bluuebunny/crossref_metadata_2025 huggingface dataset:
The Contribution of Germline Predisposition Gene Mutations to Clinical Subtypes of Invasive Breast Cancer From a Clinical Genetic Testing Cohort
Authors: Hu Chunling , Polley Eric C , Yadav Siddhartha , Lilyquist Jenna , Shimelis Hermela , Na Jie , Hart Steven N , Goldgar David E , Shah Swati , Pesaran Tina , Dolinsky Jill S , LaDuca Holly , Couch Fergus J
Published: February 2020

Background

The germline cancer predisposition genes associated with increased risk of each clinical subtype of breast cancer, defined by estrogen receptor (ER), progesterone receptor (PR), and HER2, are not well defined.

Methods

A total of 54 555 invasive breast cancer patients with 56 480 breast tumors were subjected to clinical hereditary cancer multigene panel testing. Heterogeneity for predisposition genes across clinical breast cancer subtypes was assessed by comparing mutation frequencies by gene among tumor subtypes and by association studies between each tumor subtype and reference controls.

Results

Mutations in 15 cancer predisposition genes were detected in 8.6% of patients with ER+/HER2-; 8.9% with ER+/HER2+; 7.7% with ER-/HER2+; and 14.4% of ER-/PR-/HER2- tumors. BRCA1, BRCA2, BARD1, and PALB2 mutations were enriched in ER- and HER2- tumors; RAD51C and RAD51D mutations were enriched in ER- tumors only; TP53 mutations were enriched in HER2+ tumors, and ATM and CHEK2 mutations were enriched in both ER+ and/or HER2+ tumors. All genes were associated with moderate (odds ratio > 2.00) or strong (odds ratio > 5.00) risks of at least one subtype of breast cancer in case-control analyses. Mutations in ATM, BARD1, BRCA1, BRCA2, CHEK2, PALB2, RAD51C, RAD51D, and TP53 had predicted lifetime absolute risks of at least 20.0% for breast cancer.

Conclusions

Germline mutations in hereditary cancer panel genes confer subtype-specific risks of breast cancer. Combined tumor subtype, age at breast cancer diagnosis, and family history of breast and/or ovarian cancer information provides refined categorical estimates of mutation prevalence for women considering genetic testing.
Novel routes towards bioplastics from plants: elucidation of the methylperillate biosynthesis pathway from Salvia dorisiana trichomes
Authors: Jongedijk Esmer , Müller Sebastian , van Dijk Aalt D J , Schijlen Elio , Champagne Antoine , Boutry Marc , Levisson Mark , van der Krol Sander , Bouwmeester Harro , Beekwilder Jules
Published: February 2020

Plants produce a large variety of highly functionalized terpenoids. Functional groups such as partially unsaturated rings and carboxyl groups provide handles to use these compounds as feedstock for biobased commodity chemicals. For instance, methylperillate, a monoterpenoid found in Salvia dorisiana, may be used for this purpose, as it carries both an unsaturated ring and a methylated carboxyl group. The biosynthetic pathway of methylperillate in plants is still unclear. In this work, we identified glandular trichomes from S. dorisiana as the location of biosynthesis and storage of methylperillate. mRNA from purified trichomes was used to identify four genes that can encode the pathway from geranyl diphosphate towards methylperillate. This pathway includes a (–)-limonene synthase (SdLS), a limonene 7-hydroxylase (SdL7H, CYP71A76), and a perillyl alcohol dehydrogenase (SdPOHDH). We also identified a terpene acid methyltransferase, perillic acid O-methyltransferase (SdPAOMT), with homology to salicylic acid OMTs. Transient expression in Nicotiana benthamiana of these four genes, in combination with a geranyl diphosphate synthase to boost precursor formation, resulted in production of methylperillate. This demonstrates the potential of these enzymes for metabolic engineering of a feedstock for biobased commodity chemicals.
A comparison between short GRB afterglows and kilonova AT2017gfo: shedding light on kilonovae properties
Authors: Rossi A , Stratta G , Maiorano E , Spighi D , Masetti N , Palazzi E , Gardini A , Melandri A , Nicastro L , Pian E , Branchesi M , Dadina M , Testa V , Brocato E , Benetti S , Ciolfi R , Covino S , D’Elia V , Grado A , Izzo L , Perego A , Piranomonte S , Salvaterra R , Selsing J , Tomasella L , Yang S , Vergani D , Amati L , Stephen J B
Published: February 2020

Multimessenger astronomy received a great boost following the discovery of kilonova (KN) AT2017gfo, the optical counterpart of the gravitational wave source GW170817 associated with the short gamma-ray burst GRB 170817A. AT2017gfo was the first KN that could be extensively monitored in time using both photometry and spectroscopy. Previously, only few candidates have been observed against the glare of short GRB afterglows. In this work, we aim to search the fingerprints of AT2017gfo-like KN emissions in the optical/NIR light curves of 39 short GRBs with known redshift. For the first time, our results allow us to study separately the range of luminosity of the blue and red components of AT2017gfo-like kilonovae in short GRBs. In particular, the red component is similar in luminosity to AT2017gfo, while the blue KN can be more than 10 times brighter. Finally, we exclude a KN as luminous as AT2017gfo in GRBs 050509B and 061201.
Assessing non-linear models for galaxy clustering – II. Model validation and forecasts for Stage IV surveys
Authors: Bose Benjamin , Pourtsidou Alkistis , Markovič Katarina , Beutler Florian
Published: February 2020

