bluuebunny/crossref_metadata_2025 huggingface dataset:
HES1 is a Critical Mediator of the SHH-GLI3 Axis in Regulating Digit Number
Authors: Sharma Deepika , Mirando Anthony J. , Leinroth Abigail , Long Jason T. , Karner Courtney M. , Hilton Matthew J.
Published: June 2020

Sonic Hedgehog/GLI3 signaling is critical in regulating digit number, such that Gli3-deficiency results in polydactyly and Shh-deficiency leads to digit number reductions. Anterior-posterior SHH/GLI3 signaling gradients regulate cell cycle factors controlling mesenchymal cell proliferation, while simultaneously regulatingGrem1to coordinate BMP-induced chondrogenesis. SHH/GLI3 also coordinates the expression of additional genes, however their importance in digit formation remain unknown. Utilizing genetic and molecular approaches, we identified HES1 as a key transcriptional regulator downstream of SHH/GLI signaling capable of inducing preaxial polydactyly (PPD), required for Gli3-deficient PPD, and capable of overcoming digit number constraints of Shh-deficiency. Our data indicate that HES1, a direct SHH/GLI signaling target, induces mesenchymal cell proliferation via suppression ofCdkn1b, while inhibiting chondrogenic genes and the anterior autopod boundary regulator,Pax9. These findings fill gaps in knowledge regarding digit number and patterning, while creating a comprehensive framework for our molecular understanding of critical mediators of SHH/GLI3 signaling.
Antibiotic concentrations in the sinonasal secretions and tissue in CRS patients after oral therapy: a randomized trial
Authors: Siu Joey , Klingler Lilian , Wang Yi , Hung Cheung-Tak , Jeong Soo Hee , Smith Susan , Tingle Malcolm , Mackenzie Brett Wagner , Biswas Kristi , Douglas Richard
Published: June 2020

Background

Despite the widespread prescription of antibiotics for patients with chronic rhinosinusitis (CRS), the extent to which drug distribution to the sinonasal mucosa influences their efficacy remains largely undefined.

Methods

Thirty subjects undergoing functional endoscopic sinus surgery (FESS) for bilateral CRS were randomized to one of three groups: 1) doxycycline (100 mg daily for seven days) 2) roxithromycin (300 mg daily for seven days) and 3) control (no antibiotics given). Drug levels were measured using liquid chromatography-tandem mass spectrometry in sinonasal secretions, sinonasal tissues and serum at steady state. Nasal endoscopy (Modified Lund-Kennedy) and Gastrointestinal Symptom Rating Scale (GSRS) scores were recorded.

Results

Antibiotic concentrations in the nasal secretions were significantly lower compared to those in the serum and tissue (mean mucus/serum ratio at steady state = 0.16 and 0.37 for doxycycline and roxithromycin respectively;p<0.01). A short course of antibiotic intake did not correlate with any difference in clinical outcomes except where slightly higher GSRS scores were reported in the roxithromycin group (p=0.04).

Conclusions

Although the efficacy of doxycycline and roxithromycin in sinonasal mucusin vivocannot be predicted solely from reported minimum inhibitory concentrations, given the added complication of bacterial biofilm antimicrobial tolerance, these results suggest that low mucosal penetration of antibiotics may be one of the factors contributing to the limited efficacy of these agents in the treatment of CRS.
A new system in qualitative RT-PCR detecting SARS-CoV-2 in biological samples: an Italian experience
Authors: Favaro Marco , Mattina Walter , Pistoia Enrico Salvatore , Gaziano Roberta , Di Francesco Paolo , Middleton Simon , D’Angelo Silvia , Altarozzi Tullio , Fontana Carla
Published: June 2020

In the last moths the world was faced with the pandemic of a new severe acute respiratory syndrome coronavirus (SARS-CoV) and the majority of the Nations have yet to come out of it. Numerous assays have emerged to meet SARS-CoV-2 diagnostic needs. A clear knowledge of these assays’ parameters is essential to choose the proper test by clinical microbiologists. Unfortunately, the latter cannot be the unique criterion that guides test selection as - given the great demand - shortcomings of commercial kits is also a great issue. Aimed by the intention of overcoming both difficulties we have developed a new qualitative RT-PCR probe based for COVID-19 detection. The system detects three genes of SARS-CoV-2: RNA-dependent RNA polymerase (RdRp), envelope (E) and nucleocapsid (N) and β-actin gene used as endogenous internal control. The results of our assay show a total agreement with those obtained using a commercially available kit, with the exception of two specimens which did not pass the endogenous internal control. Moreover, our kit was designed to be open either for nucleic acid extraction step or on the RT-PCR assay to be carried out on several instruments. Thus, it is free from the industrial production logics of closed systems and conversely it is hypothetically available for distribution on large numbers in any microbiological laboratories. Presently, the kit is currently distributed worldwide
GWAS of stool frequency reveals genes, pathways, and cell types relevant to human gastrointestinal motility and irritable bowel syndrome
Authors: Bonfiglio Ferdinando , Liu Xingrong , Smillie Christopher , Pandit Anita , Kurilshikov Alexander , Bacigalupe Rodrigo , Zheng Tenghao , Nim Hieu , Garcia-Etxebarria Koldo , Bujanda Luis , Andreasson Anna , Agreus Lars , Walter Susanna , Abecasis Goncalo , Eijsbouts Chris , Jostins Luke , Parkes Miles , Hughes David A , Timpson Nicholas , Raes Jeroen , Franke Andre , Kennedy Nicholas A , Regev Aviv , Zhernakova Alexandra , Simren Magnus , Camilleri Michael , D’Amato Mauro
Published: June 2020

Objective

Gut dysmotility is associated with constipation, diarrhea, and functional gastrointestinal disorders (FGID) like irritable bowel syndrome (IBS). Its molecular underpinnings, and their anomalies in FGID disorders are poorly characterized, hence we sought to gain mechanistic insight through a large-scale genetic investigation.

