bluuebunny/crossref_metadata_2025_split huggingface dataset:
β1 subunit stabilises sodium channel Nav1.7 against mechanical stress
Authors: Körner Jannis , Meents Jannis , Machtens Jan‐Philipp , Lampert Angelika 
Published: April 2018

Key points
The voltage‐gated sodium channel Nav1.7 is a key player in neuronal excitability and pain signalling. In addition to voltage sensing, the channel is also modulated by mechanical stress.
Using whole‐cell patch‐clamp experiments, we discovered that the sodium channel subunit β1 is able to prevent the impact of mechanical stress on Nav1.7.
An intramolecular disulfide bond of β1 was identified to be essential for stabilisation of inactivation, but not activation, against mechanical stress using molecular dynamics simulations, homology modelling and site‐directed mutagenesis.
Our results highlight the role of segment 6 of domain IV in fast inactivation.
We present a candidate mechanism for sodium channel stabilisation against mechanical stress, ensuring reliable channel functionality in living systems.
AbstractVoltage‐gated sodium channels are key players in neuronal excitability and pain signalling. Precise gating of these channels is crucial as even small functional alterations can lead to pathological phenotypes such as pain or heart failure. Mechanical stress has been shown to affect sodium channel activation and inactivation. This suggests that stabilising components are necessary to ensure precise channel gating in living organisms. Here, we show that mechanical shear stress affects voltage dependence of activation and fast inactivation of the Nav1.7 channel. Co‐expression of the β1 subunit, however, protects both gating modes of Nav1.7 against mechanical shear stress. Using molecular dynamics simulation, homology modelling and site‐directed mutagenesis, we identify an intramolecular disulfide bond of β1 (Cys21‐Cys43) which is partially involved in this process: the β1‐C43A mutant prevents mechanical modulation of voltage dependence of activation, but not of fast inactivation. Our data emphasise the unique role of segment 6 of domain IV for sodium channel fast inactivation and confirm previous reports that the intracellular process of fast inactivation can be modified by interfering with the extracellular end of segment 6 of domain IV. Thus, our data suggest that physiological gating of Nav1.7 may be protected against mechanical stress in a living organism by assembly with the β1 subunit.
Genetic Diversity among Staphylococcus aureus Isolates Showing Oxacillin and/or Cefoxitin Resistance Not Linked to the Presence of
mec
Genes
Authors: Argudín M. Angeles , Roisin S. , Nienhaus L. , Dodémont M. , de Mendonça R. , Nonhoff C. , Deplano A. , Denis O. 
Published: April 2018

ABSTRACT

Methicillin-resistant
Staphylococcus aureus
isolates lacking
mec
genes (
n
= 32), collected from Belgian hospitals, were characterized for their β-lactamase production and the presence of mutations in
pbp
genes, the
pbp4
promoter, and genes involved in penicillin-binding protein 4 overproduction (
gdpP
and
yjbH
). Twelve isolates were β-lactamase hyperproducers (BHPs), while 12 non-BHP isolates might produce an incomplete GdpP protein. Most isolates showed nucleotide missense mutations in
pbp
genes. A few isolates also showed mutations in the
pbp4
promoter.
Incomplete Downregulation of CD4 Expression Affects HIV-1 Env Conformation and Antibody-Dependent Cellular Cytotoxicity Responses
Authors: Prévost Jérémie , Richard Jonathan , Medjahed Halima , Alexander Audrey , Jones Jennifer , Kappes John C. , Ochsenbauer Christina , Finzi Andrés 
Published: April 2018

