bluuebunny/crossref_metadata_2025 huggingface dataset:
Soluble ECM promotes organotypic formation in lung alveolar model
Authors: Valdoz Jonard C. , Franks Nicholas A. , Cribbs Collin G. , Jacobs Dallin J. , Dodson Ethan L. , Knight Connor J. , Poulson P. Daniel , Garfield Seth R. , Johnson Benjamin C. , Hemeyer Brandon M. , Sudo Miranda T. , Saunooke Jordan A. , Kartchner Braden C. , Saxton Aubrianna , Vallecillo-Zuniga Mary L. , Santos Matheus , Chamberlain Brandon , Christensen Kenneth A. , Nordin Greg P. , Narayanan A. Sampath , Raghu Ganesh , Van Ry Pam M.
Published: September 2021

Micropatterned suspension culture creates consistently sized and shaped cell aggregates but has not produced organotypic structures from stable cells, thus restricting its use in accurate disease modeling. Here, we show that organotypic structure is achieved in hybrid suspension culture via supplementation of soluble extracellular matrix (ECM). We created a viable lung organoid from epithelial, endothelial, and fibroblast human stable cell lines in suspension culture. We demonstrate the importance of soluble ECM in organotypic patterning with the emergence of lumen-like structures with airspace showing feasible gas exchange units, formation of branching, perfusable vasculature, and long-term 70-day maintenance of lumen structure. Our results show a dependent relationship between enhanced fibronectin fibril assembly and the incorporation of ECM in the organoid. We successfully applied this technology in modeling lung fibrosis via bleomycin induction and test a potential antifibrotic drug in vitro while maintaining fundamental cell-cell interactions in lung tissue. Our human fluorescent lung organoid (hFLO) model represents features of pulmonary fibrosis which were ameliorated by fasudil treatment. We also demonstrate a 3D culture method with potential of creating organoids from mature cells, thus opening avenues for disease modeling and regenerative medicine, enhancing understanding of lung cell biology in health and lung disease.
Cardiotoxic effects of methamphetamine associated with electrophysiological and epigenetic aberrations in zebrafish
Authors: Zhang Jimmy , Nguyen Anh H. , Schmiess-Heine Lauren , Le Tai , Xia Xing , Lau Michael P.H. , Lee Juhyun , Cao Hung
Published: September 2021

Long-term methamphetamine (Meth) abuse damages functional and molecular changes in the brain that causes chronic and relapsing disease. In this study, we sought to investigate a relationship between cardiotoxicity and arrhythmia with associated Meth abuse in zebrafish to identify and to understand the adverse cardiac symptoms associated with Meth as well as to assess the applicability of zebrafish as an appropriate model for cardiac-related drug screening studies. Over a two-week duration, zebrafish were first treated with various concentrations of Meth, ranging from 0 to 50 μM. Immediately after treatment, zebrafish underwent electrocardiogram (ECG) measurement for electrophysiological analysis. Results show that a higher incidence of increased heart rate over the duration of the experiment, corroborating with results from previous human case studies involving Meth users. However, abnormalities commonly cited in those same case studies, such as prolongation of QTc, were not significantly presented in obtained ECG recordings. We have also conducted genetic, epigenetic, and histochemical analysis in an attempt to understand the cardiotoxic effects of Meth on zebrafish cardiac function. These results suggested myocardial damage and decrease in gene expression associated with normal physiological function. Finally, this paper provides insights into potential reasons for the apparent discrepancies in our data with prior research as well as an outlook of zebrafish cardiotoxic drug screening studies.
Combi-Seq: Multiplexed transcriptome-based profiling of drug combinations using deterministic barcoding in single-cell droplets
Authors: Mathur L , Szalai B , Utharala R , Ballinger M , Landry JJM , Ryckelynck M , Benes V , Saez-Rodriguez J , Merten CA
Published: September 2021

