bluuebunny/crossref_metadata_2025 huggingface dataset:
Data driven phenotyping and COVID-19 case definitions: a pattern recognition approach
Authors: Vavougios George D. , Konstantatos Christoforos , Sinigalias Pavlos-Christoforos , Zarogiannis Sotirios G. , Kolomvatsos Konstantinos , Stamoulis George , Gourgoulianis Konstantinos I.
Published: May 2021

Introduction

COVID-19 has pathological pulmonary as well as several extrapulmonary manifestations and thus many different symptoms may arise in patients. The aim of our study was to determine COVID-19 syndromic phenotypes in a data driven manner using survey results extracted from Carnegie Mellon University’s Delphi Group.

Methods

Monthly survey results (>1 million responders per month; 320.326 responders with positive COVID-19 test and disease duration <30 days were included in this study) were used sequentially in identifying and validating COVID-19 syndromic phenotypes. Logistic Regression Weighted Multiple Correspondence Analysis (LRW-MCA) was used as a preprocessing procedure, in order to weight and transform symptoms recorded by the survey to eigenspace coordinates (i.e. object scores per case / dimension), with a goal of capturing a total variance of > 75%. These scores along with symptom duration were subsequently used by the Two Step Clustering algorithm to produce symptom clusters. Post-hoc logistic regression models adjusting for age, gender and comorbidities and confirmatory linear principal components analyses were used to further explore the data. The model created from 66.165 included responders in August, was subsequently validated in data from March – December 2020.

Results

Five validated COVID-19 syndromes were identified in August: 1. Afebrile (0%), Non-Coughing (0%), Oligosymptomatic (ANCOS) 2. Febrile (100%) Multisymptomatic (FMS) 3. Afebrile (0%) Coughing (100%) Oligosymptomatic (ACOS), 4. Oligosymptomatic with additional self-described symptoms (100%; OSDS) and 5. Olfaction / Gustatory Impairment Predominant (100%; OGIP).

Discussion

We present 5 distinct symptom phenotypes within the COVID-19 spectrum that remain stable within 9 – 12 days of first symptom onset. The typical febrile respiratory phenotype is presented as a minority among identified syndromes, a finding that may impact both epidemiological surveillance norms and transmission dynamics.
Age-dependent association between SARS-CoV-2 cases reported by passive surveillance and viral load in wastewater
Authors: Omori Ryosuke , Miura Fuminari , Kitajima Masaaki
Published: May 2021

The true number of individuals infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is difficult to estimate using a case-reporting system (i.e., passive surveillance) alone because of asymptomatic infection. While wastewater-based epidemiology has been implemented as an alternative/additional monitoring tool to reduce reporting bias, the relationship between passive and wastewater surveillance data has yet to be explicitly examined. Since there is strong age dependency in the symptomatic ratio of SARS-CoV-2 infections, this study aimed to estimate i) an age-dependent association between the number of reported cases and the viral load in wastewater and ii) the time lag between those time series. The viral load in wastewater was modeled as a combination of contributions from different age groups’ virus shedding, incorporating the delay, and fitted with daily case count data collected from the Massachusetts Department of Public Health and SARS-CoV-2 RNA concentrations in wastewater collected by the Massachusetts Water Resources Authority. The estimated lag between the time series of viral loads in wastewater and of reported cases was 10.8 days (95% confidence interval =[10.2, 11.6]) for wastewater treatment plant’s northern area and 8.8 days [8.4, 9.1] for southern area. The estimated contribution rate of a reported case to the viral load in wastewater in the 0–19 yr age group was 0.38 [0.35, 0.41] for northern area and 0.40 [0.37, 0.43] for southern area, that in the 80+ yr age group was 0.67 [0.65, 0.69] for northern area and 0.51 [0.49, 0.52] for southern area. The estimated lag between those time series suggested the predictability of reported cases ten days later using viral loads in wastewater. The contribution of a reported case in passive surveillance to the viral load in wastewater differed by age, suggesting a large variation in viral shedding kinetics among age groups.
N440K variant of SARS-CoV-2 has Higher Infectious Fitness
Authors: Tandel Dixit , Gupta Divya , Sah Vishal , Harinivas Harshan Krishnan
Published: May 2021

Several variants of SARS-CoV-2 have been emerging across the globe, continuing to threaten the efforts to end COVID-19 pandemic. Recent data indicate the prevalence of variants with N440K Spike substitution in several parts of India, which is under the second wave of the pandemic. Here, we first analyze the prevalence of N440K variants within the sequences submitted from India and identify a rising trend of its spread across various clusters. We then compare the replicative fitness and infectivity of a prototype of this variant with two other previously prevalent strains. The N440K variant produced ten times higher infectious viral titers than a prevalent A2a strain, and over 1000 folds higher titers than a much less prevalent A3i strain prototype in Caco2 cells. Similar results were detected in Calu-3 cells as well, confirming the increased potency of the N440K variant. Interestingly, A3i strain showed the highest viral RNA levels, but the lowest infectious titers in the culture supernatants, indicating the absence of correlation between the RNA content and the infectivity of the sample. N440K mutation has been reported in several viral sequences across India and based on our results, we predict that the higher infectious titers achieved by N440K variant could possibly lead to its higher rate of transmission. Availability of more sequencing data in the immediate future would help understand the potential spread of this variant in more detail.
Induced neural progenitor cells and iPS-neurons from major depressive disorder patients show altered bioenergetics and electrophysiological properties
Authors: Triebelhorn Julian , Cardon Iseline , Kuffner Kerstin , Bader Stefanie , Jahner Tatjana , Meindl Katrin , Rothhammer-Hampl Tanja , Riemenschneider Markus J. , Drexler Konstantin , Berneburg Mark , Nothdurfter Caroline , Manook André , Brochhausen Christoph , Baghai Thomas C. , Hilbert Sven , Rupprecht Rainer , Milenkovic Vladimir M. , Wetzel Christian H.
Published: April 2021

