bluuebunny/crossref_metadata_2025_split huggingface dataset:
A new window into the molecular physiology of membrane proteins
Authors: Landreh Michael , Robinson Carol V. 
Published: December 2014

AbstractIntegral membrane proteins comprise ∼25% of the human proteome. Yet, our understanding of their molecular physiology is still in its infancy. This can be attributed to two factors: the experimental challenges that arise from the difficult chemical nature of membrane proteins, and the unclear relationship between their activity and their native environment. New approaches are therefore required to address these challenges. Recent developments in mass spectrometry have shown that it is possible to study membrane proteins in a solvent‐free environment and provide detailed insights into complex interactions, ligand binding and folding processes. Interestingly, not only detergent micelles but also lipid bilayer nanodiscs or bicelles can serve as a means for the gentle desolvation of membrane proteins in the gas phase. In this manner, as well as by direct addition of lipids, it is possible to study the effects of different membrane components on the structure and function of the protein components allowing us to add functional data to the least accessible part of the proteome.
Essential amino acid transporter Lat4 (Slc43a2) is required for mouse development
Authors: Guetg Adriano , Mariotta Luca , Bock Lukas , Herzog Brigitte , Fingerhut Ralph , Camargo Simone M. R. , Verrey François 
Published: December 2014

Key points
Lat4 (Slc43a2) transports branched‐chain amino acids, phenylalanine and methionine, and is expressed in kidney tubule and small intestine epithelial cells.
Using a new knockout model as a negative control, it is shown that Lat4 is expressed at the basolateral side of small intestine enterocytes and kidney epithelial cells of the proximal tubule, thick ascending limb and distal convoluted tubule.
In the Xenopus oocyte expression system, Lat4 is shown to function as a uniporter with symmetric intracellular and extracellular apparent affinities for phenylalanine.
Mice lacking Lat4 display a slight intrauterine growth restriction, postnatal malnutrition and early death, presumably as a result of defective amino acid (re)absorption.
These results demonstrate the crucial role that the uniporter Lat4 plays for amino acid transport across cellular barriers and mouse development.
AbstractAmino acid (AA) uniporter Lat4 (Slc43a2) mediates facilitated diffusion of branched‐chain AAs, methionine and phenylalanine, although its physiological role and subcellular localization are not known. We report that Slc43a2 knockout mice were born at expected Mendelian frequency but displayed an ∼10% intrauterine growth retardation and low amniotic fluid AAs, suggesting defective transplacental transport. Postnatal growth was strongly reduced, with premature death occurring within 9 days such that further investigations were made within 3 days of birth. Lat4 immunofluorescence showed a strong basolateral signal in the small intestine, kidney proximal tubule and thick ascending limb epithelial cells of wild‐type but not Slc43a2 null littermates and no signal in liver and skeletal muscle. Experiments using Xenopus laevis oocytes demonstrated that Lat4 functioned as a symmetrical low affinity uniporter with a K0.5 of ∼5 mm for both in‐ and efflux. Plasma AA concentration was decreased in Slc43a2 null pups, in particular that of non‐essential AAs alanine, serine, histidine and proline. Together with an increased level of plasma long chain acylcarnitines and a strong alteration of liver gene expression, this indicates malnutrition. Attempts to rescue pups by decreasing the litter size or by nutrients injected i.p. did not succeed. Radioactively labelled leucine but not lysine given per os accumulated in the small intestine of Slc43a2null pups, suggesting the defective transcellular transport of Lat4 substrates. In summary, Lat4 is a symmetrical uniporter for neutral essential AAs localizing at the basolateral side of (re)absorbing epithelia and is necessary for early nutrition and development.
Proton transfer unlocks inactivation in cyclic nucleotide‐gated A1 channels
Authors: Marchesi Arin , Arcangeletti Manuel , Mazzolini Monica , Torre Vincent 
Published: December 2014

