-C chemokine receptor type 5 gene with 32-bp deletions (CCR5Δ32) are resistant to HIV-1 infection. In this study, we generated induced pluripotent stem cells (iPSCs) homozygous for the naturally occurring CCR5Δ32 mutation through genome editing of wild-type iPSCs using a combination of transcription activator-like effector nucleases (TALENs) or RNA-guided clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 together with the piggyBac technology. Remarkably, TALENs or CRISPR-Cas9-mediated double-strand DNA breaks resulted in up to 100% targeting of the colonies on one allele of which biallelic targeting occurred at an average of 14% with TALENs and 33% with CRISPR. Excision of the piggyBac using transposase seamlessly reproduced exactly the naturally occurring CCR5Δ32 mutation without detectable exogenous sequences. We differentiated these modified iPSCs into monocytes/macrophages and demonstrated their resistance to HIV-1 challenge. We propose that this strategy may provide an approach toward a functional cure of HIV-1 infection.

Structure and Function of p97 and Pex1/6 Type II AAA+ Complexes.

Science.gov (United States)

Saffert, Paul; Enenkel, Cordula; Wendler, Petra

2017-01-01

Protein complexes of the Type II AAA+ (ATPases associated with diverse cellular activities) family are typically hexamers of 80-150 kDa protomers that harbor two AAA+ ATPase domains. They form double ring assemblies flanked by associated domains, which can be N-terminal, intercalated or C-terminal to the ATPase domains. Most prominent members of this family include NSF (N-ethyl-maleimide sensitive factor), p97/VCP (valosin-containing protein), the Pex1/Pex6 complex and Hsp104 in eukaryotes and ClpB in bacteria. Tremendous efforts have been undertaken to understand the conformational dynamics of protein remodeling type II AAA+ complexes. A uniform mode of action has not been derived from these works. This review focuses on p97/VCP and the Pex1/6 complex, which both structurally remodel ubiquitinated substrate proteins. P97/VCP plays a role in many processes, including ER- associated protein degradation, and the Pex1/Pex6 complex dislocates and recycles the transport receptor Pex5 from the peroxisomal membrane during peroxisomal protein import. We give an introduction into existing knowledge about the biochemical and cellular activities of the complexes before discussing structural information. We particularly emphasize recent electron microscopy structures of the two AAA+ complexes and summarize their structural differences.

A PCR-based genotyping method to distinguish between wild-type and ornamental varieties of Imperata cylindrica.

Science.gov (United States)

Cseke, Leland J; Talley, Sharon M

2012-02-20

Wild-type I. cylindrica (cogongrass) is one of the top ten worst invasive plants in the world, negatively impacting agricultural and natural resources in 73 different countries throughout Africa, Asia, Europe, New Zealand, Oceania and the Americas(1-2). Cogongrass forms rapidly-spreading, monodominant stands that displace a large variety of native plant species and in turn threaten the native animals that depend on the displaced native plant species for forage and shelter. To add to the problem, an ornamental variety [I. cylindrica var. koenigii (Retzius)] is widely marketed under the names of Imperata cylindrica 'Rubra', Red Baron, and Japanese blood grass (JBG). This variety is putatively sterile and noninvasive and is considered a desirable ornamental for its red-colored leaves. However, under the correct conditions, JBG can produce viable seed (Carol Holko, 2009 personal communication) and can revert to a green invasive form that is often indistinguishable from cogongrass as it takes on the distinguishing characteristics of the wild-type invasive variety(4) (Figure 1). This makes identification using morphology a difficult task even for well-trained plant taxonomists. Reversion of JBG to an aggressive green phenotype is also not a rare occurrence. Using sequence comparisons of coding and variable regions in both nuclear and chloroplast DNA, we have confirmed that JBG has reverted to the green invasive within the states of Maryland, South Carolina, and Missouri. JBG has been sold and planted in just about every state in the continental U.S. where there is not an active cogongrass infestation. The extent of the revert problem in not well understood because reverted plants are undocumented and often destroyed. Application of this molecular protocol provides a method to identify JBG reverts and can help keep these varieties from co-occurring and possibly hybridizing. Cogongrass is an obligate outcrosser and, when crossed with a different genotype, can produce

Differential Activity of the Oral Glucan Synthase Inhibitor SCY-078 against Wild-Type and Echinocandin-Resistant Strains of Candida Species.