The molecular pathomechanisms of major depressive disorder (MDD) are still not completely understood. Here, we follow the hypothesis, that mitochondria dysfunction which is inevitably associated with bioenergetic disbalance is a risk factor that contributes to the susceptibility of an individual to develop MDD. Thus, we investigated molecular mechanisms related to mitochondrial function in induced neuronal progenitor cells (NPCs) which were reprogrammed from fibroblasts of eight MDD patients and eight non-depressed controls. We found significantly lower maximal respiration rates, altered cytosolic basal calcium levels, and smaller soma size in NPCs derived from MDD patients. These findings are partially consistent with our earlier observations in MDD patient-derived fibroblasts. Furthermore, we differentiated MDD and control NPCs into iPS-neurons and analysed their passive biophysical and active electrophysiological properties to investigate whether neuronal function can be related to altered mitochondrial activity and bioenergetics. Interestingly, MDD patient-derived iPS-neurons showed significantly lower membrane capacitance, a less hyperpolarized membrane potential, increased Na+current density and increased spontaneous electrical activity. Our findings indicate that functional differences evident in fibroblasts derived from MDD patients are partially present after reprogramming to induced-NPCs, might relate to altered function of iPS-neurons and thus might be associated with the aetiology of major depressive disorder.
Low-bias ncRNA Libraries using Ordered Two-Template Relay: Serial Template Jumping by a Modified Retroelement Reverse Transcriptase
Authors: Upton Heather E. , Ferguson Lucas , Temoche-Diaz Morayma M. , Liu Xiaoman , Pimentel Sydney C. , Ingolia Nicholas T. , Schekman Randy , Collins Kathleen
Published: May 2021

Non-long terminal repeat (non-LTR) and group II intron retroelements encode reverse transcriptases (RTs) that copy the retroelement transcript directly into host cell DNA, often at specific target sites. Biochemical characterization of these enzymes has been limited by recombinant expression and purification challenges, hampering understanding of their transposition mechanism and their exploitation for research and biotechnology. Properties of retroelement RTs substantiate their application for end-to-end RNA sequence capture. To investigate this utility, we first compared a non-LTR RT fromBombyx moriand a group II intron RT fromEubacterium rectale. Only the non-LTR RT showed processive template jumping, producing one cDNA from discontinuous templates each copied end-to-end. We also discovered an unexpected terminal deoxynucleotidyl transferase activity of the RTs that adds nucleotide(s) of choice to 3’ ends of single-stranded RNA or DNA. Combining these two types of activity with additional insights about non-templated nucleotide additions to duplexed cDNA product, we developed a streamlined protocol for linking Next Generation Sequencing (NGS) adaptors to both cDNA ends in a single RT reaction. When benchmarked using a reference pool of microRNAs (miRNAs), library production using modified non-LTR retroelment RT for Ordered Two-Template Relay (OTTR) outperformed all commercially available kits and rivaled the low bias of technically demanding home-brew protocols. We applied OTTR to inventory RNAs purified from extracellular vesicles (EVs), identifying miRNAs as well as myriad other non-coding (nc) RNAs and ncRNA fragments. Our results establish the utility of OTTR for automation-friendly, low-bias, end-to-end RNA sequence inventories of complex ncRNA samples.

Significance

Retrotransposons are non-infectious mobile genetic elements that proliferate in host genomes via an RNA intermediate that is copied into DNA by a reverse transcriptase (RT) enzyme. RTs are important for biotechnological applications involving information capture from RNA, since RNA is first converted into complementary DNA for detection or sequencing. Here, we biochemically characterize RTs from two retroelements and uncover several activities that allowed us to design a streamlined, efficient workflow for determining the inventory of RNA sequences in processed RNA pools. The unique properties of non-retroviral RT activities obviate many technical issues associated with current methods of RNA sequence analysis, with wide applications in research, biotechnology, and diagnostics.
Serum-free medium increases the replication rate of the avian coronavirus infectious bronchitis virus in chicken embryo kidney cells
Authors: Quinteros José A. , Browning Glenn F. , Noormohammadi Amir H. , Stevenson Mark A. , Coppo Mauricio J. C. , Loncoman Carlos A. , Ficorilli Nino , Lee Sang-Won , Diaz-Méndez Andrés
Published: May 2021