Anti-cancer therapies often exhibit only short-term effects. Tumors typically develop drug resistance causing relapses that might be tackled with drug combinations. Identification of the right combination is challenging and would benefit from high-content, high-throughput combinatorial screens directly on patient biopsies. However, such screens require a large amount of material, normally not available from patients. To address these challenges, we developed a scalable microfluidic workflow to screen hundreds of drug combinations in picoliter-size droplets using transcriptome changes as a readout for drug effects. We devised a deterministic combinatorial DNA barcoding approach to encode treatment conditions, enabling the gene expression-based readout of drug effects in a highly multiplexed fashion. We applied our method to screen the effect of 420 drug combinations on the transcriptome of K562 cells using only ∼250 single cell droplets per condition, to successfully predict synergistic and antagonistic drug pairs, as well as their pathway activities.
Transport among protocells via tunneling nanotubes
Authors: Schanke Ingrid Jin , Xue Lin , Spustova Karolina , Gözen Irep
Published: September 2021

We employ model protocell networks for evaluation of molecular transport through lipid nanotubes as potential means of communication among primitive cells on the early Earth. Network formation is initiated by deposition of multilamellar lipid reservoirs onto a silicon oxide surface in an aqueous environment. These reservoirs autonomously develop into surface-adhered protocells interconnected via lipid nanotubes, and encapsulate solutes from the ambient buffer in the process. We prepare networks in the presence of DNA and RNA and observe encapsulation of these molecules, and their diffusive transport between the lipid compartments via the interconnecting nanotubes. By means of an analytical model we determine key physical parameters affecting the transport, such as nanotube diameter and compartment size. We conclude that nanotube-mediated transport in self-organized nanotube-vesicle networks could have been a possible pathway of chemical communication between primitive, self-assembled protocells under early earth conditions, circumventing the necessity for crossing the membrane barrier. We suggest this transport within a closed protocell network as a feasible means of RNA and DNA exchange under primitive prebiotic conditions, possibly facilitating early replication.
Antioxidant Response Activating nanoParticles (ARAPas) localize to atherosclerotic plaque and locally activate the Nrf2 pathway
Authors: Maiocchi Sophie , Cartaya Ana , Thai Sydney , Akerman Adam , Bahnson Edward
Published: September 2021

Atherosclerotic disease is the leading cause of death world-wide with few novel therapies available despite the ongoing health burden. Redox dysfunction is a well-established driver of atherosclerotic progression; however, the clinical translation of redox-based therapies is lacking. One of the challenges facing redox-based therapies is their targeted delivery to cellular domains of redox dysregulation. In the current study, we sought to develop Antioxidant Response Activating nanoParticles (ARAPas), encapsulating redox-based interventions, that exploit macrophage biology and the dysfunctional endothelium in order to selectively accumulate in atherosclerotic plaque. We employed flash nanoprecipitation (FNP) to synthesize bio-compatible polymeric nanoparticles encapsulating the hydrophobic Nrf2 activator drug, CDDO-Methyl (CDDOMe-ARAPas). Nuclear factor erythroid 2-related factor 2 (Nrf2)-activators are a promising class of redox-active drug molecules whereby activation of Nrf2 results in the expression of several antioxidant and cyto-protective enzymes that can be athero-protective. In this study, we characterize the physiochemical properties of CDDOMe-ARAPas as well as confirm their in vitro internalization by murine macrophages. Drug release of CDDOMe was determined by Nrf2-driven GFP fluorescence. Moreover, we show that these CDDOMe-ARAPas exert anti-inflammatory effects in classically activated macrophages. Finally, we show that CDDOMe-ARAPas selectively accumulate in atherosclerotic plaque of two widely-used murine models of atherosclerosis: ApoE−/−and LDLr−/−mice, and are capable of increasing gene expression of Nrf2-transcriptional targets in the atherosclerotic aortic arch. Future work will assess the therapeutic efficacy of intra-plaque Nrf2 activation with CDDOMe-ARAPas to inhibit atherosclerotic plaque progression. Overall, our present studies underline that targeting of atherosclerotic plaque is an effective means to enhance delivery of redox-based interventions.
Multimodal Detection of Dopamine by Sniffer Cells Expressing Genetically Encoded Fluorescence Sensors
Authors: Herenbrink Carmen Klein , Støier Jonatan Fullerton , Reith William Dalseg , Dagra Abeer , Cuadrado Gregorek Miguel Alejandro , Li Yulong , Tian Lin , Gether Ulrik , Herborg Freja
Published: September 2021