Science.gov (United States)

Pfaller, Michael A; Messer, Shawn A; Rhomberg, Paul R; Borroto-Esoda, Katyna; Castanheira, Mariana

2017-08-01

SCY-078 (formerly MK-3118) is a novel orally active inhibitor of fungal β-(1,3)-glucan synthase (GS). SCY-078 is a derivative of enfumafungin and is structurally distinct from the echinocandin class of antifungal agents. We evaluated the in vitro activity of this compound against wild-type (WT) and echinocandin-resistant isolates containing mutations in the FKS genes of Candida spp. Against 36 Candida spp. FKS mutants tested, 30 (83.3%) were non-WT to 1 or more echinocandins, and only 9 (25.0%) were non-WT (MIC, >WT-upper limit) to SCY-078. Among C. glabrata isolates carrying FKS alterations, 84.0% were non-WT to the echinocandins versus only 24.0% for SCY-078. In contrast to the echinocandin comparators, the activity of SCY-078 was minimally affected by the presence of FKS mutations, suggesting that this agent is useful in the treatment of Candida infections due to echinocandin-resistant strains. Copyright © 2017 American Society for Microbiology.

CYCLOPHILIN A: STRUCTURE AND FUNCTIONS

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A. A. Kalinina

2017-01-01

Full Text Available Cyclophilins belong to a large family of ancient conservative proteins with peptidyl-prolyl-cis-trans isomerase activity. The main member of this family – cyclophilin A – was discovered as an intracellular ligand for cyclosporine A. Further investigations revealed a wide range of functions of cyclophilin A. Cyclophilin A is involved in T-cell signaling, it takes part in folding, assembly and intracellular transport of proteins, as well as acts as an antioxidant. Different cell types secrete cyclophilin A under infection or oxidative stress. Cyclophilin A is one of the main factors involved in inflammation and pathogenesis of autoimmune, cardiovascular and other diseases. This protein is thought to take part in tumor progression. In this review we describe the structure of cyclophilin A and its main known functions in health and disease.

Fructose 6-phosphate phosphoketolase activity in wild-type strains of Lactobacillus, isolated from the intestinal tract of pigs.

Science.gov (United States)

Bolado-Martínez, E; Acedo-Félix, E; Peregrino-Uriarte, A B; Yepiz-Plascencia, G

2012-01-01

Phosphoketolases are key enzymes of the phosphoketolase pathway of heterofermentative lactic acid bacteria, which include lactobacilli. In heterofermentative lactobacilli xylulose 5-phosphate phosphoketolase (X5PPK) is the main enzyme of the phosphoketolase pathway. However, activity of fructose 6-phosphate phosphoketolase (F6PPK) has always been considered absent in lactic acid bacteria. In this study, the F6PPK activity was detected in 24 porcine wild-type strains of Lactobacillus reuteri and Lactobacillus mucosae, but not in the Lactobacillus salivarius or in L. reuteri ATCC strains. The activity of F6PPK increased after treatment of the culture at low-pH and diminished after porcine bile-salts stress conditions in wild-type strains of L. reuteri. Colorimetric quantification at 505 nm allowed to differentiate between microbial strains with low activity and without the activity of F6PPK. Additionally, activity of F6PPK and the X5PPK gene expression levels were evaluated by real time PCR, under stress and nonstress conditions, in 3 L. reuteri strains. Although an exact correlation, between enzyme activity and gene expression was not obtained, it remains possible that the xpk gene codes for a phosphoketolase with dual substrate, at least in the analyzed strains of L. reuteri.

Diversity, assembly and regulation of archaeal type IV pili-like and non-type-IV pili-like surface structures.

Science.gov (United States)

Lassak, Kerstin; Ghosh, Abhrajyoti; Albers, Sonja-Verena

2012-01-01

Archaea have evolved fascinating surface structures allowing rapid adaptation to changing environments. The archaeal surface appendages display such diverse biological roles as motility, adhesion, biofilm formation, exchange of genetic material and species-specific interactions and, in turn, increase fitness of the cells. Intriguingly, despite sharing the same functions with their bacterial counterparts, the assembly mechanism of many archaeal surface structures is rather related to assembly of bacterial type IV pili. This review summarizes our state-of-the-art knowledge about unique structural and biochemical properties of archaeal surface appendages with a particular focus on archaeal type IV pili-like structures. The latter comprise not only widely distributed archaella (formerly known as archaeal flagella), but also different highly specialized archaeal pili, which are often restricted to certain species. Recent findings regarding assembly mechanisms, structural aspects and physiological roles of these type IV pili-like structures will be discussed in detail. Recently, first regulatory proteins involved in transition from both planktonic to sessile lifestyle and in assembly of archaella were identified. To conclude, we provide novel insights into regulatory mechanisms underlying the assembly of archaeal surface structures. Copyright © 2012. Published by Elsevier Masson SAS.