Infectious bronchitis virus (IBV), an avian coronavirus, can be isolated and cultured in tracheal organ cultures (TOCs), embryonated eggs and cell cultures. TOCs and embryonated eggs are commonly used for viral isolation but use of these is laborious and expensive. Cell cultures have been used only with IBV strains that have previously been adapted to grow under laboratory conditions, and not for primary isolation. Previous studies using the coronavirus porcine epidemic diarrhoea virus (PEDV) have suggested that foetal bovine serum (FBS), a common component of cell culture media, can inhibit the adsorption of coronaviruses onto the host cell membrane receptors. In the present study, the replication of IBV in primary chicken embryo kidney (CEK) cell cultures and the Leghorn hepatocellular carcinoma (LMH) cell line was examined using two different cell culture media, one containing FBS and the other containing yeast extract (YE). A reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) assay was used to quantify viral RNA copies in cell lysates. The highest concentrations of viral genomes were observed when the cell culture medium did not contain FBS. Examination of the infectivity of virus grown in CEK cell cultures was examined by titration in embryonated chicken eggs, demonstrating that the cell lysate from CEK cell cultures in medium without FBS contained a higher median embryo infectious dose (EID50) than that from CEK cell cultures in medium containing FBS. These results suggest that improved replication of IBV in cell cultures can be achieved by the omission of FBS from the cell culture medium. This may enhance the potential for production of vaccines in cell culture and facilitate the isolation of emergent IBV strains in cell cultures.
Sex- and Subtype-Specific Adaptations in Excitatory Signaling Onto Deep-Layer Prelimbic Cortical Pyramidal Neurons After Chronic Alcohol Exposure
Authors: Hughes Benjamin A. , O’Buckley Todd K. , Boero Giorgia , Herman Melissa A. , Morrow A. Leslie
Published: May 2021

Long-term alcohol use results in behavioral deficits including impaired working memory, elevated anxiety, and blunted inhibitory control that are associated with prefrontal cortical (PFC) dysfunction. Preclinical observations demonstrate multiple impairments in GABAergic neurotransmission onto deep-layer principal cells (PCs) in prelimbic cortex that suggest dependence-related cortical dysfunction is the product of elevated excitability in these cells. Despite accumulating evidence showing alcohol-induced changes in interneuron signaling onto PCs differ between sexes, there is limited data explicitly evaluating sex-specific ethanol effects on excitatory signaling onto deep-layer PCs that may further contribute to deficits in PFC-dependent behaviors. To address this, we conducted electrophysiological and behavioral tests in both male and female Sprague-Dawley rats to evaluate the effects of chronic ethanol exposure. Among our observations, we report a marked enhancement in glutamatergic signaling onto deep-layer PCs in male, but not female, rats after alcohol exposure. This phenomenon was furthermore specific to a sub-class of PC, sub-cortically projecting Type-A cells, and coincided with enhanced anxiety-like behavior, but no observable deficit in working memory. In contrast, female rats displayed an alcohol-induced facilitation in working memory performance with no change in expression of anxiety-like behavior. Together, these results suggest fundamental differences in alcohol effects on cell activity, cortical sub-circuits, and PFC-dependent behaviors across male and female rats.
Cell-free glycoengineering of the recombinant SARS-CoV-2 spike glycoprotein
Authors: Ruhnau Johannes , Grote Valerian , Juarez-Osorio Mariana , Bruder Dunja , Rapp Erdmann , Rexer Thomas F. T. , Reichl Udo
Published: May 2021

The baculovirus-insect cell expression system is readily utilized to produce viral glycoproteins for research as well as for subunit vaccines and vaccine candidates, for instance against SARS-CoV-2 infections. However, the glycoforms of recombinant proteins derived from this expression system are inherently different from mammalian cell-derived glycoforms with mainly complex-type N-glycans attached, and the impact of these differences in protein glycosylation on the immunogenicity is severely underinvestigated. This applies also to the SARS-CoV-2 spike glycoprotein, which is the antigen target of all licensed vaccines and vaccine candidates including virus like particles and subunit vaccines that are variants of the spike protein. Here, we expressed the transmembrane-deleted human β-1,2 N-acetlyglucosamintransferases I and II (MGAT1∆TM and MGAT2∆TM) and the β-1,4-galactosyltransferase (GalT∆TM) in E. coli to in-vitro remodel the N-glycans of a recombinant SARS-CoV-2 spike glycoprotein derived from insect cells. In a cell-free sequential one-pot reaction, fucosylated and afucosylated paucimannose-type N-glycans were converted to complex-type galactosylated N-glycans. In the future, this in-vitro glycoengineering approach can be used to efficiently generate a wide range of N-glycans on antigens considered as vaccine candidates for animal trials and preclinical testing to better characterize the impact of N-glycosylation on immunity and to improve the efficacy of protein subunit vaccines.
Disease-linked TDP-43 hyperphosphorylation suppresses TDP-43 condensation and aggregation
Authors: da Silva Lara Gruijs , Simonetti Francesca , Hutten Saskia , Riemenschneider Henrick , Sternburg Erin L. , Pietrek Lisa M. , Gebel Jakob , Dötsch Volker , Edbauer Dieter , Hummer Gerhard , Stelzl Lukas S. , Dormann Dorothee
Published: May 2021

Post-translational modifications (PTMs) have emerged as key modulators of protein phase separation and have been linked to protein aggregation in neurodegenerative disorders. The major aggregating protein in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), the RNA-binding protein TDP-43, is hyperphosphorylated in disease on several C-terminal serine residues, which is generally believed to promote TDP-43 aggregation. Here, we show that hyperphosphorylation by Casein kinase 1δ or C-terminal phosphomimetic mutations surprisingly reduce TDP-43 phase separation and aggregation and render TDP-43 condensates more liquid-like and dynamic. Multi-scale simulations reveal reduced homotypic interactions of TDP-43 low complexity domains through enhanced solvation of phosphomimetic residues. Cellular experiments show that phosphomimetic substitutions do not affect nuclear import or RNA regulatory functions of TDP-43, but suppress accumulation of TDP-43 in membrane-less organelles and promote its solubility in neurons. We propose that TDP-43 hyperphosphorylation may be a protective cellular response to counteract TDP-43 aggregation.
Characterization of a triad of genes in cyanophage S-2L sufficient to replace adenine by 2-aminoadenine in bacterial DNA
Authors: Czernecki D. , Bonhomme F. , Kaminski P.A. , Delarue M.
Published: May 2021