Accurate modelling of non-linear scales in galaxy clustering will be crucial for data analysis of Stage IV galaxy surveys. A selection of competing non-linear models must be made based on validation studies. We provide a comprehensive set of forecasts of two different models for the halo redshift space power spectrum, namely the commonly applied TNS model and an effective field theory of large-scale structure (EFTofLSS) inspired model. Using simulation data and a least-χ2 analysis, we determine ranges of validity for the models. We then conduct an exploratory Fisher analysis using the full anisotropic power spectrum to investigate parameter degeneracies. We proceed to perform an MCMC analysis utilizing the monopole, quadrupole, and hexadecapole spectra, with a restricted range of scales for the latter in order to avoid biasing our growth rate, f, constraint. We find that the TNS model with a Lorentzian damping and standard Eulerian perturbative modelling outperforms other variants of the TNS model. Our MCMC analysis finds that the EFTofLSS-based model may provide tighter marginalized constraints on f at z = 0.5 and z = 1 than the TNS model, despite having additional nuisance parameters. However this depends on the range of scales used as well as the fiducial values and priors on the EFT nuisance parameters. Finally, we extend previous work to provide a consistent comparison between the Fisher matrix and MCMC forecasts using the multipole expansion formalism, and find good agreement between them.
Faint rapid red transients from neutron star–CO white dwarf mergers
Authors: Zenati Yossef , Bobrick Alexey , Perets Hagai B
Published: February 2020

Mergers of neutron stars (NS) and white dwarfs (WD) may give rise to observable explosive transient events. We use 3D hydrodynamical (smoothed particle hydrodynamics – SPH) simulations and 2D hydrodynamical–thermonuclear simulations (using the flash adaptive mesh refinement code) to model the disruption of CO-WDs by NSs, which produce faint transient events. We post-process the simulations using a large nuclear network and make use of the SuperNu radiation transfer code to predict the observational signatures and detailed properties of these transients. We calculate the light curves and spectra for five models of NS–CO-WD mergers. The small yields of 56Ni (few $\times 10^{-3}\, {\rm M_{\odot }}$) result in faint, rapidly evolving reddened transients (RRTs) with B(R) peak magnitudes of at most ∼−12 (−13) to ∼−13 (−15), much shorter and fainter than both regular and faint/peculiar Type Ia supernovae. These transients are likely to be accompanied by several months long, 1–2 mag dimmer red/infrared afterglows. We show that the spectra of RRTs share some similarities with rapidly evolving transients such as SN 2010X, although RRTs are significantly fainter, especially in the I/R bands, and show far stronger Si lines. We estimate that the upcoming Large Synoptic Survey Telescope could detect RRTs at a rate of up to ∼10–70 yr−1 through observations in the R/I bands. The qualitative agreement between the SPH and flash approaches supports the earlier hydrodynamical studies of these systems.
Hierarchical chromatin organization detected by TADpole
Authors: Soler-Vila Paula , Cuscó Pol , Farabella Irene , Di Stefano Marco , Marti-Renom Marc A
Published: February 2020

The rapid development of Chromosome Conformation Capture (3C-based techniques), as well as imaging together with bioinformatics analyses, has been fundamental for unveiling that chromosomes are organized into the so-called topologically associating domains or TADs. While TADs appear as nested patterns in the 3C-based interaction matrices, the vast majority of available TAD callers are based on the hypothesis that TADs are individual and unrelated chromatin structures. Here we introduce TADpole, a computational tool designed to identify and analyze the entire hierarchy of TADs in intra-chromosomal interaction matrices. TADpole combines principal component analysis and constrained hierarchical clustering to provide a set of significant hierarchical chromatin levels in a genomic region of interest. TADpole is robust to data resolution, normalization strategy and sequencing depth. Domain borders defined by TADpole are enriched in main architectural proteins (CTCF and cohesin complex subunits) and in the histone mark H3K4me3, while their domain bodies, depending on their activation-state, are enriched in either H3K36me3 or H3K27me3, highlighting that TADpole is able to distinguish functional TAD units. Additionally, we demonstrate that TADpole's hierarchical annotation, together with the new DiffT score, allows for detecting significant topological differences on Capture Hi-C maps between wild-type and genetically engineered mouse.
The critical function of the plastid rRNA methyltransferase, CMAL, in ribosome biogenesis and plant development
Authors: Zou Meijuan , Mu Ying , Chai Xin , Ouyang Min , Yu Long-Jiang , Zhang Lixin , Meurer Jörg , Chi Wei
Published: February 2020

Methylation of nucleotides in ribosomal RNAs (rRNAs) is a ubiquitous feature that occurs in all living organisms. The formation of methylated nucleotides is performed by a variety of RNA-methyltransferases. Chloroplasts of plant cells result from an endosymbiotic event and possess their own genome and ribosomes. However, enzymes responsible for rRNA methylation and the function of modified nucleotides in chloroplasts remain to be determined. Here, we identified an rRNA methyltransferase, CMAL (Chloroplast MraW-Like), in the Arabidopsis chloroplast and investigated its function. CMAL is the Arabidopsis ortholog of bacterial MraW/ RsmH proteins and accounts to the N4-methylation of C1352 in chloroplast 16S rRNA, indicating that CMAL orthologs and this methyl-modification nucleotide is conserved between bacteria and the endosymbiont-derived eukaryotic organelle. The knockout of CMAL in Arabidopsis impairs the chloroplast ribosome accumulation and accordingly reduced the efficiency of mRNA translation. Interestingly, the loss of CMAL leads not only to defects in chloroplast function, but also to abnormal leaf and root development and overall plant morphology. Further investigation showed that CMAL is involved in the plant development probably by modulating auxin derived signaling pathways. This study uncovered the important role of 16S rRNA methylation mediated by CMAL in chloroplast ribosome biogenesis and plant development.
Vote Choice and the Nonseparability of Economic and Social Issues
Authors: Stoetzer Lukas F , Zittlau Steffen
Published: February 2020