ABSTRACTHIV-1-infected cells expressing envelope glycoproteins (Env) in the CD4-bound conformation on their surfaces are targeted by antibody-dependent cellular cytotoxicity (ADCC) mediated by CD4-induced (CD4i) antibodies and sera from HIV-1-infected individuals (HIV+sera). By downregulating the surface expression of CD4, Nef prevents Env-CD4 interaction, thus protecting HIV-1-infected cells from ADCC. HIV-1 infectious molecular clones (IMCs) are widely used to measure ADCC. In order to facilitate the identification of infected cells and high-throughput ADCC analysis, reporter genes (e.g., theRenillaluciferase [LucR] gene) are often introduced into IMC constructs. We evaluated the susceptibility of HIV-1-infected CD4+T lymphocytes to ADCC using a panel of parental IMCs and derivatives that expressed the LucR reporter gene, utilizing different molecular strategies, including one specifically designed to retain Nef expression. We found that in some of these constructs, Nef expression in CD4+T cells was suboptimal, and consequently, CD4 downregulation was incomplete. CD4 molecules remaining on the cell surface resulted in the exposure of ADCC-mediating CD4i epitopes on Env and a dramatic increase in the susceptibility of the infected cells to ADCC. Strikingly, protection from ADCC was observed when cells were infected with the parental IMC, which exhibited strong CD4 downregulation. This discrepancy between the parental and Nef-impaired viruses was independent of the strains of Env expressed, but rather, it was correlated with the levels of CD4 surface expression. Overall, our results indicate that caution should be taken when selecting IMCs for ADCC measurements and that CD4 downregulation needs to be carefully monitored when drawing conclusions about the nature and magnitude of ADCC.IMPORTANCEIn-depth understanding of the susceptibility of HIV-1-infected cells to ADCC might help establish correlates of vaccine protection and guide the development of HIV-1 vaccine strategies. Different ADCC assays have been developed, including those using infectious molecular clones (IMCs) carrying a LucR reporter gene that greatly facilitates large-scale quantitative analysis. We previously reported different molecular strategies for introducing LucR while maintaining Nef expression and function and, consequently, CD4 surface downregulation. Here, we demonstrate that utilizing IMCs that exhibit impaired Nef expression can have undesirable consequences due to incomplete CD4 downregulation. CD4 molecules remaining on the cell surface resulted in the exposure of ADCC-mediating CD4i epitopes on Env and a dramatic increase in the susceptibility of the infected cells to ADCC. Overall, our results indicate that CD4 downregulation needs to be carefully monitored when drawing conclusions about the nature and magnitude of ADCC.
Listeria monocytogenes Source Distribution Analysis Indicates Regional Heterogeneity and Ecological Niche Preference among Serotype 4b Clones
Authors: Lee Sangmi , Chen Yi , Gorski Lisa , Ward Todd J. , Osborne Jason , Kathariou Sophia 
Published: April 2018

ABSTRACT

Biodiversity analysis of the foodborne pathogen
Listeria monocytogenes
recently revealed four serotype 4b major hypervirulent clonal complexes (CCs), i.e., CC1, CC2, CC4, and CC6. Hypervirulence was indicated by overrepresentation of these clones, and serotype 4b as a whole, among human clinical isolates in comparison to food. However, data on potential source-dependent partitioning among serotype 4b clones in diverse regions are sparse. We analyzed a panel of 347 serotype 4b isolates, primarily from North America, to determine the distribution of clones in humans, other animals, food, and water. CC1, CC2, CC4, and CC6 predominated, but surprisingly, only three clones, i.e., CC2 and the singleton sequence types (STs) ST382 and ST639, exhibited significant source-dependent associations, with higher propensity for food (CC2) or water (ST382 and ST639) than other sources. Pairwise comparisons between human and food isolates identified CC4 as the only serotype 4b clone significantly overrepresented among human isolates. Our analysis also revealed several serotype 4b clones emerging in North America. Two such emerging clones, ST382 (implicated in several outbreaks since 2014) and ST639, were primarily encountered among human and water isolates. Findings suggest that in spite of the ubiquity of CC1, CC2, CC4, and CC6, regional heterogeneity in serotype 4b is substantially larger than previously surmised. Analysis of even large strain panels from one region may not adequately predict clones unique to, and emerging in, other areas. Serotype 4b clonal complexes may differ in ecological niche preference, suggesting the need to further elucidate reservoirs and vehicles, especially for emerging clones.

IMPORTANCE
In
Listeria monocytogenes
, serotype 4b strains are leading contributors to human disease, but intraserotype distributions among different sources and regions remain poorly elucidated. Analysis of 347 serotype 4b isolates from four different sources, mostly from North America, confirmed the overall predominance of the major clones CC1, CC2, CC4, and CC6 but found that only CC4 was significantly associated with human disease, while CC2 was significantly associated with food. Remarkably, several emerging clones were identified among human isolates from North America, with some of these also exhibiting a propensity for surface water. The latter included the singleton clones ST382, implicated in several outbreaks in the United States since 2014, and ST639. These clones were noticeably underrepresented among much larger panels from other regions. Though associated with North America for the time being, they may eventually become globally disseminated through the food trade or other venues.
Determining the Specificity of Cascade Binding, Interference, and Primed Adaptation
In Vivo
in the
Escherichia coli
Type I-E CRISPR-Cas System
Authors: Cooper Lauren A. , Stringer Anne M. , Wade Joseph T. 
Published: April 2018