General anesthetic octanol and related compounds activate wild-type and delF508 cystic fibrosis chloride channels

OpenAIRE

Marcet, Brice; Becq, Frédéric; Norez, Caroline; Delmas, Patrick; Verrier, Bernard

2004-01-01

Cystic fibrosis transmembrane conductance regulator (CFTR) Cl− channel is defective during cystic fibrosis (CF). Activators of the CFTR Cl− channel may be useful for therapy of CF. Here, we demonstrate that a range of general anesthetics like normal-alkanols (n-alkanols) and related compounds can stimulate the Cl− channel activity of wild-type CFTR and delF508-CFTR mutant.The effects of n-alkanols like octanol on CFTR activity were measured by iodide (125I) efflux and patch-clamp techniques o...

Wild-type male offspring of fmr-1+/- mothers exhibit characteristics of the fragile X phenotype.

Science.gov (United States)

Zupan, Bojana; Toth, Miklos

2008-10-01

Fragile X syndrome is an X-linked disorder caused by the inactivation of the FMR-1 gene with symptoms ranging from impaired cognitive functions to seizures, anxiety, sensory abnormalities, and hyperactivity. Males are more severely affected than heterozygote (H) females, who, as carriers, have a 50% chance of transmitting the mutated allele in each pregnancy. fmr-1 knockout (KO) mice reproduce fragile X symptoms, including hyperactivity, seizures, and abnormal sensory processing. In contrast to the expectation that wild-type (WT) males born to H (fmr-1(+/-)) mothers (H>WT) are behaviorally normal and indistinguishable from WT males born to WT mothers (WT>WT); here, we show that H>WT offspring are more active than WT>WT offspring and that their hyperactivity is similar to male KO mice born to H or KO (fmr-1(-/-)) mothers (H>KO/KO>KO). H>WT mice, however, do not exhibit seizures or abnormal sensory processing. Consistent with their hyperactivity, the effect of the D2 agonist quinpirole is reduced in H>WT as well as in H>KO and KO>KO mice compared to WT>WT offspring, suggesting a diminished feedback inhibition of dopamine release. Our data indicate that some aspects of hyperactivity and associated dopaminergic changes in 'fragile X' mice are a maternal fmr-1 genotype rather than an offspring fmr-1 genotype effect.

A new functional and structural generation of JK edge-triggered flip-flops

International Nuclear Information System (INIS)

Stefanescu, I.

1977-01-01

A new type of logical structure for a JK edge-triggered flip-flop is proposed by the author. The structure facilitates flip-flop realizations, named ''jk-JK edge-triggered flip-flops'', satisfying more functional requirements, and offering an increased flexibility in logical design, with respect to the conventional JK edge-triggered flip-flops. The function of new flip-flops covers the function of JK edge-triggered flip-flops, known as integrated circuits. (author)

Molecular analysis of mutant and wild type alcohol dehydrogenase alleles from Drosophila

International Nuclear Information System (INIS)

Batzer, M.A.

1988-01-01

Wild type alcohol dehydrogenase polypeptides (ADH) from Drosophila melanogaster transformants were examined using western blots and polyclonal antiserum specific for Drosophila melanogaster ADH. Mutants induced in Drosophila spermatozoa at the alcohol dehydrogenase (Adh) locus using X-rays, 1-ethyl-1-nitrosourea (ENU) or ethyl methanesulfonate (EMS) were characterized using genetic complementation tests, western blots, Southern blots, northern blots and enzymatic amplification of the Adh locus. Genetic complementation tests showed that 22/30 X-ray-induced mutants, and 3/13 ENU and EMS induced mutants were multi-locus deficiencies. Western blot analysis of the intragenic mutations showed that 4/7 X-ray-induced mutants produced detectable polypeptides, one of which was normal in molecular weight and charge. In contrast 8/10 intragenic ENU and EMS induced mutants produced normal polypeptides. Southern blot analysis showed that 5/7 intragenic X-ray induced mutants and all 10 of the intragenic ENU and EMS induced mutants were normal with respect to the alleles they were derived from

Characterization of global yeast quantitative proteome data generated from the wild-type and glucose repression Saccharomyces cerevisiae strains: The comparison of two quantitative methods

DEFF Research Database (Denmark)

Usaite, Renata; Wohlschlegel, James; Venable, John D.

2008-01-01

The quantitative proteomic analysis of complex protein mixtures is emerging as a technically challenging but viable systems-level approach for studying cellular function. This study presents a large-scale comparative analysis of protein abundances from yeast protein lysates derived from both wild......-type yeast and yeast strains lacking key components of the Snf1 kinase complex. Four different strains were grown under well-controlled chemostat conditions. Multidimensional protein identification technology followed by quantitation using either spectral counting or stable isotope labeling approaches...... labeling strategy. The stable isotope labeling based quantitative approach was found to be highly reproducible among biological replicates when complex protein mixtures containing small expression changes were analyzed. Where poor correlation between stable isotope labeling and spectral counting was found...