Design

We used stool frequency (STL-FRQ) as a (surrogate) quantitative trait to study the genetics of gut motility, exploiting questionnaire and genotype data from UK Biobank and four smaller population-based cohorts (LifeLines-Deep, Genes for Good, Flemish Gut Flora Project and PopCol), in a GWAS meta-analysis spanning 8,817,117 high-quality SNP markers and 167,875 individuals of European descent.

Results

We identify 13 genome-wide significant loci (P≤5.0×10−8) harboring prioritized genes that are: i) involved in sensory perception and neurotransmitter/neuropeptide signaling; ii) enriched for their expression in enteric motor neurons associated with the control of peristalsis (P=7.0×10−8) iii) previously linked to other traits and conditions, including GI motility and dysmotility syndromes, and the response to their pharmacological treatment. The genetic architecture of STL-FRQ most strongly correlates with that of IBS (rg=0.42; P=1.1×10−3). In UK Biobank, the risk of IBS with diarrhea was 4x higher in individuals from the top 1% of the distribution of polygenic scores (PGS) computed based on STL-FRQ GWAS summary statistics (ORs=4.14; P=1.2×10−97).

Conclusion

We identify loci harboring genes with a plausible role in GI motility, possibly acting via neurotransmission and similar pathways in specialized enteric neurons. The demonstrated relevance of these findings to IBS warrants further study for the identification of actionable pathomechanisms in the dysmotility syndromes.
Two distinct immunopathological profiles in autopsy lungs of COVID-19
Authors: Nienhold Ronny , Ciani Yari , Koelzer Viktor H. , Tzankov Alexandar , Haslbauer Jasmin D. , Menter Thomas , Schwab Nathalie , Henkel Maurice , Frank Angela , Zsikla Veronika , Willi Niels , Kempf Werner , Hoyler Thomas , Barbareschi Mattia , Moch Holger , Tolnay Markus , Cathomas Gieri , Demichelis Francesca , Junt Tobias , Mertz Kirsten D.
Published: June 2020

Coronavirus Disease 19 (COVID-19) is a respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which has grown to a worldwide pandemic with substantial mortality. Immune mediated damage has been proposed as a pathogenic factor, but immune responses in lungs of COVID-19 patients remain poorly characterized. Therefore we conducted transcriptomic, histologic and cellular profiling ofpost mortemCOVID-19 (n=34 tissues from 16 patients) and normal lung tissues (n=9 tissues from 6 patients). Two distinct immunopathological reaction patterns of lethal COVID-19 were identified. One pattern showed high local expression of interferon stimulated genes (ISGhigh) and cytokines, high viral loads and limited pulmonary damage, the other pattern showed severely damaged lungs, low ISGs (ISGlow), low viral loads and abundant infiltrating activated CD8+ T cells and macrophages. ISGhighpatients died significantly earlier after hospitalization than ISGlowpatients. Our study may point to distinct stages of progression of COVID-19 lung disease and highlights the need for peripheral blood biomarkers that inform about patient lung status and guide treatment.
Development of a SARS-CoV-2 total antibody assay and the dynamics of antibody response over time in hospitalized and non-hospitalized patients with COVID-19
Authors: Vogelzang Erik H. , Loeff Floris C. , Derksen Ninotska IL , Kruithof Simone , Heer Pleuni Ooijevaar-de , van Mierlo Gerard , Linty Federica , Mok Juk Yee , van Esch Wim , de Bruin Sanne , Vlaar Alexander P.J. ,   AmsterdamUMC COVID-19 biobank study group, Seppen Bart , Leeuw Maureen , van Oudheusden Anne J.G. , Buiting Anton G.M. , Jim Kin Ki , Vrielink Hans , Swaneveld Francis , Vidarsson Gestur , van der Schoot C. Ellen , Wever Peter , Li Wentao , van Kuppeveld Frank , Murk Jean-Luc , Bosch Berend Jan , Wolbink Gerrit-Jan , Rispens Theo
Published: June 2020

SARS-CoV-2 infections often cause only mild disease that may evoke relatively low antibody titers compared to patients admitted to hospitals. Generally, total antibody bridging assays combine good sensitivity with high selectivity. Therefore, we developed sensitive total antibody bridging assays for detection of SARS-CoV-2 antibodies to the receptor-binding domain (RBD) and nucleocapsid protein (NP), in addition to conventional isotype-specific assays. Antibody kinetics was assessed in PCR-confirmed hospitalized COVID-19 patients (n=41) and three populations of patients with COVID-19 symptoms not requiring hospital admission: PCR-confirmed convalescent plasmapheresis donors (n=182), PCR-confirmed hospital care workers (n=47), and a group of longitudinally sampled symptomatic individuals highly suspect of COVID-19 (n=14). In non-hospitalized patients, the antibody response to RBD is weaker but follows similar kinetics as has been observed in hospitalized patients. Across populations, the RBD bridging assay identified most patients correctly as seropositive. In 11/14 of the COVID-19-suspect cases, seroconversion in the RBD bridging assay could be demonstrated before day 12; NP antibodies emerged less consistently. Furthermore, we demonstrated the feasibility of finger prick sampling for antibody detection against SARS-CoV-2 using these assays. In conclusion, the developed bridging assays reliably detect SARS-CoV-2 antibodies in hospitalized and non-hospitalized patients, and are therefore well-suited to conduct seroprevalence studies.
Assessing the effect of global travel and contact reductions to mitigate the COVID-19 pandemic and resurgence
Authors: Lai Shengjie , Ruktanonchai Nick W , Carioli Alessandra , Ruktanonchai Corrine W , Floyd Jessica R , Prosper Olivia , Zhang Chi , Du Xiangjun , Yang Weizhong , Tatem Andrew J
Published: June 2020