ABSTRACT

In clustered regularly interspaced short palindromic repeat (CRISPR)-Cas (CRISPR-associated) immunity systems, short CRISPR RNAs (crRNAs) are bound by Cas proteins, and these complexes target invading nucleic acid molecules for degradation in a process known as interference. In type I CRISPR-Cas systems, the Cas protein complex that binds DNA is known as Cascade. Association of Cascade with target DNA can also lead to acquisition of new immunity elements in a process known as primed adaptation. Here, we assess the specificity determinants for Cascade-DNA interaction, interference, and primed adaptation
in vivo
, for the type I-E system of
Escherichia coli
. Remarkably, as few as 5 bp of crRNA-DNA are sufficient for association of Cascade with a DNA target. Consequently, a single crRNA promotes Cascade association with numerous off-target sites, and the endogenous
E. coli
crRNAs direct Cascade binding to >100 chromosomal sites. In contrast to the low specificity of Cascade-DNA interactions, >18 bp are required for both interference and primed adaptation. Hence, Cascade binding to suboptimal, off-target sites is inert. Our data support a model in which the initial Cascade association with DNA targets requires only limited sequence complementarity at the crRNA 5′ end whereas recruitment and/or activation of the Cas3 nuclease, a prerequisite for interference and primed adaptation, requires extensive base pairing.

IMPORTANCE
Many bacterial and archaeal species encode CRISPR-Cas immunity systems that protect against invasion by foreign DNA. In the
Escherichia coli
CRISPR-Cas system, a protein complex, Cascade, binds 61-nucleotide (nt) CRISPR RNAs (crRNAs). The Cascade complex is directed to invading DNA molecules through base pairing between the crRNA and target DNA. This leads to recruitment of the Cas3 nuclease, which destroys the invading DNA molecule and promotes acquisition of new immunity elements. We made the first
in vivo
measurements of Cascade binding to DNA targets. Thus, we show that Cascade binding to DNA is highly promiscuous; endogenous
E. coli
crRNAs can direct Cascade binding to >100 chromosomal locations. In contrast, we show that targeted degradation and acquisition of new immunity elements require highly specific association of Cascade with DNA, limiting CRISPR-Cas function to the appropriate targets.
Gender and survival from out-of-hospital cardiac arrest: a New Zealand registry study
Authors: Dicker Bridget , Conaglen Kate , Howie Graham 
Published: April 2018

ObjectiveTo determine the relationships between survival from all-cause out-of-hospital cardiac arrest (OHCA) and gender in New Zealand.MethodsA retrospective observational study was conducted using data compliant with the Utstein guidelines from the St John New Zealand OHCA Registry for adult patients who were treated for an OHCA between 1 October 2013 and 30 September 2015. Univariate logistic regression was used to investigate factors associated with return of spontaneous circulation sustained to handover at hospital and survival to 30 days. Multivariate logistic regression models were used to investigate outcome differences in survival according to gender at 30 days postevent.ResultsWomen survived to hospital handover in 29% of cases, which was not significantly different from men (31%). When adjusted for age, location, aetiology, initial rhythm and witnessed status, there was no significant difference in 30-day survival between men (16%) and women (13%) (adjusted OR 1.22, 95% CI (0.96 to 1.55), p=0.11).ConclusionNo statistical differences were found in 30-day survival between genders when adjustments for unfavourable Utstein variables were accounted for.
CM-Score: a validated scoring system to predict CDKN2A germline mutations in melanoma families from Northern Europe
Authors: Potjer Thomas P , Helgadottir Hildur , Leenheer Mirjam , van der Stoep Nienke , Gruis Nelleke A , Höiom Veronica , Olsson Håkan , van Doorn Remco , Vasen Hans F A , van Asperen Christi J , Dekkers Olaf M , Hes Frederik J 
Published: April 2018