Cyanophage S-2L is known to profoundly alter the biophysical properties of its DNA by replacing all adenines (A) with 2-aminoadenines (Z), which still pair with thymines but with a triple hydrogen bond. It was recently demonstrated that a homologue of adenylosuccinate synthase (PurZ) and a dATP triphosphohydrolase (DatZ) are two important pieces of the metabolism of 2-aminoadenine, participating in the synthesis of ZTGC-DNA. Here, we determine that S-2L PurZ can use either dATP or ATP as a source of energy, thereby also depleting the pool of nucleotides in dATP. Furthermore, we identify a conserved gene (mazZ)located betweenpurZanddatZgenes inSiphoviridaephage genomes, and show that it encodes a (d)GTP-specific diphosphohydrolase, thereby providing the substrate of PurZ in the 2-aminoadenine synthesis pathway. High-resolution crystal structures of S-2L PurZ and MazZ with their respective substrates provide a rationale for their specificities. The Z-cluster made of these three genes –datZ,mazZandpurZ– was expressed inE. coli, resulting in a successful incorporation of 2-aminoadenine in the bacterial chromosomal and plasmidic DNA. This work opens the possibility to study synthetic organisms containing ZTGC-DNA.
Post-stroke administration of the p75 neurotrophin receptor modulator, LM11A-31, attenuates chronic changes in brain metabolism, increases neurotransmitter levels, and improves recovery
Authors: Nguyen Thuy-Vi V. , Crumpacker Rachel H. , Calderon Kylie E. , Garcia Frankie G. , Zbesko Jacob C. , Frye Jennifer B. , Gonzalez Selena , Becktel Danielle A. , Yang Tao , Tavera-Garcia Marco A. , Morrison Helena W. , Schnellmann Rick G. , Longo Frank M. , Doyle Kristian P.
Published: May 2021

The aim of this study was to test whether post-stroke oral administration of a small molecule p75 neurotrophin receptor (p75NTR) modulator (LM11A-31) can augment neuronal survival and improve recovery in a mouse model of stroke. Mice were administered LM11A-31 for up to 12 weeks, beginning 1 week after stroke. Metabolomic analysis revealed that after 2 weeks of daily treatment, mice that received LM11A-31 were distinct from vehicle treated mice by principal component analysis and had higher levels of serotonin, acetylcholine, and dopamine in their ipsilateral hemisphere. LM11A-31 treatment also improved redox homeostasis by restoring reduced glutathione. It also offset a stroke induced reduction in glycolysis by increasing acetyl-CoA. There was no effect on cytokine levels in the infarct. At 13 weeks following stroke, adaptive immune cell infiltration in the infarct was unchanged in LM11A-31 treated mice, indicating that LM11A-31 does not alter the chronic inflammatory response to stroke at the site of the infarct. However, LM11A-31 treated mice had less brain atrophy, neurodegeneration, tau pathology, and microglial activation in other regions of the ipsilateral hemisphere. These findings correlated with improved recovery of motor function on a ladder test, improved sensorimotor and cognitive abilities on a nest construction test, and less impulsivity in an open field test. These data support small molecule modulation of the p75 neurotrophin receptor for preserving neuronal health and function during stroke recovery.

SIGNIFICANCE STATEMENT

The findings from this study introduce the p75 neurotrophin receptor as a novel small molecule target for promotion of stroke recovery. Given that LM11A-31 is in clinical trials as a potential therapy for Alzheimer’s disease, it could be considered as a candidate for assessment in stroke or vascular dementia studies.
Shelterin components modulate nucleic acids condensation and phase separation in the context of telomeric DNA
Authors: Soranno Andrea , Incicco J. Jeremías , De Bona Paolo , Tomko Eric J. , Galburt Eric A. , Holehouse Alex S. , Galletto Roberto
Published: May 2021

Telomeres are nucleoprotein complexes that protect the ends of chromosomes and are essential for chromosome stability in Eukaryotes. In cells, individual telomeres form distinct globules of finite size that appear to be smaller than expected for bare DNA. Moreover, upon changes in their protein composition, telomeres can cluster to form telomere-induced-foci (TIFs) or co-localize with promyelocytic leukemia (PML) nuclear bodies. The physical basis for collapse of individual telomeres and coalescence of multiple ones remains unclear, as does the relationship between these two phenomena. By combining single-molecule measurements, optical microscopy, turbidity assays, and simulations, we show that the telomere scaffolding protein TRF2 can condense individual DNA chains and drives coalescence of multiple DNA molecules, leading to phase separation and the formation of liquid-like droplets. Addition of the TRF2 binding protein hRap1 modulates phase boundaries and tunes the specificity of solution demixing while simultaneously altering the degree of DNA compaction. Our results suggest that the condensation of single telomeres and formation of biomolecular condensates containing multiple telomeres are two different outcomes driven by the same set of molecular interactions. Moreover, binding partners, such as other telomere components, can alter those interactions to promote single-chain DNA compaction over multiple-chain phase separation.
Early acquisition of threat conditioning in a selectively-bred anxiety-like rat phenotype: regulation by maternal presence and FGF2
Authors: White Amanda M. , Chang Da-Jeong , Hider Joanna H. , Hebda-Bauer Elaine K. , Turner Cortney A. , Sullivan Regina M. , Akil Huda , Debiec Jacek
Published: May 2021