Travel and physical distancing interventions have been implemented across the World to mitigate the COVID-19 pandemic, but studies are needed to quantify the effectiveness of these measures across regions and time. Timely population mobility data were obtained to measure travel and contact reductions in 135 countries or territories. During the 10 weeks of March 22 – May 30, 2020, domestic travel in study regions has dramatically reduced to a median of 59% (interquartile range [IQR] 43% - 73%) of normal levels seen before the outbreak, with international travel down to 26% (IQR 12% - 35%). If these travel and physical distancing interventions had not been deployed across the World, the cumulative number of cases might have shown a 97-fold (IQR 79 – 116) increase, as of May 31, 2020. However, effectiveness differed by the duration and intensity of interventions and relaxation scenarios, with variations in case severity seen across populations, regions, and seasons.

One Sentence Summary

Travel and physical distancing interventions across the World are key to mitigate the COVID-19 pandemic and resurgence.
Routine laboratory blood tests predict SARS-CoV-2 infection using machine learning
Authors: Yang He S. , Hou Yu , Vasovic Ljiljana V. , Steel Peter , Chadburn Amy , Racine-Brzostek Sabrina E. , Velu Priya , Cushing Melissa M. , Loda Massimo , Kaushal Rainu , Zhao Zhen , Wang Fei
Published: June 2020

Background

Accurate diagnostic strategies to rapidly identify SARS-CoV-2 positive individuals for management of patient care and protection of health care personnel are urgently needed. The predominant diagnostic test is viral RNA detection by RT-PCR from nasopharyngeal swabs specimens, however the results are not promptly obtainable in all patient care locations. Routine laboratory testing, in contrast, is readily available with a turn-around time (TAT) usually within 1-2 hours.

Method

We developed a machine learning model incorporating patient demographic features (age, sex, race) with 27 routine laboratory tests to predict an individual’s SARS-CoV-2 infection status. Laboratory test results obtained within two days before the release of SARS-CoV-2-RT-PCR result were used to train a gradient boosted decision tree (GBDT) model from 3,356 SARS-CoV-2 RT-PCR tested patients (1,402 positive and 1,954 negative) evaluated at a metropolitan hospital.

Results

The model achieved an area under the receiver operating characteristic curve (AUC) of 0.854 (95% CI: 0.829-0.878). Application of this model to an independent patient dataset from a separate hospital resulted in a comparable AUC (0.838), validating the generalization of its use. Moreover, our model predicted initial SARS-CoV-2 RT-PCR positivity in 66% individuals whose RT-PCR result changed from negative to positive within two days.

Conclusion

This model employing routine laboratory test results offers opportunities for early and rapid identification of high-risk SARS-CoV-2 infected patients before their RT-PCR results are available. It may play an important role in assisting the identification of SARS-COV-2 infected patients in areas where RT-PCR testing is not accessible due to financial or supply constraints.
Higher risk of COVID-19 hospitalization for unemployed: an analysis of 1,298,416 health insured individuals in Germany
Authors: Dragano Nico , Rupprecht Christoph J. , Dortmann Olga , Scheider Maria , Wahrendorf Morten
Published: June 2020

Background

Previous research on infectious disease has revealed that infection risk as well as the severity of diseases is related to income and poverty. In this study we investigate if unemployed persons have a higher risk to become hospitalized with a COVID-19 diagnosis compared with employed persons.

Methods

We used routine data on hospitalizations in a study population of 1,298,416persons between the ages 18 and 65 who were enrolled in a German health insurance and who were active on the labour market (either employed or unemployed). Hospital diagnosis of COVID-19 (ICD-10-GM U07.1 and U07.2) were reported on a daily basis from 01.01.2020 to 04.06.2020. We studied if the rate of persons hospitalized with a COVID-19 diagnoses differed by employment situation. Logistic regression models comparing employed with short- and long-term unemployed were calculated adjusting for age and sex.

Results

In total, we observed 1,311 persons who were hospitalized, corresponding to a rate of 100.98 cases per 100.000 in our study population. Rates varied between the groups in different employment situations with lowest rates for employed and highest for long-term unemployed. Odds ratio for a hospitalization was 1.84 (1.64 - 2.07) for long-term and 1.18 (0.75 - 1.85) for short-term unemployed compared with employed persons.

Conclusion

The results are in line with earlier (mainly ecological) studies from the USA and Great Britain which found social inequalities in hospitalization risk. The fact that differences exist in Germany, a country with a universal health care system, indicates socioeconomic differences in the COVID-19 pandemic exists across countries.
Genetic architecture of four smoking behaviors using partitioned h2SNP
Authors: Evans Luke M. , Jang Seonkyeong , Ehringer Marissa A. , Otto Jacqueline M. , Vrieze Scott I. , Keller Matthew C.
Published: June 2020

Background and Aims

Smoking is a leading cause of premature death. Although genome-wide association studies have identified many loci that influence smoking behaviors, much of the genetic variance in these traits remains unexplained. We sought to characterize the genetic architecture of four smoking behaviors through SNP-based heritability (h2SNP) analyses.

Design

We applied recently-developed partitioned h2SNP approaches to smoking behavior traits assessed in the UK Biobank.

