bluuebunny/crossref_metadata_2025 huggingface dataset:
Control of dynamic cell behaviors during angiogenesis and anastomosis by Rasip 1
Authors: Lee Minkyoung , Betz Charles , Paatero Ilkka , Schellinx Niels , Yin Jianmin , Wilson Christopher William , Ye Weilan , Affolter Markus , Belting Heinz-Georg
Published: August 2020

Organ morphogenesis is driven by a wealth of tightly orchestrated cellular behaviors, which ensure proper organ assembly and function. Many of these cell activities involve cell-cell interactions and remodeling of the F-actin cytoskeleton. Here, we analyze the requirement for Rasip1 (Ras-interacting protein 1), an endothelial-specific regulator of junctional dynamics, during blood vessel formation. Phenotype analysis ofrasip1mutants in zebrafish embryos reveal distinct requirements for Rasip1 during sprouting angiogenesis, vascular anastomosis and lumen formation. During angiogenic sprouting, Rasip1 is required for efficient cell pairing, which is essential for multicellular tube formation. High-resolution time-lapse analyses show that these cell pairing defects are caused by a destabilization of tricellular junctions suggesting that tri-cellular junctions may serve as a counterfort to tether sprouting endothelial cells during morphogenetic cell rearrangements. During anastomosis, Rasip1 is required to establish a stable apical membrane compartment;rasip1mutants display ectopic, reticulated junctions and the apical compartment is frequently collapsed. Loss of Ccm1 and Heg1 function leads to junctional defects similar to those seen inrasip1mutants. Analysis ofradil-bsingle andrasip1/radil-bdouble mutants reveal distinct and overlapping functions of both proteins. While Rasip1 and Radil-b have similar functions during angiogenic sprouting, the junction formation during anastomosis may primarily depend on Rasip1.
FAIR sharing of molecular visualization experiences: from pictures in the cloud to collaborative virtual reality exploration in immersive 3D environments
Authors: Martinez Xavier , Baaden Marc
Published: August 2020

Motivated by the current Covid-19 pandemic that has spurred a substantial flow of structural data we describe how molecular visualization experiences can be used to make these datasets accessible to a broad audience. Using a variety of technology vectors related to the cloud, 3D- and virtual reality gear, we examine how to share curated visualizations of structural biology, modeling and/or bioinformatics datasets for interactive and collaborative exploration. We discuss F.A.I.R. as overarching principle for sharing such visualizations. We provide four initial example scenes related to recent Covid-19 structural data together with a ready-to-use (and share) implementation in the UnityMol software.

Synopsis

Visualization renders structural molecular data accessible to a broad audience. We describe an approach to share molecular visualization experiences based on FAIR principles. Our workflow is exemplified with recent Covid-19 related data.
Network Medicine Framework Shows Proximity of Polyphenol Targets and Disease Proteins is Predictive of the Therapeutic Effects of Polyphenols
Authors: do Valle Italo F. , Roweth Harvey G. , Malloy Michael W. , Moco Sofia , Barron Denis , Battinelli Elisabeth , Loscalzo Joseph , Barabási Albert-László
Published: August 2020

Polyphenols, natural products present in plant-based foods, play a protective role against several complex diseases through their antioxidant activity and by diverse molecular mechanisms. Here we developed a network medicine framework to uncover the mechanistic roles of polyphenols on health by considering the molecular interactions between polyphenol protein targets and proteins associated with diseases. We find that the protein targets of polyphenols cluster in specific neighborhoods of the human interactome, whose network proximity to disease proteins is predictive of the molecule’s known therapeutic effects. The methodology recovers known associations, such as the effect of epigallocatechin 3-O-gallate on type 2 diabetes, and predicts that rosmarinic acid (RA) has a direct impact on platelet function, representing a novel mechanism through which it could affect cardiovascular health. We experimentally confirm that RA inhibits platelet aggregation and alpha granule secretion through inhibition of protein tyrosine phosphorylation, offering direct support for the predicted molecular mechanism. Our framework represents a starting point for mechanistic interpretation of the health effects underlying food-related compounds, allowing us to integrate into a predictive framework knowledge on food metabolism, bioavailability, and drug interaction.
Analysis of Inositol Phosphate Metabolism by Capillary Electrophoresis Electrospray Ionization Mass Spectrometry (CE-ESI-MS)
Authors: Qiu Danye , Wilson Miranda S. , Eisenbeis Verena B. , Harmel Robert K. , Riemer Esther , Haas Thomas M. , Wittwer Christopher , Jork Nikolaus , Gu Chunfang , Shears Stephen B. , Schaaf Gabriel , Kammerer Bernd , Fiedler Dorothea , Saiardi Adolfo , Jessen Henning J.
Published: August 2020

The analysis ofmyo-inositol phosphates (InsPs) andmyo-inositol pyrophosphates (PP-InsPs) is a daunting challenge due to the large number of possible isomers, the absence of a chromophore, the high charge density, the low abundance, and the instability of the esters and anhydrides. Given their importance in biology, an analytical approach to follow and understand this complex signaling hub is highly desirable. Here, capillary electrophoresis (CE) coupled to electrospray ionization mass spectrometry (ESI-MS) is implemented to analyze complex mixtures of InsPs and PP-InsPs with high sensitivity. Stable isotope labeled (SIL) internal standards allow for matrix-independent quantitative assignment. The method is validated in wild-type and knockout mammalian cell lines and in model organisms. SIL-CE-ESI-MS enables for the first time the accurate monitoring of InsPs and PP-InsPs arising from compartmentalized cellular synthesis pathways, by feeding cells with either [13C6]-myo-inositol or [13C6]-D-glucose. In doing so, we uncover that there must be unknown inositol synthesis pathways in mammals, highlighting the unique potential of this method to dissect inositol phosphate metabolism and signalling.
Ericoid mycorrhizal diversity increases with soil age and progressive phosphorus limitation across a 4.1 million-year chronosequence
Authors: Leopold Devin R. , Peay Kabir G. , Vitousek Peter M. , Fukami Tadashi
Published: August 2020