Dopamine serves an important role in supporting both locomotor control and higher brain functions such as motivation and learning. Dopaminergic dysfunction is implicated in an equally multidimensional spectrum of neurological and neuropsychiatric diseases. Extracellular dopamine levels are known to be tightly controlled by presynaptic dopamine transporters (DAT), which is also a main target of psychostimulants. Still, detailed data on dopamine dynamics in space and time is needed to fully understand how dopamine signals are encoded and translated into cellular and behavioral responses, and to uncover the pathological effects of dopamine-related diseases. The recently developed genetically encoded fluorescent dopamine sensors enable unprecedented monitoring of dopamine dynamics and have changed the field of in vivo dopamine recording. However, the potential of these sensors to be used for in vitro and ex vivo assays remains unexplored. Here, we demonstrate a generalizable blueprint for making “sniffer” dopamine cells for multimodal detection of dopamine in vitro and ex vivo. We generated sniffer cell lines with inducible expression of six different dopamine sensors and performed a head-to-head comparison of sensor properties to guide users in sensor selection. In proof-of-principle experiments, we show how the sniffer cells can be applied to measure release of endogenous dopamine from cultured neurons and striatal slices, and for determining total dopamine content in striatal tissue. Furthermore, we use the sniffer cells to quantify DAT-mediated dopamine uptake, and AMPH-induced and constitutive dopamine efflux as a radiotracer free, high-throughput alternative to electrochemical- and radiotracer-based assays. Importantly, the sniffer cells framework can readily be applied to other transmitter systems for which the list of genetically encoded fluorescent sensors is rapidly growing.
Development of infants’ neural speech processing and its relation to later language skills: an MEG study
Authors: Zhao T. Christina , Kuhl Patricia K.
Published: September 2021

The ‘sensitive period’ for phonetic learning (∼6-12 months) is one of the earliest milestones in language acquisition where infants start to become specialized in processing speech sounds in their native language. In the last decade, advancements in neuroimaging technologies for infants are starting to shed light on the underlying neural mechanisms supporting this important learning period. The current study reports on the largest longitudinal dataset to date with the aim to replicate and extend on two important questions: 1) what are the developmental changes during the ‘sensitive period’ for native and nonnative speech processing? 2) how does native and nonnative speech processing in infants predict later language outcomes? Fifty-four infants were recruited at 7 months of age and their neural processing of speech was measured using Magnetoencephalography (MEG). Specifically, the neural sensitivity to a native and a nonnative speech contrast was indexed by the mismatch response (MMR). They repeated the measurement again at 11 months of age and their language development was further tracked from 12 months to 30 months of age using the MacArthur-Bates Communicative Development Inventory (CDI). Using an a prior Region-of-Interest (ROI) approach, we observed significant increases for the Native MMR in the left inferior frontal region (IF) and superior temporal region (ST) from 7 to 11 months, but not for the Nonnative MMR. Complementary whole brain comparison revealed more widespread developmental changes for both contrasts. However, only individual differences in the left IF and ST for the Nonnative MMR at 11 months of age were significant predictors of individual vocabulary growth up to 30 months of age. An exploratory machine-learning based analysis further revealed that whole brain MMR for both Native and Nonnative contrasts can robustly predict later outcomes, but with very different underlying spatial-temporal patterns. The current study extends our current knowledge and suggests that native and nonnative speech processing may follow different developmental trajectories and utilize different mechanisms that are relevant for later language skills.
Developing selective FPR2 agonists can be a potential approach to treat moderate to severe asthma
Authors: Visaga Senthil A. , Kalonia Harikesh , Verma Vinay , Sinha Sandeep , Singh Shashi Kant , Upadhyay Swati , Sahdev Sudhir , Pansari Amita , Kumar Rajesh , Bandgar Mahadev , Karanjule Narayan , Shirumalla Raj Kumar , Morishita Kaoru , Tandon Ruchi
Published: September 2021