Setting

UK Biobank.

Participants

UK Biobank participants of European ancestry. The number of participants varied depending on the trait, from 54,792 to 323,068.

Measurements

Smoking initiation, age of initiation, cigarettes per day (CPD; count, log-transformed, binned, and dichotomized into heavy versus light), and smoking cessation. Imputed genome-wide SNPs.

Findings

We estimated h2SNP(SE)=0.18(0.01) for smoking initiation and 0.12(0.02) for smoking cessation, which were more than twice the previously reported estimates. Estimated age of initiation h2SNP=0.05(0.01) and binned CPD h2SNP=0.1(0.01) were similar to previous reports. These estimates remained substantially below published twin-based h2 of roughly 50%. CPD encoding strongly influenced estimates, with dichotomized CPD h2SNP=0.28. We found significant contributions of low-frequency variants and variants in low linkage-disequilibrium (LD) with surrounding genomic regions. Functional annotations related to LD, allele frequency, sequence conservation, and selective constraint also contributed significantly to the partitioned heritability. We found no evidence of dominance genetic variance for any trait.

Conclusion

h2SNP of these four specific smoking behaviors is modest overall. The patterns of partitioned h2SNP for these highly polygenic traits is consistent with negative selection. We found a predominant contribution of common variants, and our results suggest a role of low-frequency or rare variants, poorly tagged by surrounding regions. Deep sequencing of large samples and/or improved imputation will be required to fully assess the role of rare variants.
Anti-phospholipid antibodies in COVID-19 are different from those detectable in the anti-phospholipid syndrome
Authors: Borghi Maria Orietta , Beltagy Asmaa , Garrafa Emirena , Curreli Daniele , Cecchini Germana , Bodio Caterina , Grossi Claudia , Blengino Simonetta , Tincani Angela , Franceschini Franco , Andreoli Laura , Lazzaroni Maria Grazia , Piantoni Silvia , Masneri Stefania , Crisafulli Francesca , Brugnoni Duilio , Muiesan Maria Lorenza , Salvetti Massimo , Parati Gianfranco , Torresani Erminio , Mahler Michael , Heilbron Francesca , Pregnolato Francesca , Pengo Martino , Tedesco Francesco , Pozzi Nicola , Meroni Pier Luigi
Published: June 2020

Background

Critically ill patients with coronavirus disease 2019 (COVID-19) have a profound hypercoagulable state and often develop coagulopathy which leads to organ failure and death. Because of a prolonged activated partial-thromboplastin time (aPTT), a relationship with anti-phospholipid antibodies (aPL) has been proposed, but results are controversial. Functional assays for aPL (i.e., lupus anticoagulant) can be influenced by concomitant anticoagulation and/or high levels of C reactive protein. The presence of anti-cardiolipin (aCL), anti-beta2-glycoprotein I (anti-β2GPI) and anti-phosphatidylserine/prothrombin (aPS/PT) antibodies was not investigated systematically. Epitope specificity of anti-β2GPI antibodies was not reported.

Objective

To evaluate the prevalence and the clinical association of aPL in a large cohort of COVID-19 patients, and to characterize the epitope specificity of anti-β2GPI antibodies.

Methods

ELISA and chemiluminescence assays were used to test 122 sera of patients suffering from severe COVID-19. Of them, 16 displayed major thrombotic events.

Results

Anti-β2GPI IgG/IgA/IgM were the most frequent in 15.6/6.6/9.0% of patients, while aCL IgG/IgM were detected in 5.7/6.6% by ELISA. Comparable values were found by chemiluminescence. aPS/PT IgG/IgM were detectable in 2.5 and 9.8% by ELISA. No association between thrombosis and aPL was found. Reactivity against domain 1 and 4-5 of β2GPI was limited to 3/58 (5.2%) tested sera for each domain and did not correlate with aCL/anti-β2GPI nor with thrombosis.

Conclusions

aPL show a low prevalence in COVID-19 patients and are not associated with major thrombotic events. aPL in COVID-19 patients are mainly directed against β2GPI but display an epitope specificity different from antibodies in antiphospholipid syndrome.
Extrapolation of Infection Data for the CoVid-19 Virus in 21 Countries and States and Estimate of the Efficiency of Lock Down
Authors: Langel Walter
Published: June 2020

Predictions about the further development of the Corona pandemic are of great public interest but many approaches demand a large number of country specific parameters and are not easily transferable. A special interest of simulations on the pandemic is to trace the effect of politics for reducing the virus spread, since these measures have had an enormous impact on economy and daily life.

Here a simple yet powerful algorithm is introduced for fitting the infection numbers by simple analytic functions. This way, the increase of the case numbers in periods with different regulations can be distinguished, and by extrapolating the fit functions, a forecast for the maximum numbers and time scales are possible. The effect of the restraints such as lock down are demonstrated by comparing the resulting infection history with the likely unconstrained virus spread, and it is shown that a delay of 1-4 weeks before imposing measures aiming at social distancing could have led to a complete infection of the respective populations.

The approach is simply transferable to many different states. Here data from six E.U. countries, the UK, Russia, two Asian countries, the USA and ten states inside the USA with significant case numbers are analyzed, and striking qualitative similarities are found.