Ericaceous plants rely on ericoid mycorrhizal fungi for nutrient acquisition. However, the factors that affect the composition and structure of these fungal communities remain largely unknown. Here, we use a 4.1-myr soil chronosequence in Hawaii to test the hypothesis that changes in nutrient availability with soil age determine the diversity and species composition of fungi associated with ericoid roots. We sampled roots of a native Hawaiian plant,Vaccinium calycinum, and used DNA metabarcoding to quantify changes in fungal diversity and species composition. We also used a fertilization experiment at the youngest and oldest sites to assess the importance of nutrient limitation. We found an increase in diversity and a clear pattern of species turnover across the chronosequence, driven largely by putative ericoid mycorrhizal fungi. Fertilization with nitrogen at the youngest site and phosphorus at the oldest site reduced total fungal diversity, suggesting a direct role of nutrient limitation. Our results also reveal the presence of novel fungal species associated with Hawaiian Ericaceae and suggest a greater importance of phosphorus availability for communities of ericoid mycorrhizal fungi than is generally assumed.
Three-dimensional residual channel attention networks denoise and sharpen fluorescence microscopy image volumes
Authors: Chen Jiji , Sasaki Hideki , Lai Hoyin , Su Yijun , Liu Jiamin , Wu Yicong , Zhovmer Alexander , Combs Christian A. , Rey-Suarez Ivan , Chang Hungyu , Huang Chi Chou , Li Xuesong , Guo Min , Nizambad Srineil , Upadhyaya Arpita , Lee Shih-Jong J. , Lucas Luciano A.G. , Shroff Hari
Published: August 2020

We demonstrate residual channel attention networks (RCAN) for restoring and enhancing volumetric time-lapse (4D) fluorescence microscopy data. First, we modify RCAN to handle image volumes, showing that our network enables denoising competitive with three other state-of-the-art neural networks. We use RCAN to restore noisy 4D super-resolution data, enabling image capture over tens of thousands of images (thousands of volumes) without apparent photobleaching. Second, using simulations we show that RCAN enables class-leading resolution enhancement, superior to other networks. Third, we exploit RCAN for denoising and resolution improvement in confocal microscopy, enabling ∼2.5-fold lateral resolution enhancement using stimulated emission depletion (STED) microscopy ground truth. Fourth, we develop methods to improve spatial resolution in structured illumination microscopy using expansion microscopy ground truth, achieving improvements of ∼1.4-fold laterally and ∼3.4-fold axially. Finally, we characterize the limits of denoising and resolution enhancement, suggesting practical benchmarks for evaluating and further enhancing network performance.
Lipoprotein particles interact with membranes and transfer their cargo without receptors
Authors: Plochberger Birgit , Sych Taras , Weber Florian , Novacek Jiri , Axmann Markus , Stangl Herbert , Sezgin Erdinc
Published: August 2020

Lipid transfer from lipoprotein particles to cells is essential for lipid homeostasis. High density lipoprotein (HDL) particles are mainly captured by cell-membrane-associated scavenger receptor class B type 1 (SR-B1) from the blood stream while low and very low density lipoprotein (LDL, VLDL) particles are mostly taken up by receptor-mediated endocytosis. However, the role of the target lipid membrane itself in the transfer process has been largely neglected so far. Here, we study how lipoprotein particles (HDL, LDL and VLDL) interact with synthetic lipid bilayers and cell-derived membranes and transfer their cargo subsequently. Employing cryo-electron microscopy, spectral imaging and fluorescence (cross) correlation spectroscopy allowed us to observe integration of all major types of lipoprotein particles into the membrane and delivery of their cargo in a receptor-independent manner. Importantly, biophysical properties of the target cell membranes change upon cargo delivery. The concept of receptor-independent interaction of lipoprotein particles with membranes helps to better understand lipoprotein particle biology and can be exploited for novel treatments of dyslipidemia diseases.
Ribonucleotide incorporation characteristics around yeast autonomously replicating sequences reveal the labor division of replicative DNA polymerases
Authors: Xu Penghao , Storici Francesca
Published: August 2020

Ribonucleoside monophosphate (rNMP) incorporation in DNA is a natural and prominent phenomenon resulting in DNA structural change and genome instability. While DNA polymerases have different rNMP incorporation rates, little is known whether these enzymes incorporate rNMPs following specific sequence patterns. In this study, we analyzed a series of rNMP incorporation datasets, generated from three rNMP mapping techniques, and obtained fromSaccharomyces cerevisiaecells expressing wild-type or mutant replicative DNA polymerase and ribonuclease H2 genes. We performed computational analyses of rNMP sites around early and late firing autonomously replicating sequences (ARS’s) of the yeast genome, from which bidirectional, leading and lagging DNA synthesis starts. We find the preference of rNMP incorporation on the leading strand in wild-type DNA polymerase yeast cells. The leading/lagging-strand ratio of rNMP incorporation changes dramatically within 500 nt from ARS’s, highlighting the Pol δ - Pol ε handoff during early leading-strand synthesis. Furthermore, the pattern of rNMP incorporation is markedly distinct between the leading the lagging strand. Overall, our results show the different counts and patterns of rNMP incorporation during DNA replication from ARS, which reflects the different labor of division and rNMP incorporation pattern of Pol δ and Pol ε.
Ecological fitness cost associated with the AHAS Trp574Leu mutation in feralRaphanus sativus
Authors: Vercellino Roman B. , Hernández Fernando , Pandolfo Claudio E. , Cantamutto Miguel , Presotto Alejandro
Published: August 2020