How voters use political issues to elect political candidates is of central importance to our understanding of democratic representation. Research on voting behavior often assumes that American voters hold distinct economic and cultural issue preferences. In this research note, we point out that this does not necessarily imply that preferences for candidates’ positions on the two issue dimensions are also additively separable in voters’ decisions. Analyzing survey data on US presidential elections from 1996 to 2016, we estimate to what extent voters’ economic and socio-cultural preferences are nonseparable and find that the two general dimensions act as substitutes in their decisions. Our finding implies that voting decisions are partially structured by an underlying single dimension, as liberal deviations from a voter’s ideal point on social issues can be compensated by conservative deviations on economic issues.
Charged spherically symmetric Taub–NUT black hole solutions in $f(R)$ gravity
Authors: Nashed G G L , Bamba Kazuharu
Published: February 2020

$f(R)$ theory is a modification of Einstein’s general relativity which has provided many interesting results in cosmology and astrophysics. To derive a black hole solution in this theory is difficult due to the fact that it contains fourth-order differential equations. In this study, we use the first reliable deviation from general relativity which is given by the quadratic form of $f(R)=R+\beta R^2$, where $\beta$ is a dimensional parameter. We calculate the energy conditions of charged black holes and show that they are all satisfied for the Taub–NUT spacetime. Finally, we study some thermodynamic quantities such as entropy, temperature, specific heat, and Gibbs free energy. The calculations of heat capacity and free energy show that the charged Taub–NUT black hole has positive values, which means that it has thermal stability.
Short-term effects of air pollution on daily asthma-related emergency department visits in an industrial city
Authors: AlBalawi Salem M , Namdeo Anil , Hodgson Susan , Pless-Mulloli Tanja , McNally Richard J Q
Published: February 2020

Background

Epidemiological studies from Europe and North America have provided evidence that exposure to air pollution can aggravate symptoms in asthmatic patients.

Methods

Daily number of AEDv, air pollution levels (PM10, PM2.5, SO2, NO2 and CO) and meteorological variables was obtained from Jubail Industrial City, Saudi Arabia, for the period of 2007–11. Data were analyzed using a time-series approach. Relative risks (RRs) were estimated using Poisson regression.

Results

The associations between AEDv and PM10, PM2.5, SO2 and NO2 remained positive and statistically significant after mutual adjustment in the multi-pollutant model.

The RR of AEDv increased by 5.4, 4.4, 3.4 and 2.2% per an inter-quartile range increase in SO2 (2.0 ppb), PM2.5 (36 μg/m3), NO2 (7.6 ppb) and PM10 (140 μg/m3), respectively. No significant associations between AEDv and CO were found.

Conclusions

Current levels of ambient air pollution are associated with AEDv in this industrial setting in the Middle East. Greater awareness of environmental health protection and the implementation of effective measures to improve the quality of air in such settings would be beneficial to public health.
Molecular epidemiology, in vitro susceptibility and exoenzyme screening of Malassezia clinical isolates
Authors: Li Wei , Zhang Zi-Wei , Luo Yun , Liang Ni , Pi Xiao-Xue , Fan Yi-Ming
Published: February 2020

Introduction.Malasseziafolliculitis (MF) and pityriasis versicolor (PV) are common dermatoses caused byMalasseziaspecies. Their molecular epidemiology, drug susceptibility and exoenzymes are rarely reported in China.

Aim.To investigate the molecular epidemiology, drug susceptibility and enzymatic profile ofMalasseziaclinical isolates.

Methodology.Malasseziastrains were recovered from MF and PV patients and healthy subjects (HS) and identified by sequencing analysis. The minimum inhibitory concentrations (MICs) of nine antifungals (posaconazole, voriconazole, itraconazole, fluconazole, ketoconazole, miconazole, bifonazole, terbinafine and caspofungin) and tacrolimus, the interactions between three antifungals (itraconazole, ketoconazole and terbinafine) and tacrolimus, and the extracellular enzyme profile were evaluated using broth and checkerboard microdilution and the Api-Zym system, respectively.

Results.Among 392Malasseziaisolates from 729 subjects (289 MF, 218 PV and 222 HS),Malassezia furfurandMalassezia globosaaccounted for 67.86 and 18.88 %, respectively.M. furfurwas the major species in MF and PV patients and HS. Among 60M.furfurand 50M.globosastrains, the MICs for itraconazole, posaconazole, voriconazole and ketoconazole were <1 μg ml−1.M. furfurwas more susceptible to itraconazole, terbinafine and bifonazole but tolerant to miconazole compared withM. globosa(P<0.05). Synergistic effects between terbinafine and itraconazole or between tacrolimus and itraconazole, ketoconazole or terbinafine occurred in 6, 7, 6 and 9 out of 37 strains, respectively. Phosphatases, lipases and proteases were mainly secreted in 51 isolates.