Key points
Desensitization and inactivation provide a form of short‐term memory controlling the firing patterns of excitable cells and adaptation in sensory systems.
Unlike many of their cousin K+ channels, cyclic nucleotide‐gated (CNG) channels are thought not to desensitize or inactivate.
Here we report that CNG channels do inactivate and that inactivation is controlled by extracellular protons.
Titration of a glutamate residue within the selectivity filter destabilizes the pore architecture, which collapses towards a non‐conductive, inactivated state in a process reminiscent of the usual C‐type inactivation observed in many K+ channels.
These results indicate that inactivation in CNG channels represents a regulatory mechanism that has been neglected thus far, with possible implications in several physiological processes ranging from signal transduction to growth cone navigation.
AbstractIon channels control ionic fluxes across biological membranes by residing in any of three functionally distinct states: deactivated (closed), activated (open) or inactivated (closed). Unlike many of their cousin K+ channels, cyclic nucleotide‐gated (CNG) channels do not desensitize or inactivate. Using patch recording techniques, we show that when extracellular pH (pHo) is decreased from 7.4 to 6 or lower, wild‐type CNGA1 channels inactivate in a voltage‐dependent manner. pHo titration experiments show that at pHo < 7 the I–V relationships are outwardly rectifying and that inactivation is coupled to current rectification. Single‐channel recordings indicate that a fast mechanism of proton blockage underlines current rectification while inactivation arises from conformational changes downstream from protonation. Furthermore, mutagenesis and ionic substitution experiments highlight the role of the selectivity filter in current decline, suggesting analogies with the C‐type inactivation observed in K+ channels. Analysis with Markovian models indicates that the non‐independent binding of two protons within the transmembrane electrical field explains both the voltage‐dependent blockage and the inactivation. Low pH, by inhibiting the CNGA1 channels in a state‐dependent manner, may represent an unrecognized endogenous signal regulating CNG physiological functions in diverse tissues.
Development of a Reverse Genetic System for Infectious Salmon Anemia Virus: Rescue of Recombinant Fluorescent Virus by Using Salmon Internal Transcribed Spacer Region 1 as a Novel Promoter
Authors: Toro-Ascuy Daniela , Tambley Carolina , Beltran Carolina , Mascayano Carolina , Sandoval Nicolas , Olivares Eduardo , Medina Rafael A. , Spencer Eugenio , Cortez-San Martín Marcelo 
Published: December 2014

ABSTRACT

Infectious salmon anemia (ISA) is a serious disease of marine-farmed Atlantic salmon (
Salmo salar
) caused by ISA virus (ISAV), belonging to the genus
Isavirus
, family
Orthomyxoviridae
. There is an urgent need to understand the virulence factors and pathogenic mechanisms of ISAV and to develop new vaccine approaches. Using a recombinant molecular biology approach, we report the development of a plasmid-based reverse genetic system for ISAV, which includes the use of a novel fish promoter, the Atlantic salmon internal transcribed spacer region 1 (ITS-1). Salmon cells cotransfected with pSS-URG-based vectors expressing the eight viral RNA segments and four cytomegalovirus (CMV)-based vectors that express the four proteins of the ISAV ribonucleoprotein complex allowed the generation of infectious recombinant ISAV (rISAV). We generated three recombinant viruses, wild-type rISAV
901_09
and rISAVr
S6-NotI-HPR
containing a NotI restriction site and rISAV
S6/EGFP-HPR
harboring the open reading frame of enhanced green fluorescent protein (EGFP), both within the highly polymorphic region (HPR) of segment 6. All rescued viruses showed replication activity and cytopathic effect in Atlantic salmon kidney-infected cells. The fluorescent recombinant viruses also showed a characteristic cytopathic effect in salmon cells, and the viruses replicated to a titer of 6.5 × 10
5
PFU/ml, similar to that of the wild-type virus. This novel reverse genetics system offers a powerful tool to study the molecular biology of ISAV and to develop a new generation of ISAV vaccines to prevent and mitigate ISAV infection, which has had a profound effect on the salmon industry.
Escherichia coli Survival in, and Release from, White-Tailed Deer Feces
Authors: Guber Andrey K. , Fry Jessica , Ives Rebecca L. , Rose Joan B. 
Published: December 2014