Evolution and Adaptation of Wild Emmer Wheat Populations to Biotic and Abiotic Stresses.

Science.gov (United States)

Huang, Lin; Raats, Dina; Sela, Hanan; Klymiuk, Valentina; Lidzbarsky, Gabriel; Feng, Lihua; Krugman, Tamar; Fahima, Tzion

2016-08-04

The genetic bottlenecks associated with plant domestication and subsequent selection in man-made agroecosystems have limited the genetic diversity of modern crops and increased their vulnerability to environmental stresses. Wild emmer wheat, the tetraploid progenitor of domesticated wheat, distributed along a wide range of ecogeographical conditions in the Fertile Crescent, has valuable "left behind" adaptive diversity to multiple diseases and environmental stresses. The biotic and abiotic stress responses are conferred by series of genes and quantitative trait loci (QTLs) that control complex resistance pathways. The study of genetic diversity, genomic organization, expression profiles, protein structure and function of biotic and abiotic stress-resistance genes, and QTLs could shed light on the evolutionary history and adaptation mechanisms of wild emmer populations for their natural habitats. The continuous evolution and adaptation of wild emmer to the changing environment provide novel solutions that can contribute to safeguarding food for the rapidly growing human population.

Hyperspectral remote sensing of wild oyster reefs

Science.gov (United States)

Le Bris, Anthony; Rosa, Philippe; Lerouxel, Astrid; Cognie, Bruno; Gernez, Pierre; Launeau, Patrick; Robin, Marc; Barillé, Laurent

2016-04-01

The invasion of the wild oyster Crassostrea gigas along the western European Atlantic coast has generated changes in the structure and functioning of intertidal ecosystems. Considered as an invasive species and a trophic competitor of the cultivated conspecific oyster, it is now seen as a resource by oyster farmers following recurrent mass summer mortalities of oyster spat since 2008. Spatial distribution maps of wild oyster reefs are required by local authorities to help define management strategies. In this work, visible-near infrared (VNIR) hyperspectral and multispectral remote sensing was investigated to map two contrasted intertidal reef structures: clusters of vertical oysters building three-dimensional dense reefs in muddy areas and oysters growing horizontally creating large flat reefs in rocky areas. A spectral library, collected in situ for various conditions with an ASD spectroradiometer, was used to run Spectral Angle Mapper classifications on airborne data obtained with an HySpex sensor (160 spectral bands) and SPOT satellite HRG multispectral data (3 spectral bands). With HySpex spectral/spatial resolution, horizontal oysters in the rocky area were correctly classified but the detection was less efficient for vertical oysters in muddy areas. Poor results were obtained with the multispectral image and from spatially or spectrally degraded HySpex data, it was clear that the spectral resolution was more important than the spatial resolution. In fact, there was a systematic mud deposition on shells of vertical oyster reefs explaining the misclassification of 30% of pixels recognized as mud or microphytobenthos. Spatial distribution maps of oyster reefs were coupled with in situ biomass measurements to illustrate the interest of a remote sensing product to provide stock estimations of wild oyster reefs to be exploited by oyster producers. This work highlights the interest of developing remote sensing techniques for aquaculture applications in coastal

Location of Primary Tumor and Benefit From Anti-Epidermal Growth Factor Receptor Monoclonal Antibodies in Patients With RAS and BRAF Wild-Type Metastatic Colorectal Cancer

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Moretto, Roberto; Cremolini, Chiara; Rossini, Daniele; Pietrantonio, Filippo; Battaglin, Francesca; Mennitto, Alessia; Bergamo, Francesca; Loupakis, Fotios; Marmorino, Federica; Berenato, Rosa; Marsico, Valentina Angela; Caporale, Marta; Antoniotti, Carlotta; Masi, Gianluca; Salvatore, Lisa; Borelli, Beatrice; Fontanini, Gabriella; Lonardi, Sara; De Braud, Filippo