Formyl peptide receptor (FPR) family members have been reported to play important role in the resolution of inflammation. A few FPR2/FPR1 dual agonists are reported in the public domain for their anti-inflammatory properties. None of these molecules, however, have been successful as a therapy yet. Recent reports bring forward the ambiguous role of FPR1 in inflammation. These include both positive and negative outcomes. We, therefore, aimed to develop selective FPR2 agonists and evaluated their potential in mitigating the non-resolving inflammation in mouse models of moderate to severe asthma. Extensive structure-activity-relationship (SAR) studies were conducted on the imidazole and benzimidazole chemotype series to identify potent and selective FPR2 agonists. A few molecules were shortlisted based on their in vitro profile and absorption, distribution, metabolism and excretion (ADME) properties and were further evaluated in mouse models of asthma. We report herewith identification of 3 RCI compounds with low nanomolar potency for FPR2 agonism and >10,000 fold selectivity over FPR1 in Ca2+ release assay. These molecules also showed potency in other in vitro assays and potent efficacy in three distinct animal models of asthma. Our data suggest that FPR2 agonism can be a potential therapeutic approach to treat asthma. Our findings also propose that FPR1 can be spared to achieve the desired pharmacological activity.
Deletion of serine palmitoyl transferase 2 in hepatocytes impairs ceramide/sphingomyelin balance, prevents obesity and leads to liver damage in mice
Authors: Lallement Justine , Raho Ilyès , Merlen Grégory , Rainteau Dominique , Croyal Mikael , Schiffano Melody , Kassis Nadim , Doignon Isabelle , Soty Maud , Lachkar Floriane , Krempf Michel , Foufelle Fabienne , Amouyal Chloé , Le Stunff Hervé , Magnan Christophe , Tordjmann Thierry , Cruciani-Guglielmacci Céline
Published: September 2021

Ceramides (Cer) have been shown as lipotoxic inducers, which disturb numerous cell signalling pathways especially insulin signalling pathway leading to metabolic disorders such as type 2 diabetes. In this study, we aimed to determine the role of de novo hepatic Cer synthesis on energy and liver homeostasis in mice. We generated mice lacking serine palmitoyltransferase 2 (Sptlc2), the rate limiting enzyme of Cer de novo synthesis, in hepatocytes.

Despite lower expression of hepatic Sptlc2, we observed an increased concentration of hepatic Cer, especially C16:0-Cer and C18:0-Cer associated with an increased neutral sphingomyelinase 2 expression, and a decreased sphingomyelin content in the liver. Sptlc2ΔHep mice were protected against obesity induced by high fat diet. Bile acid (BA) hydrophobicity was drastically decreased in KO mice, and was associated with a defect in lipid absorption. In addition, an important increase of tauro-muricholic acid in BA pool composition was associated with a downregulation of the nuclear BA receptor FXR target genes. Sptlc2 deficiency also enhanced glucose tolerance and attenuated hepatic glucose production. Finally, Sptlc2 disruption promoted apoptosis, inflammation and progressive development of hepatic fibrosis worsening with age.

Our data suggest a compensatory mechanism to regulate hepatic Cer content from sphingomyelin hydrolysis, with deleterious impact on liver homeostasis. In addition, our results show the implication of hepatic sphingolipid modulation on BA metabolism and hepatic glucose production in an insulinin-dependent manner, which demonstrates the role of Cer in many metabolic functions still under-researched.
Acoustic array biochip combined with allele-specific PCR for multiple cancer mutation analysis in tissue and liquid biopsy
Authors: Naoumi Nikoletta , Michaelidou Kleita , Papadakis George , Simaiaki Agapi E. , Fernández Román , Calero Maria , Arnau Antonio , Tsortos Achilleas , Agelaki Sofia , Gizeli Electra
Published: September 2021