Temperament is an innate, stable predisposition towards particular emotional and behavioral responses. In humans, certain temperaments are associated with a heightened risk of developing anxiety later in life. Non-human animals, including rodents, also exhibit innate, stable dispositions; these are referred to as behavioral phenotypes. The interaction between behavioral phenotype and early life adverse events is critical for the development of maladaptive anxiety. Rodent studies of typically developing animals have identified a number of mechanisms that protect against aversive experiences in early life. One such mechanism is an early life quiescence of threat learning, which protects against the effects of stress and facilitates safety and attachment learning. However, little is known about the factors that alleviate the effects of early life aversive events on phenotypes vulnerable to pathological anxiety. Here, we examined threat learning and the stress response in selectively-bred infant rats that show an anxiety-like phenotype relative to typically developing animals. We investigated the potential roles of maternal presence and the anxiolytic neurotrophic factor fibroblast growth factor 2 (FGF2) in regulating threat learning and the stress response in infant anxiety-like phenotype animals. We observed that rats selectively-bred for anxiety-like behaviors could acquire conditioned freezing earlier in life than typically developing animals. FGF2 administration on postnatal day 1 (PND 1) and maternal presence during threat conditioning were both capable of suppressing this early emergence of conditioned freezing. However, neither FGF2 nor maternal presence during threat conditioning were associated with reduced corticosterone levels during threat conditioning. Our results suggest that although an anxiety-like phenotype may be associated with early threat learning, environmental factors (such as maternal presence) and pharmacological intervention (such as modulation of the FGF2 system) may be capable of counteracting that early aversive learning. Interventions in vulnerable infants may thus decrease the impact of aversive events.
The sperm protein SPACA4 is required for efficient fertilization in mice
Authors: Herberg Sarah , Fujihara Yoshitaka , Blaha Andreas , Panser Karin , Kobayashi Kiyonori , Larasati Tamara , Novatchkova Maria , Theußl H. Christian , Olszanska Olga , Ikawa Masahito , Pauli Andrea
Published: May 2021

Fertilization is the fundamental process that initiates the development of a new individual in all sexually reproducing species. Despite its importance, our understanding of the molecular players that govern mammalian sperm-egg interaction is incomplete, partly because many of the essential factors found in non-mammalian species do not have obvious mammalian homologs. We have recently identified the Ly6/uPAR protein Bouncer as a new, essential fertilization factor in zebrafish (Herberg et al., 2018). Here, we show that Bouncer’s homolog in mammals, SPACA4, is also required for efficient fertilization in mice. In contrast to fish, where Bouncer is expressed specifically in the egg, SPACA4 is expressed exclusively in the sperm. Male knockout mice are severely sub-fertile, and sperm lacking SPACA4 fail to fertilize wild-type eggsin vitro. Interestingly, removal of the zona pellucida rescues the fertilization defect ofSpaca4-deficient spermin vitro, indicating that SPACA4 is not required for the interaction of sperm and the oolemma but rather of sperm and zona pellucida. Our work identifies SPACA4 as an important sperm protein necessary for zona pellucida penetration during mammalian fertilization.
Oxidized alkyl phospholipids stimulate sodium transport in proximal tubules via PPARγ-dependent pathway
Authors: Mizuno Tomohito , Satoh Nobuhiko , Horita Shoko , Tsukada Hiroyuki , Sato Yusuke , Kume Haruki , Nangaku Masaomi , Nakamura Motonobu
Published: May 2021

The pleiotropic effects of oxidized phospholipids (oxPLs) have been identified. 1-O-hexadecyl-2-azelaoyl-sn-glycero-3-phosphocholine (azPC), an oxPL formed from alkyl phosphatidylcholines, is a potent peroxisome proliferator-activated receptor γ (PPARγ) agonist. Although it has been reported that thiazolidinediones can induce volume expansion by enhancing renal sodium and water retention, the role of azPC, an endogenous PPARγ agonist, in renal transport functions is unknown. In the present study, we investigated the effect of azPC on renal proximal tubule (PT) transport using isolated PTs and kidney cortex tissues. We showed that azPC rapidly stimulated Na+/HCO3-cotransporter 1 activity and luminal Na+/H+exchanger (NHE) activities in a dose-dependent manner, at submicromolar concentrations, in isolated PTs from rats and humans. Additionally, the stimulatory effects were completely blocked by a specific PPARγ antagonist, 2-chloro-5-nitro-N-phenylbenzamide (GW9662), and a mitogen-activated protein/extracellular signal-regulated kinase (MEK) inhibitor, PD98059. Treatment with an siRNA against PPARγ significantly suppressed the expression of PPARγ mRNA, and it completely blocked the stimulation of both Na+/HCO3-cotransporter 1 and NHE activities by azPC. Moreover, azPC induced extracellular signal-regulated kinase (ERK) phosphorylation in rat and human kidney cortex tissues, and the induced ERK phosphorylation by azPC was completely suppressed by GW9662 and PD98059. These results suggest that azPC stimulates renal PT sodium-coupled bicarbonate transport via the PPARγ/MEK/ERK pathway. The stimulatory effects of azPC on PT transport may be partially involved in the development of volume expansion.
Simple and efficient heterologous expression of necrosis-inducing effectors using the model plantNicotiana benthamiana
Authors: Dagvadorj Bayantes , Solomon Peter S.
Published: May 2021