Conclusions.Itraconazole, posaconazole, voriconazole and ketoconazole are theagents against which there is greatest susceptibility. Synergistic effects between terbinafine and itraconazole or tacrolimas and antifungals may be irrelevant to clinical application. Overproduction of lipases could enhance the skin inhabitation ofM. furfur.
Can Search Query Forecast successfully in China’s novel coronavirus (2019-nCov) pneumonia?
Authors: Xiaoxuan LI , Qi WU , Bbenfu Lv
Published: February 2020

Recently the novel coronavirus (2019-nCov) pneumonia outbreak in China then the world, and the Number of infections and death continues to increases. Search Query performs well in forecasting the epidemics. It is still a question whether search engine data can forecast the drift and the inflexion in 2019-nCov pneumonia. Based on the Baidu Search Index, we propose three prediction models: composite Index, composite Index with filtering and suspected NCP(Novel Coronavirus Pneumonia). The result demonstrates that the predictive model of composite index with filtering performs the best while the model of suspected NCP has the highest forecast error. We further predict the out-of-the-set NCP confirmed cases and monitor that the next peak of new diagnoses will occur on February 16thand 17th.
Estimating the distribution of the incubation period of 2019 novel coronavirus (COVID-19) infection between travelers to Hubei, China and non-travelers
Authors: Leung Char
Published: February 2020

Objectives

Amid the continuing spread of the novel coronavirus (COVID-19), the incubation period of COVID-19 should be regularly re-assessed as more information is available upon the increase in reported cases. The present work estimated the distribution of incubation periods of patients infected in and outside Hubei province of China.

Methods

Clinical data were collected from the individual cases reported by the media as they were not fully available on the official pages of the Chinese health authorities. MLE was used to estimate the distributions of the incubation period.

Results

It was found that the incubation period of patients with no travel history to Hubei was longer and more volatile.

Conclusion

It is recommended that the duration of quarantine should be extended to at least 3 weeks.
A simple laboratory parameter facilitates early identification of COVID-19 patients
Authors: Li Qilin , Ding Xiuli , Xia Geqing , Geng Zhi , Chen Fenghua , Wang Lin , Wang Zheng
Published: February 2020

The total number of COVID-19 patients since the outbreak of this infection in Wuhan, China has reached 40000 and are still growing. To facilitate triage or identification of the large number of COVID-19 patients from other patients with similar symptoms in designated fever clinics, we set to identify a practical marker that could be conveniently utilized by first-line health-care workers in clinics. To do so, we performed a case-control study by analyzing clinical and laboratory findings between PCR-confirmed SARS-CoV-2 positive patients (n=52) and SARS-CoV-2 negative patients (n=53). The patients in two cohorts all had similar symptoms, mainly fever and respiratory symptoms. The rates of patients with leukocyte counts (normal or decreased number) or lymphopenia (two parameters suggested by current National and WHO COVID-19 guidelines) had no differences between these two cohorts, while the rate of eosinopenia (decreased number of eosinophils) in SARS-CoV-2 positive patients (79%) was much higher than that in SARS-CoV-2 negative patients (36%). When the symptoms were combined with eosinopenia, this combination led to a diagnosis sensitivity and specificity of 79% and 64%, respectively, much higher than 48% and 53% when symptoms were combined with leukocyte counts (normal or decreased number) and/ or lymphopenia. Thus, our analysis reveals that eosinopenia may be a potentially more reliable laboratory predictor for SARS-CoV-2 infection than leukocyte counts and lymphopenia recommended by the current guidelines.
Cohort Profile: The China Multi-Ethnic Cohort (CMEC) Study
Authors: Zhao Xing , Hong Feng , Yin Jianzhong , Tang Wenge , Zhang Gang , Liang Xian , Li Jingzhong , Cui Chaoying , Li Xiaosong
Published: February 2020

Cohort purpose

The China Multi-Ethnic Cohort (CMEC) is a community population-based prospective observational study aiming to address the urgent need for understanding NCD prevalence, risk factors and associated conditions in resource-constrained settings for ethnic minorities in China.

Cohort Basics

A total of 99 556 participants aged 30 to 79 years (Tibetan populations include those aged 18 to 30 years) from the Tibetan, Yi, Miao, Bai, Bouyei, and Dong ethnic groups in Southwest China were recruited between May 2018 and September 2019.

Follow-up and attrition

All surviving study participants will be invited for re-interviews every 3-5 years with concise questionnaires to review risk exposures and disease incidence. Furthermore, the vital status of study participants will be followed up through linkage with established electronic disease registries annually.

Design and Measures

The CMEC baseline survey collected data with an electronic questionnaire and face-to-face interviews, medical examinations and clinical laboratory tests. Furthermore, we collected biological specimens, including blood, saliva and stool, for long-term storage. In addition to the individual level data, we also collected regional level data for each investigation site.