Gene mutations endowing herbicide resistance may have negative pleiotropic effects on plant fitness. Quantifying these effects is critical for predicting the evolution of herbicide resistance and developing management strategies for herbicide-resistant weeds. This study reports the effects of the acetohydroxyacid synthase (AHAS) Trp574Leu mutation throughout the life cycle of the weed feral radish (Raphanus sativusL.). Resistant and susceptible biotypes responded differently to light and water treatments in relation to germination and emergence. Under light exposure, the resistant biotype showed higher germination and emergence, but no differences were found in seed dormancy, germination in darkness and emergence from buried seeds or pods. The resistant biotype showed delayed and reduced seedling emergence relative to the susceptible biotype under rainfed conditions, but these differences between the biotypes were not detected in irrigated soil. The biotypes showed similar relative growth rates and vegetative biomass. However, under wheat interference, resistant plants had 36–46% less total above-ground biomass, 26–47% less seeds per plant, and 36–53% less plant yield than susceptible ones, and these differences were more evident at higher plant density. This study provides a better understanding of the ecological fitness cost associated with the AHAS Trp574Leu mutation in feralR. sativus. The fitness costs could reduce the frequency of the resistant allele in areas untreated with AHAS inhibiting herbicides.
The Pharmacodynamic-Toxicodynamic Relationship of AUC and CMAX in Vancomycin Induced Kidney Injury in an Animal Model
Authors: Avedissian Sean N. , Pais Gwendolyn , Liu Jiajun , O’Donnell J. Nicholas , Lodise Thomas P. , Neely Michael , Prozialeck Walter C. , Lamar Peter C. , Becher Leighton , Scheetz Marc H.
Published: August 2020

Background

Vancomycin induces exposure-related acute kidney injury. However, the pharmacokinetic-toxicodynamic (PK-TD) relationship remains unclear.

Methods

Sprague-Dawley rats received IV vancomycin doses of 300mg/kg/day and 400mg/kg/day, divided once, twice, thrice or 4xdaily (i.e., QD, BID, TID or QID) over 24-hours. Up to 8-samples were drawn during the 24-hour dosing period. Twenty-four-hour urine was collected and assayed for kidney injury molecule-1 (KIM-1). Vancomycin was quantified via LC-MS/MS. Following terminal sampling, nephrectomy and histopathologic analyses were conducted. PK analyses were conducted using Pmetrics. PK exposures (i.e. AUC0-24h, CMAX0-24h,) were calculated for each rat, and PK-TD relationships were discerned.

Results

A total of 53-rats generated PK-TD data. A 2-compartment model fit the data well (Bayesian observed vs. predicted concentrations, R2=0.96). KIM-1 values were greater in QD and BID groups (P-values: QD vs TID:<0.002, QD vs QID:<0.004, BID vs TID:<0.002, and BID vs QID:<0.004). Exposure–response relationships were observed between KIM-1 vs CMAX0–24hand AUC0-24h(R2□=□ 0.7 and 0.68). Corrected Akaike’s information criterion showed CMAX0-24has most predictive PK-TD driver for vancomycin-induced kidney injury (VIKI) (−5.28 versus −1.95).

Conclusions

While PK-TD indices are often inter-correlated, maximal concentrations and fewer doses (for the same total daily amount) resulted in increased VIKI in our rat model.
Elucidation of remdesivir cytotoxicity pathways through genome-wide CRISPR-Cas9 screening and transcriptomics
Authors: Akinci Ersin , Cha Minsun , Lin Lin , Yeo Grace , Hamilton Marisa C. , Donahue Callie J. , Bermudez-Cabrera Heysol C. , Zanetti Larissa C. , Chen Maggie , Barkal Sammy A. , Khowpinitchai Benyapa , Chu Nam , Velimirovic Minja , Jodhani Rikita , Fife James D. , Sovrovic Miha , Cole Philip A. , Davey Robert A. , Cassa Christopher A. , Sherwood Richard I.
Published: August 2020

The adenosine analogue remdesivir has emerged as a frontline antiviral treatment for SARS-CoV-2, with preliminary evidence that it reduces the duration and severity of illness1. Prior clinical studies have identified adverse events1,2, and remdesivir has been shown to inhibit mitochondrial RNA polymerase in biochemical experiments7, yet little is known about the specific genetic pathways involved in cellular remdesivir metabolism and cytotoxicity. Through genome-wide CRISPR-Cas9 screening and RNA sequencing, we show that remdesivir treatment leads to a repression of mitochondrial respiratory activity, and we identify five genes whose loss significantly reduces remdesivir cytotoxicity. In particular, we show that loss of the mitochondrial nucleoside transporterSLC29A3mitigates remdesivir toxicity without a commensurate decrease in SARS-CoV-2 antiviral potency and that the mitochondrial adenylate kinaseAK2is a remdesivir kinase required for remdesivir efficacy and toxicity. This work elucidates the cellular mechanisms of remdesivir metabolism and provides a candidate gene target to reduce remdesivir cytotoxicity.
Development and cell cycle dynamics of the root apical meristem in the fernCeratopteris richardii
Authors: Aragón-Raygoza Alejandro , Vasco Alejandra , Blilou Ikram , Herrera-Estrella Luis , Cruz-Ramírez Alfredo
Published: August 2020

Ferns are a representative clade in plant evolution although underestimated in the genomic era.Ceratopteris richardiiis an emergent model for developmental processes in ferns, yet a complete scheme of the different growth stages is necessary. Here, we present a developmental analysis, at the tissue and cellular levels, of the first shoot-borne root of Ceratopteris. We followed early stages and emergence of the root meristem in sporelings. While assessing root growth, the first shoot-borne root ceases its elongation between the emergence of the fifth and sixth roots, suggesting Ceratopteris roots follow a determinate developmental program. We report cell division frequencies in the stem cell niche after detecting labeled nuclei in the root apical cell (RAC) and derivatives after 8 hours of exposure. These results demonstrate the RAC has a continuous mitotic activity during root development. Detection of cell cycle activity in the RAC at early times suggests this cell acts as a non-quiescent organizing center. Overall, our results provide a framework to study root function and development in ferns and to better understand the evolutionary history of this organ.