Keywords: Covid-19, forecast, analytic fit, France, Germany, Italy, Spain South Korea, New York, Washington, Florida, Michigan, Poland, Sweden, USA, Pennsylvania, China, Russia, UK, California, Illinois, Indiana, Maryland, North Carolina.
Predicting walking response to ankle exoskeletons using data-driven models
Authors: Rosenberg Michael C. , Banjanin Bora S. , Burden Samuel A. , Steele Katherine M.
Published: June 2020

II

Abstract

Despite recent innovations in exoskeleton design and control, predicting subject-specific impacts of exoskeletons on gait remains challenging. We evaluated the ability of three classes of subject-specific phase-varying models to predict kinematic and myoelectric responses to ankle exoskeletons during walking, without requiring prior knowledge of specific user characteristics. Each model – phase-varying (PV), linear phase-varying (LPV), and nonlinear phase-varying (NPV) – leveraged Floquet Theory to predict deviations from a nominal gait cycle due to exoskeleton torque, though the models differed in complexity and expected prediction accuracy. For twelve unimpaired adults walking with bilateral passive ankle exoskeletons, we predicted kinematics and muscle activity in response to three exoskeleton torque conditions. The LPV model’s predictions were more accurate than the PV model when predicting less than 12.5% of a stride in the future and explained 49–70% of the variance in hip, knee, and ankle kinematic responses to torque. The LPV model also predicted kinematic responses with similar accuracy to the more-complex NPV model. Myoelectric responses were challenging to predict with all models, explaining at most 10% of the variance in responses. This work highlights the potential of data-driven phase-varying models to predict complex subject-specific responses to ankle exoskeletons and inform device design and control.
Phylogenetic Studies and Inhibitor Design Targeting Protein Interacting Interface of Nucleoid-Associated Protein HU
Authors: Dey Debayan , Ramakumar Suryanarayanarao
Published: June 2020

The formations of nucleoprotein structures by promiscuous DNA binding proteins like HU are assisted with their protein protein interaction capability with other proteins. InE. coliGal repressosome assembly, GalR piggybacks HU to the critical position on the DNA (hbs site) through a specific GalR–HU interaction using an interface at the bottom of alpha helical region, which we termed as HUpb interface. Similarly, MtbHU also interact with Topoisomerase I with the same interface to enhance its relaxation activity. In an earlier study, we determined the crystal structure of MtbHU, inhibited it using stilbene derivatives which inhibited the cell growth. It motivated us to understand the evolutionary and structural characteristics of the HUpb interface, which has not been investigated previously for HU or for any other NAPs. Our analyses found residue positions corresponding to MtbHU Thr11 to Gln20 form the interface while Ala23 serves the pocket lining residue. Due to ancestral mutations in the duplication event before the HU and IHF split, physicochemical properties of the interface vary among clades. Thus, this interface could engage different proteins in different HU oligomeric states inProteobacteria. Protein-protein interfaces are usually a challenging target due to its flatter surface. In case of MtbHUpb interface, we observed that due to the presence of a partially hydrophobic pocket, small molecule scaffolds could fit into it, while the ligand can be further designed to interact with D17, which is the crucial residue for Topoisomerase I interaction. We used a two-step virtual screening protocol with known drug like molecules as starting set to an aim to re-purpose drugs. Our docking results showed compounds like Maltotetraose, Valrubicin, Iodixanol, Enalkiren, indinavir, Carfilzomib, oxytetracycline, quinalizarin, Raltitrexed, Epigallocatechin and their analogues exhibit high scoring binding at MtbHUpb interface. Our present report gives a model example of an evolutionary study of an interface of nucleoid associated protein, which is used to computationally design inhibitors. This strategy could be in general useful for designing inhibitors for various types of protein-protein interfaces using evolutionary guided design.
Inhibiting actin polymerization does not prevent the fast block to polyspermy in the African clawed frog,Xenopus laevis
Authors: Tembo Maiwase , Sauer Monica L. , Wisner Bennett W. , Beleny David O. , Napolitano Marc A. , Carlson Anne E.
Published: June 2020

Fertilization of an egg by more than one sperm presents one of the earliest and most prevalent obstacles to successful reproduction. As such, eggs employ multiple mechanisms to prevent sperm entry into the nascent zygote. The fast block to polyspermy is a depolarization of the egg membrane initiated by sperm entry and is employed by diverse external fertilizers including frogs and sea urchins. For some external fertilizers, sperm entry is associated with actin polymerization during the initiation of the fast block. We therefore sought to determine whether the fast block to polyspermy in the African clawed frog,Xenopus laevis, requires actin polymerization. Although actin polymerization is required for sperm entry into eggs from diverse external fertilizers, including sea urchins and zebrafish, here we demonstrate that actin polymerization is not required for the fast block to polyspermy inX. laevis.
Pretreatment with LCK Inhibitors Chemosensitizes Cisplatin-Resistant Endometrioid Ovarian Tumors
Authors: Crean-Tate Katie K. , Braley Chad , Dey Goutam , Esakov Emily , Saygin Caner , Trestan Alexandria , Silver Daniel J. , Turaga Soumya M. , Connor Elizabeth , DeBernardo Robert , Michener Chad M. , Rose Peter G. , Lathia Justin , Reizes Ofer
Published: June 2020

Objective

To evaluate LCK inhibitors (LCKi) as chemosensitizing agents for platinum-resistant endometrioid ovarian carcinoma.

Methods

KM Plotter survival data was obtained for endometrioid ovarian cancer based on LCK mRNA expression. Cisplatin resistant endometrioid ovarian carcinoma cell lines were cultured and treated first with LCKi or vehicle, then combination LCKi-cisplatin. Cell viability was assessed via CellTiter-Glo, and apoptosis with Caspase 3/7 Assay. Protein lysates were isolated from treated cells, withγ-H2Ax, a DNA adduct marker, assessed.In vivostudy compared mice treated with vehicle or LCK inhibitor followed by LCK inhibitor, cisplatin, or combination therapy. One-way ANOVA and two sample t-test were used to assess statistical significance with GraphPad Prism.