Plant fungal pathogens cause devastating diseases on cereal plants and threaten global food security. During infection, these pathogens secrete proteinaceous effectors that promote disease. Some of these effectors from necrotrophic plant pathogens induce a cell death response (necrosis), which facilitates pathogen growthin planta. Characterisation of these effectors typically requires heterologous expression and microbial expression systems such as bacteria and yeast are the predominantly used. However, microbial expression systems often require optimization for any given effector and are, in general, not suitable for effectors involving cysteine bridges and posttranslational modifications for activity. Here, we describe a simple and efficient method for expressing such effectors in the model plantNicotiana benthamiana. Briefly, an effector protein is transiently expressed and secreted into the apoplast ofN. benthamianaby Agrobacterium-mediated infiltration. Two-to-three days subsequent to agroinfiltration, the apoplast from the infiltrated leaves is extracted and can be directly used for phenotyping on host plants. The efficacy of this approach was demonstrated by expressing the ToxA, Tox3 and Tox1 necrosis-inducing effectors fromParastagonospora nodorum. All three effectors produced inN. benthamianawere capable of inducing necrosis in wheat lines, and two of three showed visible bands on Coomassie-stained gel. These data suggest thatN. benthamiana-agroinfiltration system is a feasible tool to obtain fungal effectors, especially those that require disulfide bonds and posttranslational modifications. Furthermore, due to the low number of proteins typically observed in the apoplast (compared to intracellular), this simple and high-throughput approach circumvents the requirement to lyse cells and further purify the target proteins that is required in other heterologous systems. Because of its simplicity and potential for high-throughput, this method is highly amenable to the phenotyping of candidate protein effectors on host plants.
Data fusion for abundance estimation: community science augments systematically collected removal-in-time distance sampling data
Authors: Joseph Maxwell B. , Pavlacky David C. , Bartuszevige Anne M.
Published: May 2021

Ecologists use a variety of systematically and opportunistically sampled count data to estimate bird abundance, and integrating or fusing different datasets has emerged as a critical challenge in recent years. While previous work provides data integration methodology for occupancy (presence/absence) estimation, methods for abundance estimation that account for imperfect detection and disparate survey protocols remains an active area of research. Here we show how to integrate systematically collected removal-in-time distance sampling data from the Integrated Monitoring in Bird Conservation Regions (IMBCR) program with North American Breeding Bird Survey (BBS) point counts and eBird community science observations. Using the Grasshopper Sparrow (Ammodramus savannarum) in the Great Plains of the United States as a focal species, we demonstrate that BBS and eBird data improve predictive performance for IMBCR count data, providing more spatially refined and precise estimates of abundance at regional scales. Data fusion increased predictive performance even despite relatively weak spatial correlations among data sets. The methodology developed here provides a principled way to fuse data when estimating abundance with distance sampling, that accounts for imperfect detection and variable effort.
Human iPSC-derived brain endothelial microvessels in a multi-well format enable permeability screens of anti-inflammatory drugs
Authors: Fengler Sven , Kurkowsky Birgit , Kaushalya Sanjeev Kumar , Roth Wera , Fava Eugenio , Denner Philip
Published: May 2021

Optimizing drug candidates for blood-brain barrier (BBB) penetration in humans remains one of the key challenges and many devastating brain diseases including neurodegenerative diseases still do not have adequate treatments. So far, it has been difficult to establish state-of-the-art human stem cell derivedin vitromodels that mimic physiological barrier properties including a 3D microvasculature in a format that is scalable enough to screen drugs for BBB penetration in early drug development phases. To address this challenge, we established human induced pluripotent stem cell (iPSC)-derived brain endothelial microvessels in a standardized and scalable multi-well plate format. iPSC-derived brain microvascular endothelial cells (BMECs) were supplemented with primary cell conditioned media and grew to intact microvessels in 10 days of culturing. Produced microvessels show a typical BBB phenotype including endothelial protein expression, tight-junctions and polarized localization of efflux transporter. Microvessels exhibited physiological relevant trans-endothelial electrical resistance (TEER), were leak-tight for 10 kDa dextran-Alexa 647 and strongly limited the permeability of sodium fluorescein (NaF). Permeability tests with reference compounds confirmed the suitability of our model as platform to identify potential BBB penetrating anti-inflammatory drugs. In summary, the here presented brain microvessel platform recapitulates physiological properties and allows rapid screening of BBB permeable anti-inflammatory compounds that has been suggested as promising substances to cure so far untreatable neurodegenerative diseases.
Gated recurrence enables simple and accurate sequence prediction in stochastic, changing, and structured environments
Authors: Foucault Cedric , Meyniel Florent
Published: May 2021