Summary Statement

In the Ceratopteris root, the apical cell and its derivatives have a high division frequency, suggesting the apical cell acts as a non-quiescent organizing center in the stem cell niche.
A DNA nanoscope that identifies and precisely localizes over a hundred unique molecular features with nanometer accuracy
Authors: Gopalkrishnan Nikhil , Punthambaker Sukanya , Schaus Thomas E. , Church George M. , Yin Peng
Published: August 2020

Techniques that can both spatially map out molecular features and discriminate many targets would be highly valued for their utility in studying fundamental nanoscale processes. In spite of decades of development, no current technique can achieve both nanoscale resolution and discriminate hundreds of targets. Here, we report the development of a novel bottom-up technology that: (a) labels a sample with DNA barcodes, (b) measures pairwise-distances between labeled sites and writes them into DNA molecules, (c) reads the pairwise-distances by sequencing and (d) robustly integrates this noisy information to reveal the geometry of the underlying sample. We demonstrate our technology on DNA origami, which are complex synthetic nanostructures. We both spatially localized and uniquely identified over a hundred densely packed unique elements, some spaced just 6 nm apart, with an average spatial localization accuracy (RMS deviation) of ~2 nm. The bottom-up, sequencing-enabled mechanism of the DNA nanoscope is fundamentally different from top-down imaging, and hence offers unique advantages in precision, throughput and accessibility.
BNIP3 phosphorylation by JNK1/2 promotes mitophagy via enhancing its stability under hypoxia
Authors: He Yun-Ling , Gong Sheng-Hui , Cheng Xiang , Zhao Ming , Zhao Tong , Zhao Yong-Qi , Fan Ming , Zhu Ling-Ling , Wu Li-Ying
Published: August 2020

Mitophagy is an important metabolic mechanism that modulates mitochondrial quality and quantity by selectively removing damaged or unwanted mitochondria. BNIP3, a mitochondrial outer membrane protein, is a mitophagy receptor that mediates mitophagy under various stresses, particularly hypoxia, since BNIP3 is a hypoxia-responsive protein. However, the underlying mechanisms that regulate BNIP3 and thus mediate mitophagy under hypoxic conditions remain elusive. Here, we demonstrate that in hypoxia JNK1/2 phosphorylates BNIP3 at Ser 60/Thr 66, which hampers proteasomal degradation of BNIP3 and drives mitophagy by facilitating the direct binding of BNIP3 to LC3, while PP1/2A represses mitophagy by dephosphorylating BNIP3 and triggering its proteasomal degradation. These findings reveal the intrinsic mechanisms cells use to regulate mitophagy via the JNK1/2-BNIP3 pathway in response to hypoxia. Thus, the JNK1/2-BNIP3 signaling pathway strongly links mitophagy to hypoxia and may be a promising therapeutic target for hypoxia-related diseases.
A subset of pediatric thalamic gliomas share a distinct DNA methylation profile, H3K27me3 loss and frequent alteration ofEGFR
Authors: Sievers Philipp , Sill Martin , Schrimpf Daniel , Stichel Damian , Reuss David E. , Sturm Dominik , Hench Jürgen , Frank Stephan , Krskova Lenka , Vicha Ales , Zapotocky Michal , Bison Brigitte , Harter Patrick N. , Snuderl Matija , Kramm Christof M. , Reifenberger Guido , Korshunov Andrey , Jabado Nada , Wesseling Pieter , Wick Wolfgang , Solomon David A. , Perry Arie , Jacques Thomas S. , Jones Chris , Witt Olaf , Pfister Stefan M. , von Deimling Andreas , Jones David T. W. , Sahm Felix
Published: August 2020

Background

Malignant astrocytic gliomas in children show a remarkable biological and clinical diversity. Small in-frame insertions or missense mutations in theEGFRgene have recently been identified in a distinct subset of pediatric bithalamic gliomas with a unique DNA methylation pattern.

Methods

Here, we investigated an epigenetically homogeneous cohort of malignant gliomas (n=58) distinct from other subtypes and enriched for pediatric cases and thalamic location, in order to elucidate the overlap with this recently identified subtype of pediatric bithalamic gliomas.

Results

EGFRgene amplification was detected in 16/58 (27%) tumors, and missense mutations or small in-frame insertions inEGFRwere found in 20/30 tumors with available sequencing data (67%; five of them co-occurring withEGFRamplification). Additionally, eight of the 30 tumors (27%) harbored an H3.1 or H3.3 K27M mutation (six of them with a concomitantEGFRalteration). All tumors tested showed loss of H3K27me3 staining, with evidence ofEZHIPoverexpression in the H3 wildtype cases. Although some tumors indeed showed a bithalamic growth pattern, a significant proportion of tumors occurred in the unilateral thalamus or in other (predominantly midline) locations.