Results

KM plotter data indicated LCK expression is associated with significantly worse median progression-free survival (HR 3.19, p=0.02), and a trend toward decreased overall survival in endometrioid ovarian tumors with elevated LCK expression (HR 2.45, p=0.41).In vitro, cisplatin resistant ovarian endometrioid cells treated first with LCKi followed by combination LCKi-cisplatin treatment showed decreased cell viability and increased apoptosis. Immunoblot studies revealed inhibition of LCK led to increased expression ofγ-H2AX.In vivoresults demonstrate treatment with LCKi followed by LCKi-cisplatin leads to significantly slowed tumor growth.

Conclusions

We identified a targetable pathway for chemosensitization of platinum resistant endometrioid ovarian cancer with initial treatment of LCKi followed by co-treatment with LCKi-cisplatin.
The validation of new phase-dependent gait stability measures: a modeling approach
Authors: Jin Jian , Kistemaker Dinant , van Dieën Jaap H. , Daffertshofer Andreas , Bruijn Sjoerd M.
Published: June 2020

Identification of individuals at risk of falling is important when designing fall prevention methods. Current measures that estimate gait stability and robustness appear limited in predicting falls in older adults. Inspired by recent findings on changes in phase-dependent local stability within a gait cycle, we devised several phase-dependent stability measures and tested for their usefulness to predict gait robustness in compass walker models. These measures are closely related to the often-employed maximum finite-time Lyapunov exponent and maximum Floquet multiplier that both assess a system’s response to infinitesimal perturbations. As such, they entail linearizing the system but this is realized in a rotating hypersurface orthogonal to the period-one solution followed by estimating the trajectory-normal divergence rate of the swing phases and the foot strikes. We correlated the measures with gait robustness, i.e. the largest perturbation a walker can handle, in two compass walker models with either point or circular feet to estimate their prediction accuracy. To also test for the dependence of the measures under state space transform, we represented the point-feet walker in both Euler-Lagrange and Hamiltonian canonical form. Our simulations revealed that for most of the measures their correlation with gait robustness differs between models and between different state space forms. In particular the latter may jeopardize many stability measures’ predictive capacity for gait robustness. The only exception that consistently displayed strong correlations is the divergence of foot strike. Our results admit challenges of using phase-dependent stability measures as objective means to estimate the risk of falling.
Characterization of nuclear pore complex targeting domains in Pom152 inSaccharomyces cerevisiae
Authors: Brown Jacqueline T. , Haraczy Alexandra J. , Wilhelm Christopher M. , Belanger Kenneth D.
Published: June 2020

Pom152 is a transmembrane protein within the nuclear pore complex (NPC) of fungi that is important for NPC assembly and structure. Pom152 is comprised of a short amino-terminal region that remains on the cytosolic side of the nuclear envelope (NE) and interacts with NPC proteins, a transmembrane domain, and a large, glycosylated carboxy-terminal domain within the NE lumen that self-assembles to form the NPC membrane ring. Here we show that the N-terminal 200 amino acids of Pom152 that include only the amino-terminal and transmembrane regions of the protein are sufficient for localization to the NPC. Full-length, glycosylation-deficient, and truncated Pom152-GFP chimeras expressed in cells containing endogenous Pom152 localize to both NPCs and cortical endoplasmic reticulum (ER). Expression of Pom152-GFP fusions in cells lacking endogenous Pom152 results in detectable localization at only the NE by full-length and amino-terminal Pom152-GFP fusions, but continued retention at both the NE and ER for a chimera lacking just the carboxy-terminal 377 amino acids. Targeted mutations in the amino-terminal and transmembrane domains did not alter Pom152 localization and neither deletion of Pom152 nor its carboxy-terminal glycosylation sites altered the nuclear protein export rate of an Msn5/Kap142 protein cargo. These data narrow the Pom152 region sufficient for NPC localization and provide evidence that alterations in other domains may impact Pom152 targeting or affinity for the NPC.
Loss of genetic variation and sex determination system in North American northern pike characterized by whole-genome resequencing
Authors: Johnson Hollie A , Rondeau Eric B , Sutherland Ben J G , Minkley David R , Leong Jong S , Whitehead Joanne , Despins Cody A , Gowen Brent E , Collyard Brian J , Whipps Christopher M , Farrell John M , Koop Ben F
Published: June 2020

The northern pike Esox lucius is a freshwater fish renowned for having low genetic diversity but ecological success throughout the Northern Hemisphere. Here we generate an annotated chromosome-level genome assembly of 941 Mbp in length with 25 chromosome-length scaffolds using long-reads and chromatin capture technology. We then align whole-genome resequencing data against this reference to genotype northern pike from Alaska through New Jersey (n = 47). A striking decrease in genetic diversity occurs along the sampling range, whereby samples to the west of the North American Continental Divide have substantially higher diversity than populations to the east. As an example, individuals from Interior Alaska in the west and St. Lawrence River in the east have on average 181K and 64K heterozygous SNPs per individual, respectively (i.e., a SNP variant every 3.2 kbp and 11.2 kbp, respectively). Even with such low diversity, individuals clustered with strong support within each population, and this may be related to numerous private alleles in each population. Evidence for recent population expansion was observed for a Manitoba hatchery and the St. Lawrence population (Tajima's D = -1.07 and -1.30, respectively). Non-uniform patterns of diversity were observed across the genome, with large regions showing elevated diversity in several chromosomes, including LG24. In populations with the master sex determining gene amhby still present in the genome, amhby is in LG24. As expected, amhby was largely male-specific in Alaska and the Yukon and absent southeast to these populations, but we also document some amhby(-) males in Alaska and amhby(+) males in the Columbia River. This indicates that rather than a discrete boundary after which amhby was lost in North America, there is a patchwork of presence of this system in the western region. These results support the theory that northern pike recolonized North America from refugia in Alaska and expanded following deglaciation from west to east, with probable founder effects resulting in loss of both neutral and functional diversity including the loss of the sex determination system.
The structure of autocatalytic networks, with application to early biochemistry
Authors: Steel Mike , Xavier Joana C. , Huson Daniel H.
Published: June 2020