Regular screening of cancerous point mutations is of importance to cancer management and treatment selection. Although excellent techniques like next generation sequencing and droplet digital PCR are available, these are still lacking in speed, simplicity and cost-effectiveness. Here a new approach is presented where allele-specific PCR (AS-PCR) is combined with a novel High Fundamental Frequency Quartz Crystal Microbalance (HFF-QCM) array biosensor for the amplification and detection, respectively, of cancer point mutations. For the proof-of-concept, the method was applied to the screening of the BRAF V600E and KRAS G12D mutations in spiked-in and clinical samples. Regarding the BRAF target, an analytical sensitivity of 0.01%, i.e., detection of 1 mutant copy of genomic DNA in an excess of 104 wild type molecules, was demonstrated; moreover, quantitative results during KRAS detection were obtained when an optimized assay was employed with a sensitivity of 0.05%. The assays were validated using tissue and plasma samples obtained from melanoma, colorectal and lung cancer patients. Results are in full agreement with Sanger sequencing and droplet digital PCR, demonstrating efficient detection of BRAF and KRAS mutations in samples having an allele frequency below 1%. The high sensitivity and technology-readiness level of the methodology, together with the ability for multiple sample analysis (24 array biochip), cost-effectiveness and compatibility with routine work-flow, hold promise for the implementation of this AS-PCR/acoustic methodology in clinical oncology as a tool for tissue and liquid biopsy.
3D virtual Histopathology of Cardiac Tissue from Covid-19 Patients based on Phase-Contrast X-ray Tomography
Authors: Reichardt Marius , Jensen Patrick Møller , Dahl Vedrana Andersen , Dahl Anders Bjorholm , Ackermann Maximilian , Shah Harshit , Länger Florian , Werlein Christopher , Kühnel Mark , Jonigk Danny , Salditt Tim
Published: September 2021

For the first time, we have used phase-contrast x-ray tomography to characterize the three-dimensional (3d) structure of cardiac tissue from patients who succumbed to Covid-19. By extending conventional histopatholocigal examination by a third dimension, the delicate pathological changes of the vascular system of severe Covid-19 progressions can be analyzed, fully quantified and compared to other types of viral myocarditis and controls. To this end, cardiac samples with a cross section of 3.5mm were scanned at the synchrotron in a parallel beam configuration. The vascular network was segmented by a deep learning architecture suitable for 3d datasets (V-net), trained by sparse manual annotations. Pathological alterations of vessels, concerning the variation of diameters and the amount of small holes, were observed, indicative of elevated occurrence of intussusceptive angiogenesis, also confirmed by scanning electron microscopy. Further, we implemented a fully automated analysis of the tissue structure in form of shape measures based on the structure tensor. The corresponding distributions show that the histopathology of Covid-19 differs from both influenza and typical coxsackie virus myocarditis.
Androglobin, a chimeric mammalian globin, is required for male fertility
Authors: Keppner Anna , Correia Miguel , Santambrogio Sara , Koay Teng Wei , Maric Darko , Osterhof Carina , Winter Denise V , Clerc Angèle , Stumpe Michael , Chalmel Frédéric , Dewilde Sylvia , Odermatt Alex , Kressler Dieter , Hankeln Thomas , Wenger Roland H. , Hoogewijs David
Published: September 2021