ABSTRACTWhite-tailed deer are an important reservoir for pathogens that can contribute a large portion of microbial pollution in fragmented agricultural and forest landscapes. The scarcity of experimental data on survival of microorganisms in and release from deer feces makes prediction of their fate and transport less reliable and development of efficient strategies for environment protection more difficult. The goal of this study was to estimate parameters for modelingEscherichia colisurvival in and release from deer (Odocoileus virginianus) feces. Our objectives were as follows: (i) to measure survival ofE. coliin deer pellets at different temperatures, (ii) to measure kinetics ofE. colirelease from deer pellets at different rainfall intensities, and (iii) to estimate parameters of models describing survival and release of microorganisms from deer feces. Laboratory experiments were conducted to studyE. colisurvival in deer pellets at three temperatures and to estimate parameters of Chick's exponential model with temperature correction based on the Arrhenius equation. Kinetics ofE. colirelease from deer pellets were measured at two rainfall intensities and used to derive the parameters of Bradford-Schijven model of bacterial release. The results showed that parameters of the survival and release models obtained forE. coliin this study substantially differed from those obtained by using other source materials, e.g., feces of domestic animals and manures. This emphasizes the necessity of comprehensive studies of survival of naturally occurring populations of microorganisms in and release from wildlife animal feces in order to achieve better predictions of microbial fate and transport in fragmented agricultural and forest landscapes.
Resistance of Trichoplusia ni Populations Selected by Bacillus thuringiensis Sprays to Cotton Plants Expressing Pyramided Bacillus thuringiensis Toxins Cry1Ac and Cry2Ab
Authors: Kain Wendy , Song Xiaozhao , Janmaat Alida F. , Zhao Jian-Zhou , Myers Judith , Shelton Anthony M. , Wang Ping 
Published: December 2014

ABSTRACT

Two populations of
Trichoplusia ni
that had developed resistance to
Bacillus thuringiensis
sprays (Bt sprays) in commercial greenhouse vegetable production were tested for resistance to Bt cotton (BollGard II) plants expressing pyramided Cry1Ac and Cry2Ab. The
T. ni
colonies resistant to
Bacillus thuringiensis
serovar kurstaki formulations were not only resistant to the Bt toxin Cry1Ac, as previously reported, but also had a high frequency of Cry2Ab-resistant alleles, exhibiting ca. 20% survival on BollGard II foliage. BollGard II-resistant
T. ni
strains were established by selection with BollGard II foliage to further remove Cry2Ab-sensitive alleles in the
T. ni
populations. The BollGard II-resistant strains showed incomplete resistance to BollGard II, with adjusted survival values of 0.50 to 0.78 after 7 days. The resistance to the dual-toxin cotton plants was conferred by two genetically independent resistance mechanisms: one to Cry1Ac and one to Cry2Ab. The 50% lethal concentration of Cry2Ab for the resistant strain was at least 1,467-fold that for the susceptible
T. ni
strain. The resistance to Cry2Ab in resistant
T. ni
was an autosomally inherited, incompletely recessive monogenic trait. Results from this study indicate that insect populations under selection by Bt sprays in agriculture can be resistant to multiple Bt toxins and may potentially confer resistance to multitoxin Bt crops.
Genome-Wide Identification of Genes Necessary for Biofilm Formation by Nosocomial Pathogen Stenotrophomonas maltophilia Reveals that Orphan Response Regulator FsnR Is a Critical Modulator
Authors: Kang Xiu-Min , Wang Fang-Fang , Zhang Huan , Zhang Qi , Qian Wei 
Published: December 2014

ABSTRACT

Stenotrophomonas maltophilia
is a Gram-negative bacterial pathogen of increasing concern to human health. Most clinical isolates of
S. maltophilia
efficiently form biofilms on biotic and abiotic surfaces, making this bacterium resistant to a number of antibiotic treatments and therefore difficult to eliminate. To date, very few studies have investigated the molecular and regulatory mechanisms responsible for
S. maltophilia
biofilm formation. Here we constructed a random transposon insertion mutant library of
S. maltophilia
ATCC 13637 and screened 14,028 clones. A total of 46 nonredundant genes were identified. Mutants of these genes exhibited marked changes in biofilm formation, suggesting that multiple physiological pathways, including extracellular polysaccharide production, purine synthesis, transportation, and peptide and lipid synthesis, are involved in bacterial cell aggregation. Of these genes, 20 putatively contributed to flagellar biosynthesis, indicating a critical role for cell motility in
S. maltophilia
biofilm formation. Genetic and biochemical evidence demonstrated that an orphan response regulator, FsnR, activated transcription of at least two flagellum-associated operons by directly binding to their promoters. This regulatory protein plays a fundamental role in controlling flagellar assembly, cell motility, and biofilm formation. These results provide a genetic basis to systematically study biofilm formation of
S. maltophilia
.
Zoonotic Potential of Escherichia coli Isolates from Retail Chicken Meat Products and Eggs
Authors: Mitchell Natalie M. , Johnson James R. , Johnston Brian , Curtiss Roy , Mellata Melha 
Published: December 2014