Conclusions

Our findings present a distinct molecular class of pediatric malignant gliomas largely overlapping with the recently reported bithalamic gliomas characterized byEGFRalteration, but additionally showing a broader spectrum ofEGFRalterations and tumor localization. Global H3K27me3 loss in this group appears to be mediated by either H3 K27 mutation orEZHIPoverexpression. EGFR inhibition may represent a potential therapeutic strategy in these highly aggressive gliomas.

Key points

This study confirms a distinct new subset of pediatric diffuse midline glioma with H3K27me3 loss, with or without H3 K27 mutation

The poor outcome of these tumors is in line with the broader family of pediatric diffuse midline gliomas with H3 K27 mutation orEZHIPoverexpression

FrequentEGFRalterations in these tumors may represent a therapeutic target in this subset

Importance of the Study

Malignant astrocytic gliomas in children show a remarkable biological and clinical diversity. Here, we highlight a distinct molecular class of pediatric malignant gliomas characterized byEGFRalteration and global H3K27me3 loss that appears to be mediated by either H3 K27 mutation orEZHIPoverexpression. EGFR inhibition may represent a potential therapeutic strategy in these highly aggressive gliomas.
Microbiota Accelerates Age-Related CpG Island Methylation in Colonic Mucosa
Authors: Sun Ang , Cole Lauren , Maegawa Shinji , Park Pyoung Hwa , Keith Kelsey , Madzo Jozef , Jelinek Jaroslav , Jobin Christian , Issa Jean-Pierre J.
Published: August 2020

DNA methylation is an epigenetic mark that is altered in cancer and aging tissues. The effects of extrinsic factors on DNA methylation remain incompletely understood. Microbial dysbiosis is a hallmark of colorectal cancer, and infections have been linked to aberrant DNA methylation in cancers of the GI tract. To determine the microbiota’s impact on DNA methylation, we studied the methylomes of colorectal mucosa in germ-free (no microbiota) and specific-pathogen-free (controlled microbiota) mice, as well as in Il-10 KO mice (Il10−/−) which are prone to inflammation and tumorigenesis in the presence of microbiota. The presence of microbiota was associated with changes in 5% of the methylome andIl10−/−mice showed alterations in 4.1% of the methylome. These changes were slightly more often hypo than hypermethylation and affected preferentially CpG sites located in gene bodies and intergenic regions. Mice with both Il-10 KO and microbiota showed much more pronounced alterations, affecting 18% of the methylome. When looking specifically at CpG island methylation alterations, a hallmark of aging and cancer, 0.4% were changed by the microbiota, 0.4% were changed byIl10−/−, while 4% were changed by both simultaneously. These effects are comparable to what is typically seen when comparing colon cancer to normal. We next compared these methylation changes to those seen in aging, and after exposure to the colon carcinogen Azoxymethane (AOM). Aging was associated with alterations in 18% of the methylome, and aging changes were accelerated in theIl10−/−/SPF mice. By contrast, AOM induced profound hypomethylation that was distinct from the effects of aging or of the microbiota. CpG sites modified by the microbiota were over-represented among DNA methylation changes in colorectal cancer. Thus, the microbiota affects the DNA methylome of colorectal mucosa in patterns reminiscent of what is observed in aging and in colorectal cancer.
Nanoencapsulation of clove oil and study of physico-chemical properties, cytotoxic, haemolytic and antioxidant activities
Authors: Nagaraju Pramod G , Sengupta Parineeta , Poornima Priyadarshini C. G. , Rao Pooja J
Published: August 2020

The therapeutic properties of clove oil is known for centuries, however, the pungent nature, chemical instability and low water solubility impose limitations in harnessing its therapeutic potential. Hence, nanoencapsulation of clove oil was performed to overcome the above constraints and control its in-vitro release. The stability of nanoemulsion depends on various factors where the surfactant and its hydrophile/lipofile balance (HLB) play a key role. The non-ionic surfactants Tween 20, 40 and 80 with HLB of 16.7, 15.6 and 15, respectively, were used to study the stability of clove oil nanoemulsion (CON). The creaming index of CON prepared with Tween 20, 40 and 80 was 22.75 and 17.5 and 1.5%, respectively, after 8 days of storage at room temperature. Tween 20 and 40 produced particles > 300 nm while Tween 80 resulted in particles of size ∼150 nm. Transmission electron microscopic image of spray dried CON prepared with Tween 80 showed particle size in the range 150-190 nm after one month of storage at room temperature. The in vitro release studies showed 76% and 42% cumulative release of CON and native clove oil (NC), respectively at pH 7.4. The cellular toxicity of CON was significantly reduced by four fold compared to NC at a concentration of 60 µg/mL when tested on Caco2 cells. Similarly, haemolytic activity on red blood cells revealed less than 10% haemolysis signifying the compatibility of CON for its nutraceutical applications. In addition, CON also exhibited higher in-vitro antioxidant compared to NC as shown by DPPH and ABTS radical scavenging activity. Collectively, we have developed a unique method for NC nanoencapsulation using cost effective polysaccharide (maltodextrin) and surfactant for stabilizing the nanoemulsion for increased bioactivity.
De NovoMutation in an Enhancer ofEBF3in simplex autism
Authors: Padhi Evin M. , Hayeck Tristan J. , Mannion Brandon , Chatterjee Sumantra , Byrska-Bishop Marta , Musunuri Rajeeva , Narzisi Giuseppe , Abhyankar Avinash , Cheng Zhang , Hunter Riana D. , Akiyama Jennifer , Fries Lauren E. , Ng Jeffrey , Stong Nick , Allen Andrew S. , Dickel Diane E. , Bernier Raphael A. , Gorkin David U. , Pennacchio Len A. , Zody Michael C. , Turner Tychele N.
Published: August 2020

Previous research in autism and other neurodevelopmental disorders (NDDs) has indicated an important contribution ofde novoprotein-coding variants within specific genes. The role ofde novononcoding variation has been observable as a general increase in genetic burden but has yet to be resolved to individual functional elements. In this study, we assessed whole-genome sequencing data in 2,671 families with autism, with a specific focus onde novovariation in enhancers with previously characterizedin vivoactivity. We identified three independentde novomutations limited to individuals with autism in the enhancer hs737. These mutations result in similar phenotypic characteristics, affect enhancer activityin vitro, and preferentially occur in AAT motifs in the enhancer with predicted disruptions of transcription factor binding. We also find that hs737 is enriched for copy number variation in individuals with NDDs, is dosage sensitive in the human population, is brain-specific, and targets the NDD geneEBF3that is genome-wide significant for protein codingde novovariants, demonstrating the importance of understanding all forms of variation in the genome.