From decision making to perception to language, predicting what is coming next is crucial. It is also challenging in stochastic, changing, and structured environments; yet the brain makes accurate predictions in many situations. What computational architecture could enable this feat? Bayesian inference makes optimal predictions but is prohibitively difficult to compute. Here, we show that a specific recurrent neural network architecture enables simple and accurate solutions in several environments. This architecture relies on three mechanisms: gating, lateral connections, and recurrent weight training. Like the optimal solution and the human brain, such networks develop internal representations of their changing environment (including estimates of the environment’s latent variables and the precision of these estimates), leverage multiple levels of latent structure, and adapt their effective learning rate to changes without changing their connection weights. Being ubiquitous in the brain, gated recurrence could therefore serve as a generic building block to predict in real-life environments.
Sex-specific multi-level 3D genome dynamics in the mouse brain
Authors: Rocks Devin , Shukla Mamta , Finnemann Silvia C. , Kalluchi Achyuth , Rowley M. Jordan , Kundakovic Marija
Published: May 2021

The female mammalian brain exhibits sex-hormone-driven plasticity during the reproductive period. Evidence implicates chromatin dynamics in gene regulation underlying this plasticity. However, whether ovarian hormones impact higher-order chromatin organization in post-mitotic neuronsin vivois unknown. Here, we mapped 3D genome of ventral hippocampal neurons across the estrous cycle and by sex in mice. In females, we found cycle-driven dynamism in 3D chromatin organization, including in estrogen-response-elements-enriched X-chromosome compartments, autosomal CTCF loops, and enhancer-promoter interactions. With rising estrogen levels, the female 3D genome becomes more similar to the male genome. Cyclical enhancer-promoter interactions are partially associated with gene expression and enriched for brain disorder-relevant genes. Our study reveals unique 3D genome dynamics in the female brain relevant to female-specific gene regulation, neuroplasticity, and disease risk.
Muscle fatigue revisited - Insights from optically pumped magnetometers
Authors: Sometti Davide , Semeia Lorenzo , Chen Hui , Dax Juergen , Kronlage Cornelius , Baek Sangyeob , Kirchgässner Milena , Romano Alyssa , Heilos Johanna , Staber Deborah , Oppold Julia , Righetti Giulia , Middelmann Thomas , Braun Christoph , Broser Philip , Marquetand Justus
Published: May 2021

Muscle fatigue is well characterized electromyographically, nevertheless only information about summed potential differences is detectable. In contrast, recently developed quantum sensors optically pumped magnetometers (OPMs) offer the advantage of recording both the electrical current propagation in the muscle as well as its geometry, by measuring the magnetic field generated by the muscular action potentials. Magnetomyographic investigation of muscle fatigue is still lacking and it is an open question whether fatigue is characterized similarly in magnetomyography (MMG) compared to electromyography (EMG). Herein, we investigated the muscle fatigue during a 3×1-min strong isometric contraction of the rectus femoris muscle of 12 healthy subjects using simultaneous EMG-MMG (4-channel surface EMG and 4 OPM along the rectus femoris muscle).

Both EMG and MMG showed the characteristic frequency decrease in the signal magnitude during isometric contraction, which is typical for muscle fatigue. In addition, it was shown that the main part of this frequency decrease seems to occur in the circular component of the magnetic field around the muscle fibers and less longitudinally along the muscle fibers. Overall, these results show not only that magnetomyography is capable of reproducing the electromyographic standards in identifying muscular fatigue, but it also adds relevant information about the spatial characterization of the signal. Therefore, OPM-MMG offers new insights for the study of muscular activity and might serve as a new, supplementary neurophysiological method.
Orthology clusters from gene trees withPossvm
Authors: Grau-Bové Xavier , Sebé-Pedrós Arnau
Published: May 2021

Possvm(PhylogeneticOrthologSorting withSpecies oVerlap andMCL) is a tool that automates the process of classifying clusters of orthologous genes from precomputed phylogenetic trees. It identifies orthology relationships between genes using the species overlap algorithm to infer taxonomic information from the gene tree topology, and then uses the Markov Clustering Algorithm (MCL) to identify orthology clusters and provide annotated gene family classifications. Our benchmarking shows that this approach, when provided with accurate phylogenies, is able to identify manually curated orthogroups with high precision and recall. Overall,Possvmautomates the routine process of gene tree inspection and annotation in a highly interpretable manner, and provides reusable outputs that can be used to obtain phylogeny-informed gene annotations and inform comparative genomics and gene family evolution analyses.
Giant ankyrin-B mediates transduction of axon guidance and collateral branch pruning factor Sema 3A
Authors: Creighton Blake A. , Ajit Deepa , Afriyie Simone , Bay Julia , Lorenzo Damaris
Published: May 2021

Variants in the high confident autism spectrum disorder geneANK2target both ubiquitously expressed 220-kDa ankyrin-B and neurospecific 440-kDa ankyrin-B (AnkB440) isoforms. Previous work showed that knock-in mice expressing an ASD-linkedAnk2variant yielding a truncated AnkB440 product exhibit ectopic brain connectivity and behavioral abnormalities. Expression of this variant or loss of AnkB440 caused axonal hyperbranchingin vitro, which implicated AnkB440 microtubule bundling activity in suppressing collateral branch formation. Leveraging multiple mouse models, cellular assays, and live microscopy, we show that AnkB440 also modulates axon collateral branching stochastically by reducing the number of F-actin-rich branch initiation points. Additionally, we show that AnkB440 enables growth cone (GC) collapse in response to chemorepellent factor semaphorin 3A (Sema 3A) by stabilizing its receptor complex L1 cell adhesion molecule/neuropilin-1. ASD-linkedANK2variants failed to rescue Sema 3A-induced GC collapse. We propose that impaired response to repellent cues due to AnkB440 deficits leads to axonal guidance and branch pruning defects and may contribute to the pathogenicity ofANK2variants.
A conserved Guided Entry of Tail-anchored pathway is involved in the trafficking of tail-anchored membrane proteins inPlasmodium falciparum
Authors: Kumar Tarkeshwar , Maitra Satarupa , Rahman Abdur , Bhattacharjee Souvik
Published: May 2021