Spermatogenesis is a highly specialised process, involving multiple dedicated pathways and regulatory check-points. Defects ultimately lead to male sub-fertility or sterility, and numerous aspects of mammalian sperm formation remain unknown. The predominant expression of the latest globin family member, androglobin (Adgb) in mammalian testis tissue prompted us to assess its physiological function in spermatogenesis. Adgb knockout mice display male infertility, reduced testis weight, impaired maturation of elongating spermatids, abnormal sperm shape and ultrastructural defects in microtubule and mitochondrial organisation. Epididymal sperm from Adgb knockout animals display multiple flagellar malformations including coiled, bifide or shortened flagella, and erratic acrosomal development. Following immunoprecipitation and mass spectrometry, we could identify septin 10 (Sept10) as interactor of Adgb. The Sept10-Adgb interaction was confirmed both in vivo using testis lysates, and in vitro by reciprocal co-immunoprecipitation experiments. Furthermore, absence of Adgb leads to mislocalisation of Sept10 in sperm, indicating defective manchette and sperm annulus formation. Finally, in vitro data suggest that Adgb contributes to Sept10 proteolysis in a calmodulin (CaM)-dependent manner. Collectively, our results provide evidence that Adgb is essential for murine spermatogenesis and further suggest that interdependence between Adgb and Sept10 is required for sperm head shaping via the manchette and proper flagellum formation.
Adiponectin receptor agonist AdipoRon improves skeletal muscle function in aged mice
Authors: Balasubramanian Priya , Schaar Anne E. , Gustafson Grace E. , Smith Alex B. , Howell Porsha R. , Greenman Angela , Baum Scott , Colman Ricki J. , Lamming Dudley W. , Diffee Gary , Anderson Rozalyn M.
Published: September 2021

The loss of skeletal muscle function with age, known as sarcopenia, significantly reduces independence and quality of life and can have significant metabolic consequences. Although exercise is effective in treating sarcopenia it is not always a viable option clinically, and currently there are no pharmacological therapeutic interventions for sarcopenia. Here we show that chronic treatment with pan-adiponectin receptor agonist AdipoRon improved muscle function in male mice by a mechanism linked to skeletal muscle metabolism and tissue remodeling. In aged mice, 6 weeks of AdipoRon treatment improved skeletal muscle functional measures in vivo and ex vivo. Improvements were linked to changes in fiber type, including an enrichment of oxidative fibers, and an increase in mitochondrial activity. In young mice, 6 weeks of AdipoRon treatment improved contractile force and activated the energy sensing kinase AMPK and the mitochondrial regulator PGC-1a (peroxisome proliferator activated receptor gamma coactivator 1 alpha). In cultured cells, the AdipoRon induced stimulation of AMPK and PGC-1a was associated with increased mitochondrial membrane potential, reorganization of mitochondrial architecture, increased respiration, and increased ATP production. Furthermore, the ability of AdipoRon to stimulate AMPK and PGC1a was conserved in nonhuman primate cultured cells. These data show that AdipoRon is an effective agent for the prevention of sarcopenia in mice and indicate that its effects translate to primates, suggesting it may also be a suitable therapeutic for sarcopenia in clinical application.
The Barley HvSTP13GR mutant triggers resistance against biotrophic fungi
Authors: Skoppek Ines Caroline , Punt Wilko , Heinrichs Marleen , Ordon Frank , Wehner Gwendolin , Boch Jens , Streubel Jana
Published: September 2021

High-yielding and stress resistant crops are essential to ensure future food supply. Barley is an important crop to feed livestock and to produce malt, but the annual yield is threatened by pathogen infections. Pathogens can trigger an altered sugar partitioning in the host plant, that possibly leads to an advantage for the pathogen. Hampering these processes represents a promising strategy to potentially increase resistance. We analyzed the response of the barley monosaccharide transporter HvSTP13 towards biotic stress and its potential use for plant protection. The expression of HvSTP13 increased upon bacterial and fungal PAMP application, suggesting a PAMP-triggered signaling that converged on the transcriptional induction of the gene. Promoter studies indicate a region that is likely targeted by transcription factors downstream of PAMP-triggered immunity pathways. We confirmed that the non-functional HvSTP13GR variant confers resistance against an economically relevant biotrophic rust fungus, in barley. In addition, we established targeted CRISPR/Cas9 cytosine base editing in barley protoplasts to generate alternative HvSTP13 mutants and characterized the sugar transport activity and subcellular localization of the proteins. These mutants represent promising variants for future resistance analysis. Our experimental setup provides basal prerequisites to further decode the role of HvSTP13 in response to biological stress. Moreover, in line with other studies, our experiments indicate that the alteration of sugar partitioning pathways, in a host pathogen interaction, is a promising approach to achieve broad and durable resistance in plants.
Molecular Basis for the Interaction of Catalase with D-Penicillamine : Rationalization of some of its Deleterious Effects
Authors: Padovani Dominique , Galardon Erwan
Published: September 2021