One Sentence Summary

Whole-genome sequencing in thousands of families reveals variants relevant to simplex autism in a brain enhancer of the well-established neurodevelopmental disorder geneEBF3.
Ion-water coupling controls class A GPCR signal transduction pathways
Authors: Thomson Neil J. , Vickery Owen N. , Ives Callum M. , Zachariae Ulrich
Published: August 2020

G-protein-coupled receptors (GPCRs) transmit signals across the cell membrane, forming the largest family of membrane proteins in humans. Most GPCRs activate through an evolutionarily conserved mechanism, which involves reorientation of helices and key residues, rearrangement of a hydrogen bonding network mediated by water molecules, and the expulsion of a sodium ion from a protonatable binding site. However, how these components interplay to engage the signal effector binding site remains elusive. Here, we applied information theory to molecular dynamics simulations of pharmaceutically important GPCRs to trace concerted conformational variations across the receptors. We discovered a conserved communication pathway that includes protein residues and cofactors and enables the exchange of information between the extracellular sodium binding site and the intracellular G-protein binding region, coupling the most highly conserved protonatable residues at long distance. Reorientation of internal water molecules was found to be essential for signal transmission along this pathway. By inhibiting protonation, sodium decoupled this connectivity, identifying the ion as a master switch that determines the receptors’ ability to move towards active conformations.
Comprehensive benchmarking of software for mapping whole genome bisulfite data: from read alignment to DNA methylation analysis
Authors: Nunn Adam , Otto Christian , Stadler Peter F. , Langenberger David
Published: August 2020

Whole genome bisulfite sequencing is currently at the forefront of epigenetic analysis, facilitating the nucleotide-level resolution of 5-methylcytosine (5mC) on a genome-wide scale. Specialised software have been developed to accommodate the unique difficulties in aligning such sequencing reads to a given reference, building on the knowledge acquired from model organisms such as human, orArabidopsis thaliana. As the field of epigenetics expands its purview to non-model plant species, new challenges arise which bring into question the suitability of previously established tools.

Herein, nine short-read aligners are evaluated:Bismark,BS-Seeker2,BSMAP,BWA-meth,ERNE-BS5,GEM3,GSNAP,Last, andsegemehl. Precision-recall of simulated alignments, in comparison to real sequencing data obtained from three natural accessions, reveals on-balance thatBWA-methandBSMAPare able to make the best use of the data during mapping. The influence of difficult-to-map regions, characterised by deviations in sequencing depth over repeat annotations, is evaluated in terms of the mean absolute deviation of the resulting methylation calls in comparison to a realistic methylome. Downstream methylation analysis is responsive to the handling of multi-mapping reads relative to mapping quality (MAPQ), and potentially susceptible to bias arising from the increased sequence complexity of densely-methylated reads.
Synaptic vesicle fusion along axons is driven by myelination and subsequently accelerates sheath growth in an activity-regulated manner
Authors: Almeida Rafael G , Williamson Jill M , Madden Megan E , Early Jason J , Voas Matthew G , Talbot William S , Bianco Isaac H , Lyons David A
Published: August 2020

To study activity-regulated myelination, we imaged synaptic vesicle fusion along single axons in living zebrafish, and found, surprisingly, that axonal synaptic vesicle fusion is driven by myelination. This myelin-induced axonal vesicle fusion was enriched along the unmyelinated domains into which newly-formed sheaths grew, and was promoted by neuronal activity, which in turn accelerated sheath growth. Our results indicate that neuronal activity consolidates sheath growth along axons already selected for myelination.
Kinesin-4 KIF21B limits microtubule growth to allow rapid centrosome polarization in T cells
Authors: Hooikaas Peter Jan , Damstra Hugo G.J. , Gros Oane J. , van Riel Wilhelmina E. , Martin Maud , Smits Yesper T.H. , van Loosdregt Jorg , Kapitein Lukas C. , Berger Florian , Akhmanova Anna
Published: August 2020

When a T cell and an antigen-presenting cell form an immunological synapse, rapid dynein-driven translocation of the centrosome towards the contact site leads to reorganization of microtubules and associated organelles. Currently, little is known about how the regulation of microtubule dynamics contributes to this process. Here, we show that the knockout of KIF21B, a kinesin-4 linked to autoimmune disorders, causes microtubule overgrowth and perturbs centrosome translocation. KIF21B restricts microtubule length by inducing microtubule pausing typically followed by catastrophe. Catastrophe induction with vinblastine prevented microtubule overgrowth and was sufficient to rescue centrosome polarization in KIF21B-knockout cells. Biophysical simulations showed that a relatively small number of KIF21B molecules can restrict microtubule length and promote an imbalance of dynein-mediated pulling forces that allows the centrosome to translocate past the nucleus. We conclude that proper control of microtubule length is important for allowing rapid remodeling of the cytoskeleton and efficient T cell polarization.
Hericerin derivatives fromHericium erinaceusexert BDNF-like neurotrophic activity in central hippocampal neurons and enhance memory
Authors: Martínez-Mármol Ramón , Chai YeJin , Khan Zahra , Kim Seon Beom , Hong Seong Min , Gormal Rachel S. , Lee Dae Hee , Lee Jae Kang , Lee Mi Kyeong , Kim Sun Yeou , Meunier Frédéric A.
Published: August 2020