BackgroundSeveral factors have been reported that influence the probability of a germline CDKN2A mutation in a melanoma family. Our goal was to create a scoring system to estimate this probability, based on a set of clinical features present in the patient and his or her family.MethodsFive clinical features and their association with CDKN2A mutations were investigated in a training cohort of 1227 Dutch melanoma families (13.7% with CDKN2A mutation) using multivariate logistic regression. Predefined features included number of family members with melanoma and with multiple primary melanomas, median age at diagnosis and presence of pancreatic cancer or upper airway cancer in a family member. Based on these five features, a scoring system (CDKN2A Mutation(CM)-Score) was developed and subsequently validated in a combined Swedish and Dutch familial melanoma cohort (n=421 families; 9.0% with CDKN2A mutation).ResultsAll five features were significantly associated (p<0.05) with a CDKN2A mutation. At a CM-Score of 16 out of 49 possible points, the threshold of 10% mutation probability is approximated (9.9%; 95% CI 9.8 to 10.1). This probability further increased to >90% for families with ≥36 points. A CM-Score under 16 points was associated with a low mutation probability (≤4%). CM-Score performed well in both the training cohort (area under the curve (AUC) 0.89; 95% CI 0.86 to 0.92) and the external validation cohort (AUC 0.94; 95% CI 0.90 to 0.98).ConclusionWe developed a practical scoring system to predict CDKN2A mutation status among melanoma-prone families. We suggest that CDKN2A analysis should be recommended to families with a CM-Score of ≥16 points.
Mutations in plasmalemma vesicle-associated protein cause severe syndromic protein-losing enteropathy
Authors: Broekaert Ilse Julia , Becker Kerstin , Gottschalk Ingo , Körber Friederike , Dötsch Jörg , Thiele Holger , Altmüller Janine , Nürnberg Peter , Hünseler Christoph , Cirak Sebahattin 
Published: April 2018

BackgroundProtein-losing enteropathy (PLE) is characterised by gastrointestinal protein leakage due to loss of mucosal integrity or lymphatic abnormalities. PLE can manifest as congenital diarrhoea and should be differentiated from other congenital diarrhoeal disorders. Primary PLEs are genetically heterogeneous and the underlying genetic defects are currently emerging.ObjectivesWe report an infant with fatal PLE for whom we aimed to uncover the underlying pathogenic mutation.MethodsWe performed whole exome sequencing (WES) for the index patient. Variants were classified based on the American College of Medical Genetics and Genomics guidelines. WES results and our detailed clinical description of the patient were compared with the literature.ResultsWe discovered a novel homozygous stop mutation (c.988C>T, p.Q330*) in the Plasmalemma Vesicle-Associated Protein (PLVAP) gene in a newborn with fatal PLE, facial dysmorphism, and renal, ocular and cardiac anomalies. The Q330* mutation is predicted to result in complete loss of PLVAP protein expression leading to deletion of the diaphragms of endothelial fenestrae, resulting in plasma protein extravasation and PLE. Recently, another single homozygous stop mutation in PLVAP causing lethal PLE in an infant was reported.ConclusionsOur findings validate PLVAP mutations as a cause of syndromic PLE. Prenatal anomalies, severe PLE and syndromic features may guide the diagnosis of this rare disease.
Changing antiepilepsy drug-prescribing trends in women with epilepsy in the UK and Ireland and the impact on major congenital malformations
Authors: Kinney Michael O , Morrow James , Patterson Chris C , Campbell Ellen , Russell Aline , Smithson Henry W , Parsons Linda , Morrison Patrick J , Bromley Rebecca , Liggan Brenda , Delanty Norman , Irwin Beth , Hunt Stephen J , Craig John J 
Published: April 2018

ObjectivesAfter 20 years of data collection, pregnancy registers have informed prescribing practice. Various populations show trends for a reduction in valproate prescribing, which is associated with an increased risk of anatomical teratogenesis and neurodevelopmental effects in those exposed in utero. Our aim was to determine if any shifts in prescribing trends have occurred in the UK and Ireland Epilepsy and Pregnancy Register cohort and to assess if there had been any change in the overall major congenital malformation (MCM) rate over time.MethodsThe UK and Ireland Epilepsy and Pregnancy Register, a prospective, observational, registration and follow-up study established in 1996, was used to determine the changes in antiepileptic drugs (AEDs) utilised during pregnancy and the MCM rate between 1996 and 2016. Linear regression analysis was used to assess changes in AED utilisation, and Poisson regression was used for the analysis of trends in the MCM rates.ResultsOutcome data for 9247 pregnancies showed a stable percentage of monotherapy to polytherapy prescribing habits over time. After Bonferroni correction, statistically significant (p<0.003) changes were found in monotherapy prescribing with increases in lamotrigine and levetiracetam and decreases in valproate and carbamazepine use. Between 1996 and 2016, the total MCM rate showed a 2.1% reduction per year (incidence risk ratio 0.979 (95% CIs 0.956 to 1.002) but Poisson regression analysis showed that this was not statistically significant p=0.08).ConclusionSignificant changes are seen in the prescribing habits in this cohort over 20 years, but a statistically significant change in the MCM rate was not detected. This work should be replicated on a larger scale to determine if significant changes are occurring in the MCM rate, which would allow a robust economic estimate of the benefits of improvements in prescribing practice and the personal effect of such changes.
Sustained Adrenergic Signaling Promotes Intratumoral Innervation through BDNF Induction
Authors: Allen Julie K. , Armaiz-Pena Guillermo N. , Nagaraja Archana S. , Sadaoui Nouara C. , Ortiz Tatiana , Dood Robert , Ozcan Merve , Herder Danielle M. , Haemmerle Monika , Gharpure Kshipra M. , Rupaimoole Rajesha , Previs Rebecca A. , Wu Sherry Y. , Pradeep Sunila , Xu Xiaoyun , Han Hee Dong , Zand Behrouz , Dalton Heather J. , Taylor Morgan , Hu Wei , Bottsford-Miller Justin , Moreno-Smith Myrthala , Kang Yu , Mangala Lingegowda S. , Rodriguez-Aguayo Cristian , Sehgal Vasudha , Spaeth Erika L. , Ram Prahlad T. , Wong Stephen T.C. , Marini Frank C. , Lopez-Berestein Gabriel , Cole Steve W. , Lutgendorf Susan K. , De Biasi Mariella , Sood Anil K. 
Published: April 2018