2016-01-01

Introduction. Right- and left-sided colorectal cancers (CRCs) differ in clinical and molecular characteristics. Some retrospective analyses suggested that patients with right-sided tumors derive less benefit from anti-epidermal growth factor receptor (EGFR) antibodies; however, molecular selection in those studies was not extensive. Patients and Methods. Patients with RAS and BRAF wild-type metastatic CRC (mCRC) who were treated with single-agent anti-EGFRs or with cetuximab-irinotecan (if refractory to previous irinotecan) were included in the study. Differences in outcome between patients with right- and left-sided tumors were investigated. Results. Of 75 patients, 14 and 61 had right- and left-sided tumors, respectively. None of the right-sided tumors responded according to RECIST, compared with 24 left-sided tumors (overall response rate: 0% vs. 41%; p = .0032), and only 2 patients with right-sided tumors (15%) versus 47 patients with left-sided tumors (80%) achieved disease control (p < .0001). The median duration of progression-free survival was 2.3 and 6.6 months in patients with right-sided and left-sided tumors, respectively (hazard ratio: 3.97; 95% confidence interval: 2.09–7.53; p < .0001). Conclusion. Patients with right-sided RAS and BRAF wild-type mCRC seemed to derive no benefit from single-agent anti-EGFRs. Implications for Practice: Right- and left-sided colorectal tumors have peculiar epidemiological and clinicopathological characteristics, distinct gene expression profiles and genetic alterations, and different prognoses. This study assessed the potential predictive impact of primary tumor site with regard to anti-epidermal growth factor receptor (EGFR) monoclonal antibody treatment in patients with RAS and BRAF wild-type metastatic colorectal cancer. The results demonstrated the lack of activity of anti-EGFRs in RAS and BRAF wild-type, right-sided tumors, thus suggesting a potential role for primary tumor location in driving treatment choices

Data Presentation Structures in Specialised Dictionaries: Law Dictionaries with Communicative Functions

DEFF Research Database (Denmark)

Nielsen, Sandro

2015-01-01

data stand out, lexicographers should prioritize functional data that are directly related to and support the function(s) of dictionaries on a need-to-have/nice-to have basis, because data presentation structures with functional focus may better help users achieve their intended goals, i.e. finding......Theoretical lexicographers have developed a range of elaborate structures to describe the arrangement of data inside dictionaries, in particular in dictionary articles. However, most of these structures have been developed on the basis of detailed analyses of print dictionaries and relatively...... little has been said about the arrangement of data in e-dictionaries. The relevant data types are lexicographical data providing help concerning the function(s) and use of dictionaries on search results pages. In order to create a visual hierarchy on screen that makes the most important search result...

Draft genome sequence of Cicer reticulatum L., the wild progenitor of chickpea provides a resource for agronomic trait improvement.

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Gupta, Sonal; Nawaz, Kashif; Parween, Sabiha; Roy, Riti; Sahu, Kamlesh; Kumar Pole, Anil; Khandal, Hitaishi; Srivastava, Rishi; Kumar Parida, Swarup; Chattopadhyay, Debasis

2017-02-01

Cicer reticulatum L. is the wild progenitor of the fourth most important legume crop chickpea (C. arietinum L.). We assembled short-read sequences into 416 Mb draft genome of C. reticulatum and anchored 78% (327 Mb) of this assembly to eight linkage groups. Genome annotation predicted 25,680 protein-coding genes covering more than 90% of predicted gene space. The genome assembly shared a substantial synteny and conservation of gene orders with the genome of the model legume Medicago truncatula. Resistance gene homologs of wild and domesticated chickpeas showed high sequence homology and conserved synteny. Comparison of gene sequences and nucleotide diversity using 66 wild and domesticated chickpea accessions suggested that the desi type chickpea was genetically closer to the wild species than the kabuli type. Comparative analyses predicted gene flow between the wild and the cultivated species during domestication. Molecular diversity and population genetic structure determination using 15,096 genome-wide single nucleotide polymorphisms revealed an admixed domestication pattern among cultivated (desi and kabuli) and wild chickpea accessions belonging to three population groups reflecting significant influence of parentage or geographical origin for their cultivar-specific population classification. The assembly and the polymorphic sequence resources presented here would facilitate the study of chickpea domestication and targeted use of wild Cicer germplasms for agronomic trait improvement in chickpea. © The Author 2016. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

Two skin cell lines from wild-type and albino Japanese flounder (Paralichthys olivaceus): establishment, characterization, virus susceptibility, efficient transfection, and application to albinism study.