Tail-anchored (TA) proteins are defined by the absence of N-terminus signal sequence and the presence of a single transmembrane domain (TMD) proximal to their extreme C-terminus. They play fundamental roles in cellular processes including vesicular trafficking, protein translocation and quality control. Accordingly, TA proteins are post-translationally integrated by the Guided Entry of TA (GET) pathway to the cellular membranes; with their N-terminus oriented towards the cytosol and C-terminus facing the organellar lumen. The TA repertoire and the GET machinery have been extensively characterized in the yeast and mammalian systems, however, they remain elusive in the human malaria parasitePlasmodium falciparum.In this study, we bioinformatically predicted a total of 63 TA proteins in theP. falciparumproteome and revealed the association of their subset with theP. falciparumhomolog of Get3 (PfGet3). In addition, our proximity labelling studies either definitively identified or shortlisted the other eligible GET constituents, and ourin vitroassociation studies validated associations between PfGet3 and the corresponding homologs of Get4 and Get2 inP. falciparum. Collectively, this study reveals the presence of proteins with hallmark TA signatures and the involvement of evolutionary conserved GET trafficking pathway for their targeted delivery within the parasite.

Synopsis

Tail-anchored (TA) proteins, characterized by an absence of N-terminal signal sequence and the presence of a transmembrane domain near the C-terminus, are post-translationally inserted at organellar membranes by the conserved multi-component Guided Entry of TA (GET) pathway. Here, we identified the putative homologs of GET machinery in the human malaria parasitePlasmodium falciparumand revealed their association with a subset of bioinformatically predicted 63 putative TA proteins, thereby validating the functional existence of this trafficking pathway within the apicomplexan parasite.

Graphical Abstract
DELAY OF GERMINATION 6, encoding theANAC060transcription factor, inhibits seed dormancy
Authors: Song Shuang , He Hanzi , Gühl Kerstin , van Bolderen-Veldkamp Marieke , Buijs Gonda , Willems Leo A.J. , Bentsink Leónie
Published: May 2021

The timing of seed germination is regulated by seed dormancy. There is ample natural variation for seed dormancy among as well as within plant species. In Arabidopsis severalDELAY OF GERMINATIONquantitative trait loci have been identified, of whichDOG1is best studied. Here we report the identification ofDOG6, a quantitative trait locus with a similar strong effect on seed dormancy asDOG1. DOG6affects the timing of germination both in laboratory as well as in field conditions. Complementation cloning revealed thatDOG6encodes the membrane bound transcription factorANAC060. The absence of the ANAC060 protein or its sequestration outside the nucleus results in increased seed dormancy levels. The different natural variants ofANAC060differ for the presence of the membrane binding domain, either due to the fact that this domain is absent in the genomic sequence or because the cDNA is alternatively spliced. Our data indicates that ANAC060 regulates seed dormancy by among others binding to and regulating the expression of protein phosphatases 2C class A proteins including PROTEIN PHOSPHATASE 2CA (PP2CA), ABI FIVE BINDING PROTEIN 3 (AFP3) and HIGHLY ABA-INDUCED PP2C GENE 3 (HAI3).

Significance Statement

ANAC transcription factors are known to effect plant development as well as the response of plants to their environment. Here, we present the identification ofDELAY OF GERMINATION 6(DOG6), a seed dormancy quantitative trait locus that encodes theANAC060transcription factor. We have identified different natural alleles ofANAC060and show that these genetic variants determine the localization of the protein.ANAC060alleles that lack the membrane binding domain end up in the nucleus. Hence they affect transcription and as such attenuate seed dormancy.
Small-molecule modulators of TRMT2A decrease PolyQ aggregation and PolyQ-induced cell death
Authors: Margreiter Michael A , Witzenberger Monika , Wasser Yasmine , Davydova Elena , Janowski Robert , Sobisch Carina , Metz Jonas , Poma Benedetta , Palomino-Hernandez Oscar , Habib Pardes , Wagner Mirko , Carell Thomas , Shah N Jon , Schulz Jörg B , Niessing Dierk , Voigt Aaron , Rossetti Giulia
Published: May 2021

Polyglutamine (polyQ) diseases are characterized by an expansion of cytosine-adenine-guanine (CAG) trinucleotide repeats encoding for an uninterrupted prolonged polyQ tract. We previously identified TRMT2A as a strong modifier of polyQ-induced toxicity in an unbiased large-scale screen in Drosophila melanogaster. This work aimed at identifying and validating pharmacological TRMT2A inhibitors as treatment opportunities for polyQ diseases in humans. Computer-aided drug discovery was implemented to identify human TRMT2A inhibitors. Additionally, the crystal structure of one protein domain, the RNA recognition motif (RRM), was determined, and Biacore experiments with the RRM were performed. The identified molecules were validated for their potency to reduce polyQ aggregation and polyQ-induced cell death in human HEK293T cells and patient derived fibroblasts. Our work provides a first step towards pharmacological inhibition of this enzyme and indicates TRMT2A as a viable drug target for polyQ diseases.