Abstract
Mounting clinical and preclinical evidence supports a key role for sustained adrenergic signaling in the tumor microenvironment as a driver of tumor growth and progression. However, the mechanisms by which adrenergic neurotransmitters are delivered to the tumor microenvironment are not well understood. Here we present evidence for a feed-forward loop whereby adrenergic signaling leads to increased tumoral innervation. In response to catecholamines, tumor cells produced brain-derived neurotrophic factor (BDNF) in an ADRB3/cAMP/Epac/JNK-dependent manner. Elevated BDNF levels in the tumor microenvironment increased innervation by signaling through host neurotrophic receptor tyrosine kinase 2 receptors. In patients with cancer, high tumor nerve counts were significantly associated with increased BDNF and norepinephrine levels and decreased overall survival. Collectively, these data describe a novel pathway for tumor innervation, with resultant biological and clinical implications.
Significance: Sustained adrenergic signaling promotes tumor growth and metastasis through BDNF-mediated tumoral innervation. Cancer Res; 78(12); 3233–42. ©2018 AACR.
Differential Effects of Estrogen Receptor β Isoforms on Glioblastoma Progression
Authors: Liu Jinyou , Sareddy Gangadhara R. , Zhou Mei , Viswanadhapalli Suryavathi , Li Xiaonan , Lai Zhao , Tekmal Rajeshwar R. , Brenner Andrew , Vadlamudi Ratna K. 
Published: April 2018

Abstract
The estrogen receptor β (ERβ) functions as a tumor suppressor in glioblastoma (GBM) cells. However, the in vivo significance of endogenous ERβ and the roles of its isoforms in GBM are incompletely understood. Using ERβ isoform-specific PCR screening, we found that GBM cells predominantly express ERβ1 and ERβ5, along with low levels of ERβ2 and ERβ4. We observed greater ERβ5 expression in higher grades of glioma than in lower grades. In CRISPR-based ERβ knockout (KO) cells and ERβ KO cells uniquely expressing ERβ1 or ERβ5 only, ERβ1 significantly reduced proliferation. Compared with parental GBM cells, ERβ KO cells exhibited high migratory and invasive potentials, and reexpression of ERβ1 resulted in the reduction of this phenotype. Interestingly, ERβ5 expression increased foci formation and anchorage-independent growth of NIH3T3 cells and increased motile structure formation, including filopodia and ruffles in GBM cells. Only ERβ1-expressing tumors resulted in longer mouse survival. RNA-Seq analysis revealed unique pathways modulated by ERβ1 and ERβ5. Compared with ERβ KO cells, ERβ1 cells exhibited lower activation of mTOR signaling molecules, including p-mTOR, p-S6K, and p-S6, and ERβ5-expressing cells had enhanced mTOR downstream signaling. Unique proteins including several that function as regulators of mTOR, immunomodulatory, and apoptosis pathways bound to ERβ1 and ERβ5 isoforms. Our work confirms the tumor-suppressive potential of ERβ1 and reveals the acquired oncogenic ability of ERβ5 in GBM cells. ERβ isoform status and their unique interactions with oncogenic pathways may have important implications in GBM progression.
Significance: These findings suggest that only ERβ isoform 1 has tumor suppressor function in GBM and that ERβ isoform switching contributes to GBM progression. Cancer Res; 78(12); 3176–89. ©2018 AACR.