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Wang, Ruoqing; Zhang, Nianwei; Wang, Renkai; Wang, Shengpeng; Wang, Na

2017-12-01

In order to provide an applicable cell platform to study fish pathology and skin pigmentation, two cell lines derived from skin tissues of wild-type and albino Japanese flounder were established and named JFSK_wt and JFSK_alb, respectively. These two cell lines were cultured for 45 passages within approximately 300 days. JFSK_wt and JFSK_alb cells were maintained in Dulbecco's Modified Eagle's Medium and Ham's F-12 Nutrient Mixture (DMEM/F12) supplemented with antibiotics, fetal bovine serum (FBS), 2-mercaptoethanol (2-Me), N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES), and basic fibroblast growth factor (bFGF). The optimal growth temperature for JFSK_wt and JFSK_alb cells was 24 °C, and microscopically, the two cell lines were composed of fibroblast-like cells. Chromosomal analysis revealed that JFSK_wt and JFSK_alb cells had an identical diploid karyotype with 2n = 48t. Results of viral inoculation assays revealed that both cell lines shared similar patterns of viral susceptibility to nervous necrosis virus (NNV). High transfection efficiency was observed in JFSK_wt and JFSK_alb cells transfected with a pEGFP-N3 reporter plasmid and Cy3-siRNA. The detection of dermal marker Dermo-1 showed that these two cells were both derived from the dermis. Finally, three genes involved in the melanogenesis pathway, including adenylate cyclase type 5 (adcy5), microphthalmia-associated transcription factor (mitf), and endothelin B receptor (ednrb), were downregulated in JFSK_alb versus JFSK_wt cells. Thus, the two cell lines, sampled from skin tissue of wild-type and albino Japanese flounder will be not only helpful for fish pathogen research but also beneficial for albinism-related gene function studies.

The common structural architecture of Shigella flexneri and Salmonella typhimurium type three secretion needles.

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Jean-Philippe Demers

2013-03-01

Full Text Available The Type Three Secretion System (T3SS, or injectisome, is a macromolecular infection machinery present in many pathogenic Gram-negative bacteria. It consists of a basal body, anchored in both bacterial membranes, and a hollow needle through which effector proteins are delivered into the target host cell. Two different architectures of the T3SS needle have been previously proposed. First, an atomic model of the Salmonella typhimurium needle was generated from solid-state NMR data. The needle subunit protein, PrgI, comprises a rigid-extended N-terminal segment and a helix-loop-helix motif with the N-terminus located on the outside face of the needle. Second, a model of the Shigella flexneri needle was generated from a high-resolution 7.7-Å cryo-electron microscopy density map. The subunit protein, MxiH, contains an N-terminal α-helix, a loop, another α-helix, a 14-residue-long β-hairpin (Q51-Q64 and a C-terminal α-helix, with the N-terminus facing inward to the lumen of the needle. In the current study, we carried out solid-state NMR measurements of wild-type Shigella flexneri needles polymerized in vitro and identified the following secondary structure elements for MxiH: a rigid-extended N-terminal segment (S2-T11, an α-helix (L12-A38, a loop (E39-P44 and a C-terminal α-helix (Q45-R83. Using immunogold labeling in vitro and in vivo on functional needles, we located the N-terminus of MxiH subunits on the exterior of the assembly, consistent with evolutionary sequence conservation patterns and mutagenesis data. We generated a homology model of Shigella flexneri needles compatible with both experimental data: the MxiH solid-state NMR chemical shifts and the state-of-the-art cryoEM density map. These results corroborate the solid-state NMR structure previously solved for Salmonella typhimurium PrgI needles and establish that Shigella flexneri and Salmonella typhimurium subunit proteins adopt a conserved structure and orientation in their

Genetic diversity and population structure in cultivated sunflower and a comparison to its wild progenitor, Helianthus annuus L.

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Mandel, J R; Dechaine, J M; Marek, L F; Burke, J M

2011-09-01

Crop germplasm collections are valuable resources for ongoing plant breeding efforts. To fully utilize such collections, however, researchers need detailed information about the amount and distribution of genetic diversity present within collections. Here, we report the results of a population genetic analysis of the primary gene pool of sunflower (Helianthus annuus L.) based on a broad sampling of 433 cultivated accessions from North America and Europe, as well as a range-wide collection of 24 wild sunflower populations. Gene diversity across the cultivars was 0.47, as compared with 0.70 in the wilds, indicating that cultivated sunflower harbors roughly two-thirds of the total genetic diversity present in wild sunflower. Population structure analyses revealed that wild sunflower can be subdivided into four genetically distinct population clusters throughout its North American range, whereas the cultivated sunflower gene pool could be split into two main clusters separating restorer lines from the balance of the gene pool. Use of a maximum likelihood method to estimate the contribution of the wild gene pool to the cultivated sunflower germplasm revealed that the bulk of the cultivar diversity is derived from two wild sunflower population genetic clusters that are primarily composed of individuals from the east-central United States, the same general region in which sunflower domestication is believed to have occurred. We also identified a nested subset of accessions that capture as much of the allelic diversity present within the sampled cultivated sunflower germplasm collection as possible. At the high end, a core set of 288 captured nearly 90% of the alleles present in the full set of 433, whereas a core set of just 12 accessions was sufficient to capture nearly 50% of the total allelic diversity present within this sample of cultivated sunflower.