The traditional medicinal mushroomHericium erinaceushas long been known for enhancing the peripheral nerve regeneration through targeting nerve growth factor (NGF) neurotrophic activity. It was also reported to protect against ageing-dependent cognitive decline in wildtype and in Alzheimer’s disease mouse models suggesting a yet to be defined action on neurons of the central nervous system. Here, we purified and identified biologically active compounds fromH. erinaceus, based on their ability to promote neurite outgrowth in hippocampal neurons.N-de phenylethyl isohericerin (NDPIH), an isoindoline compound from this mushroom together with its hydrophobic derivative hericene A, were highly potent in inducing extensive axon outgrowth and neurite branching in the absence of serum demonstrating high neurotropic activity. NDPIH also induced enlarged growth cones suggestive of a brain-derived neurotrophic factor (BDNF)-like activity. Pharmacological inhibition of tropomyosin receptor kinase B (TrkB) by ANA12 prevented NDPIH-induced neurotrophic activity providing evidence that NDPIH actsviaTrkB receptors to mediate its neurotrophic effect in central neurons. Finally,in vivotreatment withH. erinaceuscrude extract and hericene A significantly increased BDNF and downstream pathway and enhanced learning and memory in the novel object recognition memory test. Our results suggest that hericene A can promote BDNF-like activity in neuronsin vitroandin vivothereby enhancing recognition memory.
Tropism of SARS-CoV-2 in commonly used laboratory cell lines and their proteomic landscape during infection
Authors: Saccon Elisa , Chen Xi , Mikaeloff Flora , Rodriguez Jimmy Esneider , Szekely Laszlo , Sá Vinhas Beatriz , Krishnan Shuba , Byrareddy Siddappa N. , Frisan Teresa , Végvári Ákos , Mirazimi Ali , Neogi Ujjwal , Gupta Soham
Published: August 2020

The present pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is driving intense research activities to understand the basic biology of the virus and determine effective therapeutic strategies. The commonly used laboratory cell lines of human origin are the first line of experimental models to study the pathogenicity and performing antiviral assays. Thus, to find suitable cell models to study SARS-CoV-2, we assessed the tropism and cytopathogenicity of the first Swedish isolate of SARS-CoV-2 in six different cell lines of human origin and compared their growth characteristics to other globally isolated strains. Overall, Calu-3, Caco2, Huh7, and 293FT cell lines showed a high to moderate level of susceptibility to the majority of virus isolates. In Caco2 cells the virus can achieve high titers in the absence of any prominent cytopathic effect. The protein expression profile during SARS-CoV-2 infection revealed cell-type-specific regulation of cellular pathways. Type-I interferon signaling was identified as the common dysregulated cellular response in Caco2, Calu-3 and Huh7 cells. Overall, cell-type specific variability was noted for cytopathogenicity, susceptibility and cellular response to SARS-CoV-2. This study provides important clues regarding SARS-CoV-2 pathogenesis and can represent as a guide for future studies to design therapeutics.
Structural Basis for Allosteric Control of the SERCA-Phospholamban Membrane Complex by Ca2+and cAMP-dependent Phosphorylation
Authors: Weber Daniel K. , Sanz-Hernández Máximo , Reddy U. Venkateswara , Wang Songlin , Larsen Erik K. , Gopinath Tata , Gustavsson Martin , Cornea Razvan L. , Thomas David D. , De Simone Alfonso , Veglia Gianluigi
Published: August 2020

Phospholamban (PLN) is a mini-membrane protein that directly controls the cardiac Ca2+-transport response to β-adrenergic stimulation, thus modulating cardiac output during the fight- or-flight response. In the sarcoplasmic reticulum membrane, PLN binds to the sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), keeping this enzyme’s function within a narrow physiological window. PLN phosphorylation by cAMP-dependent protein kinase A or increase in Ca2+concentration reverses the inhibitory effects through an unknown mechanism. Using oriented-sample solid-state NMR spectroscopy and replica-averaged NMR-restrained structural refinement, we reveal that phosphorylation of PLN’s cytoplasmic regulatory domain signals the disruption of several inhibitory contacts at the transmembrane binding interface of the SERCA-PLN complex that are propagated to the enzyme’s active site, augmenting Ca2+transport. Our findings address long-standing questions about SERCA regulation, epitomizing a signal transduction mechanism operated by posttranslationally-modified bitopic membrane proteins.
Thermodynamic and mechanistic insights into coupled binding and unwinding of collagen by matrix metalloproteinase 1
Authors: Manka Szymon W. , Brew Keith
Published: August 2020

Local unwinding of the collagen triple helix is a necessary step for initiating the collagen degradation cascade in extracellular matrices. A few matrix metalloproteinases (MMPs) are known to support this key process, but its energetic aspects remain unknown. Here, we captured the thermodynamics of the triple helix unwinding by monitoring interactions between a collagen peptide and MMP-1(E200A) – an active-site mutant of an archetypal vertebrate collagenase – at increasing temperatures, using isothermal titration calorimetry (ITC). Coupled binding and unwinding manifests as a curved relationship between the total enthalpy change and temperature of the reaction, producing increasingly negative heat capacity change (ΔΔCp≈ −36.3 kcal/molK2). A specially designed solid-phase binding and cleavage assay (SPBCA) reported strain in the catalytically relevant unwound state, suggesting that this state is distinct from the horizon of sampled conformations of the collagenase-susceptible site. MMP-1 appears to blend selected fit with induced fit mechanisms to catalyse collagen unwinding prior to cleavage of individual collagen chains.