Metabolism across all known living systems combines two key features. First, all of the molecules that are required are either available in the environment or can be built up from available resources via other reactions within the system. Second, the reactions proceed in a fast and synchronised fashion via catalysts that are also produced within the system. Building on early work by Stuart Kauffman, a precise mathematical model for describing such self-sustaining autocatalytic systems (RAF theory) has been developed to explore the origins and organisation of living systems within a general formal framework. In this paper, we develop this theory further by establishing new relationships between classes of RAFs and related classes of networks, and developing new algorithms to investigate and visualise RAF structures in detail. We illustrate our results by showing how it reveals further details into the structure of archaeal and bacterial metabolism near the origin of life, and provide techniques to study and visualise the core aspects of primitive biochemistry.
Microbial piggy-back: howStreptomycesspores are transported by motile soil bacteria
Authors: Muok Alise R. , Claessen Dennis , Briegel Ariane
Published: June 2020

Streptomycetes are sessile, soil-dwelling bacteria that produce diverse metabolites that impact plant health and the behavior of microbial communities. Emerging studies have demonstrated thatStreptomycesspores are distributed through a variety of mechanisms, but it remains unclear how spores are transported to their preferred micro-environments, such as plant roots. Here, we show thatStreptomycesspores are capable of utilizing the motility machinery of other soil bacteria and are transported on the centimeter scale. Motility assays and microscopy studies reveal thatStreptomycesspores are transported to plant tissues by interacting directly with the flagella of both gram-positive and gram-negative bacteria. Genetics experiments demonstrate that this form of motility, called piggy-backing, is facilitated by conserved structural proteins present on the surface ofStreptomycesspores. These results demonstrate that non-motile bacteria are capable of utilizing the motility machinery of other microbes to complete necessary stages of their lifecycle, and that this mode of transport may be ubiquitous in nature.
AvrE1 and HopR1 fromPseudomonas syringaepv.actinidiaeare additively required for full virulence on kiwifruit
Authors: Jayaraman Jay , Yoon Minsoo , Applegate Emma R. , Stroud Erin A. , Templeton Matthew D.
Published: June 2020

Pseudomonas syringaepv.actinidiaeICMP 18884 biovar 3 (Psa3) produces necrotic lesions during infection of its kiwifruit host. Bacterial growthin plantaand lesion formation are dependent upon a functional type III secretion system (T3S), which translocates multiple effector proteins into host cells. Associated with the T3S locus is the conserved effector locus (CEL), which has been characterised and shown to be essential for the full virulence in otherP. syringaepathovars. Two effectors at the CEL,hopM1andavrE1, as well as anavrE1-related non-CEL effector,hopR1, have been shown to be redundant in the model pathogenP. syringaepv.tomatoDC3000 (Pto), a close relative ofPsa. However, it is not known whether CEL-related effectors are required forPsapathogenicity. ThePsa3allele ofhopM1, and its associated chaperone,shcM, have diverged significantly from their orthologs inPto. Furthermore, the CEL effectorhopAA1-1, as well as a related non-CEL effector,hopAA1-2, have both been pseudogenised. We have shown that HopM1 does not contribute toPsa3virulence due to a truncation inshcM, a truncation conserved in thePsalineage, likely due to the need to evade HopM1-triggered immunity in kiwifruit. We characterised the virulence contribution of CEL and related effectors inPsa3and found that onlyavrE1andhopR1, additively, are required forin plantagrowth and lesion production. This is unlike the redundancy described for these effectors inPtoand indicates that these twoPsa3genes are key determinants essential for kiwifruit bacterial canker disease.
The tumor suppressor microRNA let-7 inhibits human LINE-1 retrotransposition
Authors: Tristán-Ramos Pablo , Rubio-Roldan Alejandro , Peris Guillermo , Sánchez Laura , Amador-Cubero Suyapa , Viollet Sebastien , Cristofari Gael , Heras Sara R.
Published: June 2020

Nearly half of the human genome is made of transposable elements (TEs) whose activity continues to impact its structure and function. Among them, Long INterspersed Element class 1 (LINE-1 or L1) elements are the only autonomously active TEs in humans. L1s are expressed and mobilized in different cancers, generating mutagenic insertions that could affect malignancy. Tumor suppressor microRNAs are ∼22nt RNAs that post-transcriptionally regulate oncogene expression and are frequently downregulated in cancer. Here we explore whether they also influence L1 mobilization. We found that downregulation of let-7 correlates with accumulation of L1 insertions in human lung cancer. Furthermore, we demonstrate that let-7 binds to the L1 mRNA and impairs the translation of the second L1-encoded protein, ORF2p, reducing its mobilization. Overall, our data uncover a new role for let-7, one of the most relevant microRNAs, which is to maintain somatic genome integrity by restricting L1 retrotransposition.
Tunable phenotypic variability through an autoregulatory alternative sigma factor circuit
Authors: Schwall Christian P. , Loman Torkel , Martins Bruno M.C. , Cortijo Sandra , Villava Casandra , Kusmartsev Vassili , Livesey Toby , Saez Teresa , Locke James C. W.
Published: June 2020