Development of Colle-Salvetti type electron-nucleus correlation functional for MC-DFT

Energy Technology Data Exchange (ETDEWEB)

Udagawa, Taro [Department of Chemistry and Biomolecuar Science, Faculty of Engineering, Gifu University, Yanagido 1-1, Gifu 501-1193 (Japan); Tsuneda, Takao [Fuel Cell Nanomaterials Center, University of Yamanashi, Miyamae-cho 6-43, Kofu 400-0021 (Japan); Tachikawa, Masanori [Quqnatum Chemistry Division, Graduate School of Science, Yokohama City University, Seto 22-2, Kanazawa, Yokohama 236-0027 (Japan)

2015-12-31

A Colle-Salvetti type electron-nucleus correlation functional for multicomponent density-functional theory is proposed. We demonstrate that our correlation functional quantitatively reproduces the quantum nuclear effects of protons; the mean absolute deviation value is 2.8 millihartrees for the optimized structure of hydrogen-containing molecules. We also show other practical calculations with our new electron-deuteron and electron-triton correlation functionals. Since this functional is derived without any unphysical assumption, the strategy taken in this development will be a promising recipe to make new functionals for the potentials of other particles’ interactions.

Anti-tumor activity of high-dose EGFR tyrosine kinase inhibitor and sequential docetaxel in wild type EGFR non-small cell lung cancer cell nude mouse xenografts

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Tang, Ning; Zhang, Qianqian; Fang, Shu; Han, Xiao; Wang, Zhehai

2017-01-01

Treatment of non-small-cell lung cancer (NSCLC) with wild-type epidermal growth factor receptor (EGFR) is still a challenge. This study explored antitumor activity of high-dose icotinib (an EGFR tyrosine kinase inhibitor) plus sequential docetaxel against wild-type EGFR NSCLC cells-generated nude mouse xenografts. Nude mice were subcutaneously injected with wild-type EGFR NSCLC A549 cells and divided into different groups for 3-week treatment. Tumor xenograft volumes were monitored and recorded, and at the end of experiments, tumor xenografts were removed for Western blot and immunohistochemical analyses. Compared to control groups (negative control, regular-dose icotinib [IcoR], high-dose icotinib [IcoH], and docetaxel [DTX]) and regular icotinib dose (60 mg/kg) with docetaxel, treatment of mice with a high-dose (1200 mg/kg) of icotinib plus sequential docetaxel for 3 weeks (IcoH-DTX) had an additive effect on suppression of tumor xenograft size and volume (P Icotinib-containing treatments markedly reduced phosphorylation of EGFR, mitogen activated protein kinase (MAPK), and protein kinase B (Akt), but only the high-dose icotinib-containing treatments showed an additive effect on CD34 inhibition (P icotinib plus docetaxel had a similar effect on mouse weight loss (a common way to measure adverse reactions in mice), compared to the other treatment combinations. The study indicate that the high dose of icotinib plus sequential docetaxel (IcoH-DTX) have an additive effect on suppressing the growth of wild-type EGFR NSCLC cell nude mouse xenografts, possibly through microvessel density reduction. Future clinical trials are needed to confirm the findings of this study. PMID:27852073

Brain activation by short-term nicotine exposure in anesthetized wild-type and beta2-nicotinic receptors knockout mice: a BOLD fMRI study

Energy Technology Data Exchange (ETDEWEB)

Suarez, S.V.; Changeux, J.P.; Granon, S. [Unite de Neurobiologie Integrative du Systeme Cholinergique, URA CNRS 2182, Institut Pasteur, Departement de Neuroscience, 25 rue du Dr Roux, 75015 Paris (France); Amadon, A.; Giacomini, E.; Le Bihan, D. [Service Hospitalier Frederic Joliot, 4 place du general Leclerc, 91400 Orsay (France); Wiklund, A. [Section of Anaesthesiology and Intensive Care Medicine, Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm (Sweden)