ABSTRACT

Chicken products are suspected as a source of extraintestinal pathogenic
Escherichia coli
(ExPEC), which causes diseases in humans. The zoonotic risk to humans from chicken-source
E. coli
is not fully elucidated. To clarify the zoonotic risk posed by ExPEC in chicken products and to fill existing knowledge gaps regarding ExPEC zoonosis, we evaluated the prevalence of ExPEC on shell eggs and compared virulence-associated phenotypes between ExPEC and non-ExPEC isolates from both chicken meat and eggs. The prevalence of ExPEC among egg-source isolates was low, i.e., 5/108 (4.7%). Based on combined genotypic and phenotypic screening results, multiple human and avian pathotypes were represented among the chicken-source ExPEC isolates, including avian-pathogenic
E. coli
(APEC), uropathogenic
E. coli
(UPEC), neonatal meningitis
E. coli
(NMEC), and sepsis-associated
E. coli
(SEPEC), as well as an undefined ExPEC group, which included isolates with fewer virulence factors than the APEC, UPEC, and NMEC isolates. These findings document a substantial prevalence of human-pathogenic ExPEC-associated genes and phenotypes among
E. coli
isolates from retail chicken products and identify key virulence traits that could be used for screening.
Genetic Mapping Reveals that Sinefungin Resistance in Toxoplasma gondii Is Controlled by a Putative Amino Acid Transporter Locus That Can Be Used as a Negative Selectable Marker
Authors: Behnke Michael S. , Khan Asis , Sibley L. David 
Published: December 2014

ABSTRACT

Quantitative trait locus (QTL) mapping studies have been integral in identifying and understanding virulence mechanisms in the parasite
Toxoplasma gondii
. In this study, we interrogated a different phenotype by mapping sinefungin (SNF) resistance in the genetic cross between type 2 ME49-FUDR
r
and type 10 VAND-SNF
r
. The genetic map of this cross was generated by whole-genome sequencing of the progeny and subsequent identification of single nucleotide polymorphisms (SNPs) inherited from the parents. Based on this high-density genetic map, we were able to pinpoint the sinefungin resistance phenotype to one significant locus on chromosome IX. Within this locus, a single nonsynonymous SNP (nsSNP) resulting in an early stop codon in the TGVAND_290860 gene was identified, occurring only in the sinefungin-resistant progeny. Using CRISPR/CAS9, we were able to confirm that targeted disruption of TGVAND_290860 renders parasites sinefungin resistant. Because disruption of the
SNR1
gene confers resistance, we also show that it can be used as a negative selectable marker to insert either a positive drug selection cassette or a heterologous reporter. These data demonstrate the power of combining classical genetic mapping, whole-genome sequencing, and CRISPR-mediated gene disruption for combined forward and reverse genetic strategies in
T. gondii
.
Physical and neurodevelopmental outcomes in children with single-ventricle circulation
Authors: Davidson Joseph , Gringras Paul , Fairhurst Charlie , Simpson John 
Published: December 2014

ObjectiveTo investigate longer-term physical and neurodevelopmental outcomes of patients with hypoplastic left heart syndrome (HLHS) compared with other patients with functionally single-ventricle circulation surviving beyond the age of 10 years.DesignA retrospective, observational study from a UK tertiary centre for paediatric cardiology.Results58 patients with HLHS and 44 non-HLHS patients with single-ventricle physiology were included. Subjective reduction in exercise tolerance was reported in 72% (95% CI 61% to 84%) of patients with HLHS and 45% (31% to 60%) non-HLHS patients. Compared with non-HLHS patients, educational concerns were reported more frequently in patients with HLHS, 41% (29% to 54%) vs 23% (10% to 35%), as was a diagnosis of a behaviour disorder (autism or attention deficit hyperactivity disorder) 12% (4% to 21%) vs 0%, and referral to other specialist services 67% (55% to 79%) vs 48% (33% to 63%).ConclusionsWithin a group of young people with complex congenital heart disease, those with HLHS are likely to have worse physical, psychological and educational outcomes.
Survival of patients with alcohol use disorders discharged from an emergency department: a population-based cohort study
Authors: Gunnarsdottir Anna Steinunn , Kristbjornsdottir Adalbjorg , Gudmundsdottir Ragnhildur , Gunnarsdottir Oddny Sigurborg , Rafnsson Vilhjalmur 
Published: December 2014