Collaboration and data access

Collaborations are welcome. Please send specific ideas to corresponding author at: xingzhao@scu.edu.cn.
Endosidin20 targets cellulose synthase catalytic domain to inhibit cellulose biosynthesis
Authors: Huang Lei , Li Xiaohui , Zhang Weiwei , Ung Nolan , Liu Nana , Yin Xianglin , Li Yong , Mcewan Robert E. , Dilkes Brian , Dai Mingji , Hicks Glenn R. , Raikhel Natasha V. , Staiger Christopher J. , Zhang Chunhua
Published: February 2020

Cellulose is synthesized by rosette structured cellulose synthase (CESA) complexes (CSCs), each of which is composed of multiple units of CESAs in three different isoforms. CSCs rely on vesicle trafficking for delivery to the plasma membrane where they catalyze cellulose synthesis. Although the rosette structured CSCs were observed decades ago, it remains unclear what amino acids in plant CESA that directly participate in cellulose catalytic synthesis. It is also not clear how the catalytic activity of CSCs influences their efficient transport at the subcellular level. Here we report characterization of the small molecule Endosidin20 (ES20) and present evidence that it represents a new CESA inhibitor. We show data from chemical genetic analyses, biochemical assays, structural modeling, and molecular docking to support our conclusion that ES20 targets the catalytic site of Arabidopsis CESA6. Further, chemical genetic analysis reveals important amino acids that potentially form the catalytic site of plant CESA6. Using high spatiotemporal resolution live-cell imaging, we found that inhibition of CSC catalytic activity by inhibitor treatment, or by creating missense mutation at amino acids in the predicted catalytic site, causes reduced efficiency in CSC transport to the plasma membrane. Our results show that the catalytic activity of plant CSCs is integrated with subcellular trafficking dynamics.

One sentence summary

Endosidin20 targets cellulose synthase at the catalytic site to inhibit cellulose synthesis and the inhibition of catalytic activity reduces cellulose synthase complex delivery to the plasma membrane.
Coordinated assembly and release of adhesions builds apical junctional belts duringde novopolarisation of an epithelial tube
Authors: Symonds Andrew C , Buckley Clare E , Williams Charlotte A , Clarke Jonathan DW
Published: February 2020

Using the zebrafish neural tube as a model, we uncover thein vivomechanisms allowing the generation of two opposing apical epithelial surfaces within the centre of an initially unpolarised, solid organ. We show that NOK/Pals1/Mpp5a and Rab11a play a dual role in coordinating the generation of ipsilateral junctional belts whilst simultaneously releasing contralateral adhesions across the centre of the tissue. We show that Nok and Rab11a mediated resolution of cell-cell adhesions are both necessary for midline lumen opening and contribute to later maintenance of epithelial organisation. We propose these roles for both Nok and Rab11a operate through the transmembrane protein Crumbs. In light of a recent conflicting publication, we also clarify that the junction remodelling role of Nok is not specific to dividing cells.
Role of nanoscale antigen organization on B-cell activation probed using DNA origami
Authors: Veneziano Rémi , Moyer Tyson J. , Stone Matthew B. , Shepherd Tyson R. , Schief William R. , Irvine Darrell J. , Bathe Mark
Published: February 2020

Arraying vaccine immunogens in a multivalent form on the surface of virus-like particles is an important strategy used to enhance the efficacy of subunit vaccines. However, the impacts of antigen valency, spacing, and spatial organization on B cell triggering remain poorly understood. Here, we use DNA origami nanoparticles to create precise nanoscale organizations of a clinically-relevant HIV gp120 immunogen to systematically interrogate their impact on B cell triggeringin vitro. We find that antigen dimers elicit monotonically increasing B cell receptor activation as inter-antigen spacing increases up to ~30 nm, and only 5 immunogens arrayed on the surface of a 3D particle are needed to elicit maximal B cell calcium signaling. Our results reveal design principles of viral and immunogen display that drive functional B cell responses.
High antibody titres induced by protein subunit vaccines against Buruli ulcer usingMycobacterium ulceransantigens Hsp18 and MUL_3720
Authors: Mangas Kirstie M. , Tobias Nicholas , Marion Estelle , Babonneau Jérémie , Marsollier Laurent , Porter Jessica L. , Pidot Sacha J. , Wong Chinn Yi , Jackson David C. , Chua Brendon Y. , Stinear Timothy P.
Published: February 2020

Background

Mycobacterium ulceransis the causative agent of a debilitating skin and soft tissue infection known as Buruli ulcer (BU). There is no vaccine against BU. The purpose of this study was to investigate the vaccine potential of two previously described immunogenicM. ulceransproteins, MUL_3720 and Hsp18, using a mouse tail infection model of BU.

Methods

Recombinant versions of the two proteins were each electrostatically coupled with a previously described lipopeptide adjuvant. Seven C57BL/6 and seven BALB/c mice were vaccinated and boosted with each of the formulations. Vaccinated mice were then challenged withM. ulceransvia subcutaneous tail inoculation. Vaccine performance was assessed by time-to-ulceration compared to unvaccinated mice.

Results

The MUL_3720 and Hsp18 vaccines induced high titres of antigen-specific antibodies that were predominately subtype IgG1. However, all mice developed ulcers by day-40 post-M. ulceranschallenge. No significant difference was observed in the time-to-onset of ulceration between the experimental vaccine groups and unvaccinated animals.