2009-07-01

Rationale: The behavioral effects of nicotine and the role of the beta2-containing nicotinic receptors in these behaviors are well documented. However, the behaviors altered by nicotine rely on the functioning on multiple brain circuits where the high-affinity {beta}2-containing nicotinic receptors ({beta}2*nAChRs) are located. Objectives We intend to see which brain circuits are activated when nicotine is given in animals naive for nicotine and whether the {beta}2*nAChRs are needed for its activation of the blood oxygen level dependent (BOLD) signal in all brain areas. Materials and methods: We used functional magnetic resonance imaging (fMRI) to measure the brain activation evoked by nicotine (1 mg/kg delivered at a slow rate for 45 min) in anesthetized C57BL/6J mice and {beta}2 knockout (KO) mice. Results: Acute nicotine injection results in a significant increased activation in anterior frontal, motor, and somatosensory cortices and in the ventral tegmental area and the substantia nigra. Anesthetized mice receiving no nicotine injection exhibited a major decreased activation in all cortical and subcortical structures, likely due to prolonged anesthesia. At a global level, {beta}2 KO mice were not rescued from the globally declining BOLD signal. However, nicotine still activated regions of a meso-cortico-limbic circuit likely via {alpha}7 nicotinic receptors. Conclusions: Acute nicotine exposure compensates for the drop in brain activation due to anesthesia through the meso-cortico-limbic network via the action of nicotine on {beta}2*nAChRs. The developed fMRI method is suitable for comparing responses in wild-type and mutant mice. (authors)

Brain activation by short-term nicotine exposure in anesthetized wild-type and beta2-nicotinic receptors knockout mice: a BOLD fMRI study

International Nuclear Information System (INIS)

Suarez, S.V.; Changeux, J.P.; Granon, S.; Amadon, A.; Giacomini, E.; Le Bihan, D.; Wiklund, A.

2009-01-01

Rationale: The behavioral effects of nicotine and the role of the beta2-containing nicotinic receptors in these behaviors are well documented. However, the behaviors altered by nicotine rely on the functioning on multiple brain circuits where the high-affinity β2-containing nicotinic receptors (β2*nAChRs) are located. Objectives We intend to see which brain circuits are activated when nicotine is given in animals naive for nicotine and whether the β2*nAChRs are needed for its activation of the blood oxygen level dependent (BOLD) signal in all brain areas. Materials and methods: We used functional magnetic resonance imaging (fMRI) to measure the brain activation evoked by nicotine (1 mg/kg delivered at a slow rate for 45 min) in anesthetized C57BL/6J mice and β2 knockout (KO) mice. Results: Acute nicotine injection results in a significant increased activation in anterior frontal, motor, and somatosensory cortices and in the ventral tegmental area and the substantia nigra. Anesthetized mice receiving no nicotine injection exhibited a major decreased activation in all cortical and subcortical structures, likely due to prolonged anesthesia. At a global level, β2 KO mice were not rescued from the globally declining BOLD signal. However, nicotine still activated regions of a meso-cortico-limbic circuit likely via α7 nicotinic receptors. Conclusions: Acute nicotine exposure compensates for the drop in brain activation due to anesthesia through the meso-cortico-limbic network via the action of nicotine on β2*nAChRs. The developed fMRI method is suitable for comparing responses in wild-type and mutant mice. (authors)

Molecular characterization and phylogenetic relationship of wild type 1 poliovirus strains circulating across Pakistan and Afghanistan bordering areas during 2010-2012.

Directory of Open Access Journals (Sweden)

Shahzad Shaukat

Full Text Available Pakistan and Afghanistan share a long uncontrolled border with extensive population movement on both sides. Wild poliovirus transmission has never been interrupted in this block due to war against terrorism, poor public health infrastructure, misconceptions about polio vaccines and inadequate immunization activities. All these issues complicate the eradication operations and reinforce the complexity of wiping out poliomyelitis from this region. This study illustrates the origins and routes of cross-border wild poliovirus type 1 (WPV1 transmission during 2010-2012 between Pakistan and Afghanistan. Sequence analyses were conducted based on complete VP1 capsid protein sequences for WPV1 study strains to determine the origin of poliovirus genetic lineages and their evolutionary relationships. Phylogenetic tree was constructed from VP1 gene sequences applying Maximum Likelihood method using Kimura 2- parameter model in MEGA program v 5.0. A total of 72 (14.3% out of 502 wild-type 1 polioviruses were found circulating in border areas of both countries during 2010-2012. Molecular phylogenetic analysis classified these strains in to two sub-genotypes with four clusters and 18 lineages. Genetic data confirmed that the most of WPV1 lineages (12; 66.6% were transmitted from Pakistan to Afghanistan. However, the genetic diversity was significantly reduced during 2012 as most of the lineages were completely eliminated. In conclusion, Pakistan-Afghanistan block has emerged as a single poliovirus reservoir sharing the multiple poliovirus lineages due to uncontrolled movement of people across the borders between two countries. If it is neglected, it can jeopardize the extensive global efforts done so-far to eradicate the poliovirus infection. Our data will be helpful to devise the preventive strategies for effective control of wild poliovirus transmission in this region.

A comparative study of age-related hearing loss in wild type and insulin-like growth factor I deficient mice