ObjectivesThe aim was to study the cause-specific mortality of users of the emergency department (ED) who received a diagnosis of alcohol use disorder (AUD) in comparison with mortality of other users of the department.DesignA population-based prospective cohort study.ParticipantsAll patients aged 18 years and above who were subsequently discharged home from the ED during the years 2002–2008. A total of 107 237 patients were followed by record linkage to a nationwide cause-of-death registry: 1210 patients with AUD as the main discharge diagnosis and 106 027 patients in the comparison group. HR and 95% CIs were calculated.SettingED at Landspitali—the National University Hospital of Iceland, Reykjavik, Iceland. The hospital offers tertiary care and is the number one trauma centre and community hospital for the greater Reykjavik area. According to the population registry, 78% of the inhabitants of the area attended the ED during the study period.Results72 patients died in the AUD group and 4807 in the comparison group. The adjusted HR for all causes of death was 1.91 (95% CI 1.51 to 2.42). The HR for AUDs was 47.68 (95% CI 11.56 to 196.59) while for alcohol liver disease the HR was 19.06 (95% CI 6.07 to 59.87). The HR was also elevated for diseases of the circulatory system: HR 2.52 (95% CI 1.73 to 3.68); accidental poisoning: HR=13.64, (95% CI 3.98 to 46.73); suicide: HR=2.72 (95% CI 1.08 to 6.83); and event of undetermined intent: HR=10.89 (95% CI 4.53 to 26.16).ConclusionsAUD as the discharge diagnosis at the ED, among patients who were not admitted to a hospital ward but discharged home, predicts increased mortality. As the results conclusively show the vulnerability of these patients, one can question whether their needs are adequately met at the ED.
A multistudy analysis investigating systematic differences in cardiovascular trial results between Europe and Asia
Authors: Hartley Louise C , Girling Alan J , Bowater Russell J , Lilford Richard J 
Published: December 2014

ObjectiveTo assess whether there are differences in the results of cardiovascular trials between Europe and Asia using data from an extensive collection of randomised controlled trials.Study design and settingAll meta-analyses containing randomised controlled trials (RCT's) for the treatment or prevention of cardiovascular diseases were searched for in The Cochrane Library (2000–2008) and MEDLINE (2005–2008). Analysis was then conducted within and over each meta-analysis which satisfied given criteria. Separate estimates of treatment effect were calculated for Europe and Asia in each meta-analysis and then compared. Estimates of a common inter-continental difference over all meta-analyses were also calculated and meta-regression was performed. This was performed for both fatal and non-fatal end points.ResultsThe literature search identified 59 meta-analyses that satisfied the inclusion criteria. After exclusion, the number of meta-analyses reporting greater effect sizes in Asia than in Europe was significantly more than would be expected by chance (fatal 12/14, p=0.013; non-fatal 23/32, p=0.020).ConclusionsThis study provides some evidence that for cardiovascular interventions treatment effect estimation differs between Europe and Asia, with respect to both fatal and non-fatal end points.
Cone-beam CT angiography (Dyna CT) for intraoperative localization of cerebral arteriovenous malformations
Authors: Srinivasan Visish M , Schafer Sebastian , Ghali Michael G Z , Arthur Adam , Duckworth Edward A M 
Published: December 2014

BackgroundArteriovenous malformations (AVMs) of the brain are commonly treated in multimodality fashion, with endovascular embolization followed by surgical extirpation being one of the most effective strategies. Modern endovascular suites enable rotational angiography, also known as cone-beam CT angiography (CBCT-A), using the full capability of modern C-arm digital angiography systems. This imaging modality offers a superior image quality to current options such as digital subtraction angiography, MRI, or CT angiography. Preoperative planning can be greatly aided by the resolution of angioarchitecture seen in CBCT-A images. Furthermore, these images can be used for intraoperative neuronavigation when integrated with widely used frameless stereotactic systems. The utility and outcome of the use of CBCT-A for preoperative planning and intraoperative localization of AVMs was evaluated.MethodsA retrospective review was performed of 16 patients in which CBCT-A was performed, including radiological review and all clinical data.ResultsCBCT-A was successfully employed in all cases including those with (n=9) and without (n=7) rupture. Complete resection confirmed by postoperative angiography was achieved in all cases.ConclusionsWe present a novel application of CBCT-A in the treatment of AVMs, both for preoperative surgical planning and an intraoperative reference during neuronavigation.