Conclusions

These data align with previous vaccine experiments using Hsp18 and MUL_3720 that indicated these proteins may not be appropriate vaccine antigens. This work highlights the need to explore alternative vaccine targets and different approaches to understand the role antibodies might play in controlling BU.
Adaptation of pancreatic cancer cells to nutrient deprivation is reversible and requires glutamine synthetase stabilization by mTORC1
Authors: Tsai Pei-Yun , Lee Min-Sik , Jadhav Unmesh , Naqvi Insia , Madha Shariq , Adler Ashley , Mistry Meeta , Naumenko Sergey , Lewis Caroline A. , Hitchcock Daniel S. , Roberts Frederick R. , DelNero Peter , Hank Thomas , Honselmann Kim C. , Oyarvide Vicente Morales , Mino-Kenudson Mari , Clish Clary B. , Shivdasani Ramesh A. , Kalaany Nada Y.
Published: February 2020

Pancreatic ductal adenocarcinoma (PDA) is a lethal, therapy-resistant cancer that thrives in a highly desmoplastic, nutrient-deprived microenvironment. Several studies investigated the effects of depriving PDA of either glucose or glutamine alone. However, the consequences on PDA growth and metabolism of limiting both preferred nutrients have remained largely unknown. Here, we report the selection for clonal human PDA cells that survive and adapt to limiting levels of both glucose and glutamine. We find that adapted clones exhibit increased growth in vitro and enhanced tumor-forming capacity in vivo. Mechanistically, adapted clones share common transcriptional and metabolic programs, including amino acid use for de novo glutamine and nucleotide synthesis. They also display enhanced mTORC1 activity that prevents the proteasomal degradation of glutamine synthetase (GS), the rate-limiting enzyme for glutamine synthesis. This phenotype is notably reversible, with PDA cells acquiring alterations in open chromatin upon adaptation. Silencing of GS suppresses the enhanced growth of adapted cells and mitigates tumor growth. These findings identify non-genetic adaptations to nutrient deprivation in PDA and highlight GS as a dependency that could be targeted therapeutically in pancreatic cancer patients.

Significance

Pancreatic ductal adenocarcinoma (PDA) is a highly lethal malignancy with no effective therapies. PDA aggressiveness partly stems from its ability to grow within a uniquely dense stroma restricting nutrient access. This study demonstrates that PDA clones that survive chronic nutrient deprivation acquire reversible non-genetic adaptations allowing them to switch between metabolic states optimal for growth under nutrient-replete or nutrient-deprived conditions. One contributing factor to this adaptation mTORC1 activation, which stabilizes glutamine synthetase (GS) necessary for glutamine generation in nutrient-deprived cancer cells. Our findings imply that although total GS levels may not be a prognostic marker for aggressive disease, GS inhibition is of high therapeutic value, as it targets specific cell clusters adapted to nutrient starvation, thus mitigating tumor growth.
Whole exome precision oncology targeting synthetic lethal vulnerabilities across the tumor transcriptome
Authors: Lee Joo Sang , Nair Nishanth Ulhas , Chapman Lesley , Sinha Sanju , Wang Kun , Cha Houngui , Rubin Eitan , Berger Raanan , Lazar Vladimir , Kurzrock Razelle , Gilbert Mark R. , Hannenhalli Sridhar , Lee Se-Hoon , Aldape Kenneth , Ruppin Eytan
Published: February 2020

Precision oncology has made significant advances in the last few years, mainly by targeting actionable mutations in cancer driver genes. However, the proportion of patients whose tumors can be targeted therapeutically remains limited. Recent studies have begun to explore the benefit of analyzing tumor transcriptomics data to guide patient treatment, raising the need for new approaches for systematically accomplishing that. Here we show that computationally derived genetic interactions can successfully predict patient response. Assembling a broad repertoire of 32 datasets spanning more than 1,500 patients and including both tumor transcriptomics and response data, we predicted the response in 17 out of 21 targeted and 8 out of 11 checkpoint therapy datasets across 8 different cancer types with considerable accuracy, without ever training on these datasets. Analyzing the recently published multi-arm WINTHER trial, we show that the fraction of patients benefitting from transcriptomic-based treatments could potentially be markedly increased from 15% to about 85% by targeting synthetic lethal vulnerabilities in their tumors. In summary, this is the first computational approach to obtain considerable predictive performance across many different targeted and immunotherapy datasets, providing a promising new way for guiding cancer treatment based on the tumor transcriptomics of cancer patients.
CRISPR-mediated isogenic cell-SELEX approach for generating highly specific aptamers against native membrane proteins
Authors: Rosch Jonah C. , Neal Emma H. , Balikov Daniel A. , Rahim Mohsin , Lippmann Ethan S.
Published: February 2020

Introduction

The generation of affinity reagents that bind native membrane proteins with high specificity remains challenging. Mostin vitroselection paradigms utilize different cell types for positive and negative rounds of selection (where the positive selection is against a cell that expresses the desired membrane protein and the negative selection is against a cell that lacks the protein). However, this strategy can yield affinity reagents that bind unintended membrane proteins on the target cells. To address this issue, we developed a systematic evolution of ligands by exponential enrichment (SELEX) scheme that utilizes isogenic pairs of cells generated via CRISPR techniques.

Methods

Using a Caco-2 epithelial cell line with constitutive Cas9 expression, we knocked out theSLC2A1gene (encoding the GLUT1 glucose transporter) via lipofection with synthetic gRNAs. Cell-SELEX rounds were carried out against wild-type and GLUT1-null cells using a single-strand DNA (ssDNA) library. Next-generation sequencing (NGS) was used to quantify enrichment of prospective binders to the wild-type cells.

Results

10 rounds of cell-SELEX were conducted via simultaneous exposure of ssDNA pools to wild-type and GLUT1-null Caco-2 cells under continuous perfusion. The top binders identified from NGS were validated by flow cytometry and immunostaining for their specificity to the GLUT1 receptor.