Genetically identical individuals in bacterial populations can display significant phenotypic variability. This variability can be functional, for example by allowing a fraction of stress prepared cells to survive an otherwise lethal stress. The optimal fraction of stress prepared cells depends on environmental conditions. However, how bacterial populations modulate their level of phenotypic variability remains unclear. Here we show that the alternative sigma factor σVcircuit inB. subtilisgenerates functional phenotypic variability that can be tuned by stress level, environmental history, and genetic perturbations. Using single-cell time-lapse microscopy and microfluidics, we find the fraction of cells that immediately activate σVunder lysozyme stress depends on stress level and on a memory of previous stress. Iteration between model and experiment reveals that this tunability can be explained by the autoregulatory feedback structure of thesigVoperon. As predicted by the model, genetic perturbations to the operon also modulate the response variability. The conserved sigma-anti-sigma autoregulation motif is thus a simple mechanism for bacterial populations to modulate their heterogeneity based on their environment.
Biophysical characterization of the human K0513 protein
Authors: Nemukondeni Ndivhuwo , Arowolo Afolake , Shonhai Addmore , Zininga Tawanda , Burger Adélle
Published: June 2020

Glioblastoma multiforme (GBM) is an aggressive grade IV primary malignant tumour which accounts for 78 % of all brain tumours.K0513is a GBM biomarker that is upregulated in the invasive phenotype.K0513is expressed ubiquitously and is reportedly enriched in the cerebral cortex of the brain.K0513is further implicated in signalling pathways involving neuroplasticity, cytoskeletal regulation and apoptosis. The protein encoded byK0513is a prospective biomarker for pancreatic cancer prognosis. However, the gene product ofK0153is not well characterised. This study focused on structure-function characterisation of human K0513 protein. To this end, we employed bioinformatics analysis and biophysical approaches to characterize the protein.In silicostructural characterisation of the human K0513 protein suggests the presence of a SET binding factor 2 (SBF2) domain and a transmembrane region. The SBF2 domain is found in the Myotubularin-related protein 13 (MTMR13), which may function as a nucleotide exchange factor for the RAS-associated GTPase, Rab28. K0513 was predicted to interact with RAS-associated GTPase, Rab3a. Secondary structure prediction revealed K0513 to be predominantly α-helical in nature. The predicted three-dimensional model of K0513 showed a globular fold, suggesting that the protein is water-soluble. K0513 was heterologously expressed inE. coliXL1-Blue cells and subsequent purification yielded 80 % soluble protein. Biophysical characterisation using tryptophan-based fluorescence spectroscopy and limited proteolysis showed the conformation of K0513 is mostly unperturbed in the presence of nucleotides. Interestingly, K0513 was detected in lung carcinoma, fibrosarcoma and cervical adenocarcinoma cells, supporting its possible role in cancer signalling pathways.
Selective vulnerability of aneuploid human cancer cells to inhibition of the spindle assembly checkpoint
Authors: Cohen-Sharir Yael , McFarland James M. , Abdusamad Mai , Marquis Carolyn , Tang Helen , Ippolito Marica R. , Bernhard Sara V. , Laue Kathrin , Malaby Heidi L.H. , Jones Andrew , Kazachkova Mariya , Lyons Nicholas , Nagaraja Ankur , Bass Adam J. , Beroukhim Rameen , Santaguida Stefano , Stumpff Jason , Golub Todd R. , Storchova Zuzana , Ben-David Uri
Published: June 2020

Selective targeting of aneuploid cells is an attractive strategy for cancer treatment. Here, we mapped the aneuploidy landscapes of ~1,000 human cancer cell lines and classified them by their degree of aneuploidy. Next, we performed a comprehensive analysis of large-scale genetic and chemical perturbation screens, in order to compare the cellular vulnerabilities between near-diploid and highly-aneuploid cancer cells. We identified and validated an increased sensitivity of aneuploid cancer cells to genetic perturbation of core components of the spindle assembly checkpoint (SAC), which ensures the proper segregation of chromosomes during mitosis. Surprisingly, we also found highly-aneuploid cancer cells to belesssensitive to short-term exposures to multiple inhibitors of the SAC regulatorTTK. To resolve this paradox and to uncover its mechanistic basis, we established isogenic systems of near-diploid cells and their aneuploid derivatives. Using both genetic and chemical inhibition ofBUB1B,MAD2andTTK, we found that the cellular response to SAC inhibition depended on the duration of the assay, as aneuploid cancer cells became increasingly more sensitive to SAC inhibition over time. The increased ability of aneuploid cells to slip from mitotic arrest and to keep dividing in the presence of SAC inhibition was coupled to aberrant spindle geometry and dynamics. This resulted in a higher prevalence of mitotic defects, such as multipolar spindles, micronuclei formation and failed cytokinesis. Therefore, although aneuploid cancer cells can overcome SAC inhibition more readily than diploid cells, the proliferation of the resultant aberrant cells is jeopardized. At the molecular level, analysis of spindle proteins identified a specific mitotic kinesin,KIF18A, whose levels were drastically reduced in aneuploid cancer cells. Aneuploid cancer cells were particularly vulnerable toKIF18Adepletion, andKIF18Aoverexpression restored the sensitivity of aneuploid cancer cells to SAC inhibition. In summary, we identified an increased vulnerability of aneuploid cancer cells to SAC inhibition and explored its cellular and molecular underpinnings. Our results reveal a novel synthetic lethal interaction between aneuploidy and the SAC, which may have direct therapeutic relevance for the clinical application of SAC inhibitors.