D-penicillamine (D-Pen) is a sulfur compound used in the management of rheumatoid arthritis, Wilson’s disease (WD), and alcohol dependence. Many side effects are associated with its use, particularly after long-term treatment. However, the molecular bases for such side effects are poorly understood. Based on the well-known oxidase activity of hemoproteins, and the participation of catalase in cellular H2O2redox signaling, we posit that D-Pen could inactivate catalase, thus disturbing H2O2levels. Herein, we report on the molecular bases that could partly explain the side effects associated with this drug compound, and we demonstrate that it induces the formation of compound II, a temporarily inactive state of the enzyme, through two distinct mechanisms. Initially, D-Pen reacts with native catalase and/or iron metal ions, used to mimic non heme iron overload observed in long-term treated WD patients, to generate thiyl radicals. These partake into a futile redox cycling, thus producing superoxide radical anions O2•-and hydrogen peroxide H2O2. Then, either H2O2unexpectedly reacts with native CAT-Fe(II) to produce compound II, or both aforementioned reactive oxygen species intervene into compound II generation through compound I formation then reduction. These findings support evidence that D-Pen could perturb H2O2redox homeostasis through transient but recurring catalase inactivation, which may in part rationalize some deleterious effects observed with this therapeutic agent, as discussed.

Graphical abstract
AHK5 mediates ETR1-initiated multistep phosphorelay in Arabidopsis
Authors: Szmitkowska Agnieszka , Cuyacot Abigail Rubiato , Pekárová Blanka , Žd’árská Markéta , Houser Josef , Komárek Jan , Jaseňáková Zuzana , Jayasree Aswathy , Heunemann Michael , Ubogoeva Elena , Spyroglou Ioannis , Trtilek Martin , Mironova Victoria , Harter Klaus , Zemlyanskaya Elena , Žídek Lukáš , Wimmerová Michaela , Hejátko Jan
Published: September 2021

Plants, like other sessile organisms, need to sense many different signals, and in response to them, modify their developmental programs to be able to survive in a highly changing environment. The multistep phosphorelay (MSP) in plants is a good candidate for a response mechanism that integrates multiple signal types both environmental and intrinsic in origin. Recently, ethylene was shown to control MSP activity via the histidine kinase (HK) activity of ETHYLENE RESPONSE 1 (ETR1)1,2, but the underlying molecular mechanism still remains unclear.

Here we show that although ETR1 is an active HK, its receiver domain (ETR1RD) is structurally and functionally unable to accept the phosphate from the phosphorylated His in the ETR1 HK domain (ETR1HK) to initiate the phosphorelay to ARABIDOPSIS HISTIDINE-CONTAINING PHOSPHOTRANSMITTERs (AHPs), the next link downstream members in MSP signaling. Instead, ETR1 interacts with another HK ARABIDOPSIS HISTIDINE KINASE 5 (AHK5) and transfers the phosphate from ETR1HK through the receiver domain of AHK5 (AHK5RD), and subsequently to AHP1, AHP2 and AHP3, independently of the HK activity of AHK5. We show that AHK5 is necessary for ethylene-initiated, but not cytokinin-initiated, MSP signaling in planta and that it thus mediates hormonal control of root growth.
Metabolic diversity and aero-tolerance in anammox bacteria from geochemically distinct aquifers
Authors: Mosley Olivia E. , Gios Emilie , Weaver Louise , Close Murray , Daughney Chris , van der Raaij Rob , Martindale Heather , Handley Kim M.
Published: September 2021

Background

Anaerobic ammonium oxidation (anammox) is important for converting bioavailable nitrogen into dinitrogen gas, particularly in carbon poor environments. Yet, the diversity and prevalence of anammox bacteria in the terrestrial subsurface – a typically oligotrophic environment – is little understood across different geochemical conditions. To determine the distribution and activity of anammox bacteria across a range of aquifer lithologies and physicochemistries, we analysed 16S rRNA genes, metagenomes and metatranscriptomes, and quantified hydrazine synthase genes and transcripts sampled from 59 groundwater wells distributed over 1 240 km2.