Conclusions

Our data indicate that highly specific aptamers can be isolated with a SELEX strategy that utilizes isogenic cell lines. This approach should be broadly useful for generating affinity reagents that selectively bind to membrane proteins in their native conformations on the cell surface.
S-Trap eliminates cell culture media polymeric surfactants for effective proteomic analysis of mammalian cell bioreactor supernatants
Authors: Zacchi Lucia F. , Recinos Dinora Roche , Otte Ellen , Aitken Campbell , Hunt Tony , Sandford Vanessa , Lee Yih Yean , Schulz Benjamin L. , Howard Christopher B.
Published: February 2020

Proteomic analysis of bioreactor supernatants can inform on cellular metabolic status, viability, and productivity, as well as product quality, which can in turn help optimize bioreactor operation. Incubating mammalian cells in bioreactors requires the addition of polymeric surfactants such as Pluronic F68, which reduce the sheer stress caused by agitation. However, these surfactants are incompatible with mass spectrometry proteomics and must be eliminated during sample preparation. Here, we compared four different sample preparation methods to eliminate polymeric surfactants from filtered bioreactor supernatant samples: organic solvent precipitation; filter-assisted sample preparation (FASP); S-Trap; and single-pot, solid-phase, sample preparation (SP3). We found that SP3 and S-Trap substantially reduced or eliminated the polymer(s), but S-Trap provided the most robust clean-up and highest quality data. Additionally, we observed that SP3 sample preparation of our samples and in other published datasets was associated with partial alkylation of cysteines, which could impact the confidence and robustness of protein identification and quantification. Finally, we observed that several commercial mammalian cell culture media and media supplements also contained polymers with similar mass spectrometry profiles, and we suggest that proteomic analyses in these media will also benefit from the use of S-Trap sample preparation.
Renoprotective and haemodynamic effects of adiponectin and peroxisome proliferator-activated receptor agonist, pioglitazone, in renal vasculature of diabetic Spontaneously hypertensive rats
Authors: Afzal Sheryar , Sattar Munavvar Abdul , Johns Edward James , Eseyin Olorunfemi. A.
Published: February 2020

Pioglitazone, a therapeutic drug for diabetes, possesses full PPAR-γ agonist activity and increase circulating adiponectin plasma concentration. Plasma adiponectin concentration decreases in hypertensive patients with renal dysfunctions. Present study investigated the reno-protective, altered excretory functions and renal haemodynamic responses to adrenergic agonists and ANG II following separate and combined therapy with pioglitazone in diabetic model of hypertensive rats. Pioglitazone was given orally [10mg/kg/day] for 28 days and adiponectin intraperitoneally [2.5µg/kg/day] for last 7 days. Groups of SHR received either pioglitazone or adiponectin in combination. A group of Wistar Kyoto rats [WKY] served as normotensive controls, whereas streptozotocin administered SHRs served as diabetic hypertensive rats. Metabolic data and plasma samples were taken on day 0, 8, 21 and 28. In acute studies, the renal vasoconstrictor actions of Angiotensin II [ANGII], noradrenaline [NA], phenylephrine [PE] and methoxamine [ME] were determined. Diabetic SHRs control had a higher basal mean arterial blood pressure than the WKY, lower RCBP and plasma adiponectin, higher creatinine clearance and urinary sodium excretion compared to WKY [all P<0.05] which were normalized by the individual drug treatments and to greater degree following combined treatment. Responses to intra-renal administration of NA, PE, ME and ANGII were larger in diabetic SHR than WKY and SHRs [P<0.05]. Adiponectin significantly blunted responses to NA, PE, ME and ANG II in diabetic treated SHRs by 40%, whereas the pioglitazone combined therapy with adiponectin further attenuated the responses to adrenergic agonists by 65%. [all P <0.05]. These findings suggest that adiponectin possesses renoprotective effects and improves renal haemodynamics through adiponectin receptors and PPAR-γ in diabetic SHRs, suggesting that synergism exists between adiponectin and pioglitazone. A cross-talk relationship also supposed to exists between adiponectin receptors, PPAR-γ and alpha adrenoceptors in renal vasculature of diabetic SHRs.
High-throughput discovery of trafficking-deficient variants in the cardiac potassium channelKCNH2: Deep mutational scan ofKCNH2trafficking
Authors: Kozek Krystian A. , Glazer Andrew M. , Ng Chai-Ann , Blackwell Daniel , Egly Christian L. , Vanags Loren R. , Blair Marcia , Mitchell Devyn , Matreyek Kenneth A. , Fowler Douglas M. , Knollmann Bjorn C. , Vandenberg Jamie , Roden Dan M. , Kroncke Brett M.
Published: February 2020

Background

KCHN2encodes the KV11.1 potassium channel responsible forIKr, a major repolarization current during the cardiomyocyte action potential. Variants inKCNH2that decreaseIKrcan cause Type 2 Long QT syndrome, usually due to mistrafficking to the cell surface. Accurately discriminating between variants with normal and abnormal trafficking would help clinicians identify and treat individuals at risk of a major cardiac event. The volume of reported non-synonymousKCNH2variants preclude the use of conventional electrophysiologic methods for functional study.

Objective

To report a high-throughput, multiplexed screening method forKCNH2genetic variants capable of measuring the cell surface abundance of hundreds of missense variants inKCNH2.

Methods

We develop a method to quantitateKCNH2variant trafficking on a pilot region of 11 residues in the S5 helix, and generate trafficking scores for 220/231 missense variants in this region.