Graphical Abstract

Highlights

The unwinding of the collagen triple helix by matrix metalloproteinases (MMPs) is critical for collagen catabolism, but how MMPs harness bioavailable energy for this energetically expensive process is enigmatic

Rising temperature causes linear increases of the negative heat capacity change associated with the interaction of MMP-1 with a collagenase-susceptible triple-helical peptide, indicating enrichment of hydrophobic contacts during the triple helix unwinding

The complex of MMP-1 with locally unwound collagen holds considerable structural strain, which becomes relieved upon cleavage of the unwound collagen strand(s)

MMP-1 consumes heat to drive specific (structurally distinct) collagen unwinding by increasing entropy associated with evolving hydrophobic anchor points between the enzyme and the substrate

The prototypic collagenase MMP-1 appears to blend conformational selection with induced fit, shedding light on the thermodynamic principles by which MMPs trigger collagen breakdown and supporting general mechanistic conclusions concerning conformational changes coupled to protein-protein binding
Genome-wide transcript and protein analysis reveals distinct features of aging in the mouse heart
Authors: Gyuricza Isabela Gerdes , Chick Joel M. , Keele Gregory R. , Deighan Andrew G. , Munger Steven C. , Korstanje Ron , Gygi Steven P. , Churchill Gary A.
Published: August 2020

Investigation of the molecular mechanisms of aging in the human heart is challenging due to confounding factors, such as diet and medications, as well limited access to tissues. The laboratory mouse provides an ideal model to study aging in healthy individuals in a controlled environment. However, previous mouse studies have examined only a narrow range of the genetic variation that shapes individual differences during aging. Here, we analyzed transcriptome and proteome data from hearts of genetically diverse mice at ages 6, 12 and 18 months to characterize molecular changes that occur in the aging heart. Transcripts and proteins reveal distinct biological processes that are altered through the course of natural aging. Transcriptome analysis reveals a scenario of cardiac hypertrophy, fibrosis, and reemergence of fetal gene expression patterns. Proteome analysis reveals changes in energy metabolism and protein homeostasis. We found that for many protein complexes there is a decline in correlation between their component proteins with age, indicating age-related loss of stoichiometry. Some of the most affected complexes are themselves involved in protein homeostasis, which potentially contributes to a viscious cycle of progressive breakdown in protein quality control with age. In addition, we identified genetic loci that modulate age-related changes in a variety of cellular processes, including protein degradation and sorting, suggesting that genetic variation can alter the rate of molecular aging.
A Liquid-to-Solid Phase Transition Enhances the Catalytic Activity of SARM1
Authors: Loring Heather S. , Thompson Paul R.
Published: August 2020

Sterile alpha and toll/interleukin receptor (TIR) motif–containing protein 1 (SARM1) is a neuronally expressed NAD+glycohydrolase whose activity is increased in response to various stressors. The consequent depletion of NAD+triggers axonal degeneration (i.e., Wallerian degeneration), which is a characteristic feature of neurological diseases, including peripheral neuropathies and traumatic brain injury. Notably, SARM1 knockout mice show minimal degeneration in models of peripheral neuropathy and traumatic brain injury, making SARM1 a promising therapeutic target. However, the development of SARM1 inhibitors has been challenging as the purified enzyme is largely inactive. Herein, we report that SARM1 activity is increased ∼2000–fold by a liquid-to-solid phase transition. These findings provide critical insights into SARM1 biochemistry with important implications for the situationin vivo. Moreover, they will facilitate the discovery of novel SARM1–targeted therapeutics.

Graphical Abstract
HIV-specific T-cell responses reflect substantive in vivo interactions with infected cells despite long-term therapy
Authors: Stevenson Eva M. , Ward Adam R. , Truong Ronald , Thomas Allison S. , Huang Szu-Han , Dilling Thomas R. , Terry Sandra , Bui John K. , Mota Talia M. , Danesh Ali , Lee Guinevere Q. , Gramatica Andrea , Khadka Pragya , Alberto Winiffer D. Conce , Gandhi Rajesh T. , McMahon Deborah K. , Lalama Christina M. , Bosch Ronald J. , Macatangay Bernard , Cyktor Joshua C. , Eron Joseph J. , Mellors John W. , Jones R. Brad ,   for the ACTG A5321 Team
Published: August 2020

Antiretroviral therapies (ART) durably suppress HIV replication to undetectable levels – however, infection persists in the form of long-lived reservoirs of infected cells with integrated proviruses, that re-seed systemic replication if ART is interrupted. A central tenet of our current understanding of this persistence is that infected cells are shielded from immune recognition and elimination through a lack of antigen expression from proviruses. Efforts to cure HIV infection have therefore focused on reactivating latent proviruses to enable immune-mediated clearance, but these have yet to succeed in driving reductions in viral reservoirs. Here, we revisited the question of whether HIV reservoirs are predominately immunologically silent from a new angle, by querying the dynamics of HIV-specific T-cell responses over long-term ART for evidence of ongoing recognition of HIV-infected cells. We show that T-cell responses to autologous reservoir viruses persist over years, and that the maintenance of HIV-Nef-specific responses was uniquely associated with residual frequencies of infected cells. These responses disproportionately exhibited a cytotoxic, effector functional profile, indicative of recentin vivorecognition of HIV-infected cells. These results indicate substantial visibility of the HIV reservoir to T-cells on stable ART, presenting both opportunities and challenges for the development of therapeutic approaches to curing HIV infection.