Results

For 5/5 variants, high-throughput trafficking scores validated when tested in single variant flow cytometry and confocal microscopy experiments. We additionally compare our results with planar patch electrophysiology and find that loss-of-trafficking variants do not produceIKr, but that some variants which traffic normally may still be functionally compromised.

Conclusions

Here, we describe a new method for detecting trafficking-deficient variants inKCNH2in a multiplexed assay. This new method accurately generates trafficking data for variants inKCNH2and can be readily extended to all residues in Kv11.1 and to other cell surface proteins.

CLINICAL IMPLICATIONS

Hundreds ofKCNH2variants have been observed to date, and thousands more will be found as clinical and population sequencing efforts become increasingly widespread. The major mechanism of KV11.1 loss of function is misfolding and failure to traffic to the cell surface. Deep mutational scanning ofKCNH2trafficking is a scalable, high-throughput method that can help identify new loss of function variants and decipher the large number ofKCNH2variants being found in the population.
Microfluidic platform for live cell imaging of 3D cultures with clone retrieval
Authors: Mulas Carla , Hodgson Andrew C , Kohler Timo N , Agley Chibeza C , Hollfelder Florian , Smith Austin , Chalut Kevin
Published: February 2020

Combining live imaging with the ability to retrieve individual cells of interest remains a technical challenge. These combined methods are of particular interest when studying highly dynamic or transient, asynchronous or heterogeneous cell biological and developmental processes. Here we present a method to encapsulate live cells in a 3D hydrogel matrix, via droplet compartmentalisation. Using a small-scale screen, we optimised matrix conditions for the culture and multilineage differentiation of mouse embryonic stem (ES) cells. Moreover, we designed a custom microfluidic platform that is compatible with live imaging. With this platform we are able to retain or extract individual bead/droplets by media flow only, obviating the need for enzymatic cell removal from the platform. We show that we can differentiate mES cells, monitor reporter expression by live imaging, and retrieve individual droplets for functional assays, correlating reporter expression with functional response. Overall, we present a highly flexible 3D cell encapsulation and microfluidic platform that enables both monitoring of cellular dynamics and retrieval for molecular and functional assays.
Fast qualitYconTrol meThod foRderIved diffUsionMetrics (YTTRIUM) in big data analysis: UK Biobank 18608 example
Authors: Maximov Ivan I. , van der Meer Dennis , de Lange Ann-Marie , Kaufmann Tobias , Shadrin Alexey , Frei Oleksandr , Wolfers Thomas , Westlye Lars T.
Published: February 2020

Deriving reliable information about the structural and functional architecture of the brainin vivois critical for the clinical and basic neurosciences. In the new era of large population-based datasets, when multiple brain imaging modalities and contrasts are combined in order to reveal latent brain structural patterns and associations with genetic, demographic and clinical information, automated and stringent quality control (QC) procedures are important. Diffusion magnetic resonance imaging (dMRI) is a fertile imaging technique for probing and visualising brain tissue microstructurein vivo,and has been included in most standard imaging protocols in large-scale studies. Due to its sensitivity to subject motion and technical artefacts, automated QC procedures prior to statistical analyses of dMRI data are required to minimise the influence of noise and artefacts. Here, we introduce Fast qualitY conTrol meThod foR derIved diffUsion Metrics (YTTRIUM), a computationally efficient QC method utilising structural similarity to evaluate image quality and mean diffusion metrics. As an example, we applied YTTRIUM in the context of tract-based spatial statistics to assess associations between age and kurtosis imaging and white matter tract integrity in UK Biobank data (n = 18,608). In order to assess the influence of outliers on results obtained using machine learning approaches, we tested the effects of applying YTTRIUM on brain age prediction. We demonstrated that the proposed QC pipeline represents an efficient approach for identifying poor quality datasets and artifacts and increase the accuracy of machine learning based brain age prediction.
Vps54 regulatesDrosophilaneuromuscular junction development and controls postsynaptic density composition via a Rab7-dependent mechanism
Authors: Patel Prajal H. , Wilkinson Emily C. , Starke Emily L. , McGimsey Malea R. , Blankenship J. Todd , Barbee Scott A.
Published: February 2020

Vps54 is a subunit of the Golgi-associated retrograde protein (GARP) complex, which is involved in tethering endosome-derived vesicles to thetrans-Golgi network (TGN). In the wobbler mouse, a model for human motor neuron (MN) disease, reduction in the levels of Vps54 causes neurodegeneration. However, it is unclear how disruption of GARP-mediated vesicle transport leads to MN dysfunction and ultimately neurodegeneration. To better understand the role of Vps54 in MNs, we have disrupted expression of theVps54ortholog inDrosophilaand examined the impact on the larval neuromuscular junction (NMJ). Here, we show that both null mutants and MN-specific knockdown ofVps54leads to NMJ overgrowth. Reduction ofVps54partially disrupts localization of the t-SNARE, Syntaxin-16, to the TGN but has no impact on endosomal pools. Presynaptic knockdown ofVps54in MNs combined with overexpression of the small GTPases Rab5, Rab7, or Rab11 suppresses theVps54NMJ phenotype. Conversely, knockdown ofVps54combined with overexpression of dominant negative Rab7 causes axonal and behavioral abnormalities including a decrease in postysynaptic Dlg and GluRIIB levels without any effect on GluRIIA. Taken together, these data suggest thatVps54controls larval MN axon development and postsynaptic density composition by modulating Rab7-mediated endosomal trafficking.