Results

Data indicate that anammox-associated bacteria (class Brocadiae) and the anammox process are prevalent in aquifers (identified in aquifers with sandy-gravel, sand-silt and volcanic lithologies). While Brocadiae diversity decreased with increasing DO, Brocadiae 16S rRNA genes and hydrazine synthase genes and transcripts (hydrazine synthase, hzsB) were detected across a wide range of bulk groundwater dissolved oxygen (DO) concentrations (0 – 10 mg/L). Anammox genes and transcripts (hzsB) correlated significantly with those involved in bacterial and archaeal ammonia oxidation (ammonia monooxygenase, amoA), which could represent a major source of nitrite for anammox. Differences in anammox community composition were strongly associated with DO and bore depth (and to a lesser extent pH and phosphate), revealing niche differentiation among anammox bacteria in groundwater that was largely driven by water oxygen contents, and not ammonium/nitrite. Eight Brocadiae genomes (63-95% estimated completeness) reconstructed from a subset of groundwater sites belong to 2 uncharacterized families and 6 novel species (based on average nucleotide identity). Distinct groups of these genomes dominated the anammox-associated community at dysoxic and oxic sites, further reflecting the influence of DO on Brocadiae composition. Six of the genomes (dominating dysoxic or oxic sites) have genes characteristic of anammox (hydrazine synthase and/or dehydrogenase). These genes, in addition to aerotolerance genes, belonging to four Brocadiae genomes, were transcriptionally active, although transcript numbers clearly highest in dyoxic groundwater.

Conclusions

Our findings indicate anammox bacteria contribute to loss of fixed N across diverse anoxic-to-oxic aquifer conditions, and that this is likely supported by nitrite from aerobic ammonia oxidation. Results provide an insight into the distribution and activity of anammox bacteria across distinct aquifer physicochemisties.
Modulation of optical speaker using biogenic guanine platelets floating in water
Authors: Iwasaka Masakazu
Published: September 2021

Microphones are miniature devices for sound detection. Various technologies have been developed to transfer sound properties onto other physical quantities, e.g., electricity. Over the past three decades, many studies have reported on optical sensing of sound. Most of these studies were performed via application of light interference at the edges of optical fibers. Several studies have reported on detection of sounds in the air or objects causing mechanical vibrations based on light interference. This work proposes an optical speaker which is a method to reconstruct and modulate sound from the power spectrum of light that has been reflected by guanine platelets floating in water droplet. The water droplet containing fish guanine platelets was placed on a piezoelectric membrane and acoustic vibration from the membrane propagated inside the droplet. A photomultiplier tube (PMT) then collected the light reflected from the water droplet. Without post-analysis of the measured light intensity, the analog output voltage from the PMT clearly sounded an audio speaker. In addition, it was found that the guanine platelets reflecting light operated as an audio equalizer.
Assessing the zoonotic potential of a novel bat morbillivirus
Authors: Ikegame Satoshi , Carmichael Jillian C. , Wells Heather , O’Brien Robert L. Furler , Acklin Joshua A. , Chiu Hsin-Ping , Oguntuyo Kasopefoluwa Y. , Cox Robert M. , Patel Aum R. , Kowdle Shreyas , Stevens Christian S. , Eckley Miles , Zhan Shijun , Lim Jean K. , Veit Ethan C. , Evans Matthew , Hashiguchi Takao , Durigon Edison , Schountz Tony , Epstein Jonathan H. , Plemper Richard K. , Daszak Peter , Anthony Simon J. , Lee Benhur
Published: September 2021

Morbilliviruses are amongst the most contagious viral pathogens that infect mammals. Metagenomic surveys have identified numerous morbillivirus sequences in bats, but no full-length authentic morbillivirus has been isolated or characterized from bats. Here we detail the discovery of full-length Myotis Bat Morbillivirus (MBaMV) from a bat surveillance program in Brazil. After determining that MBaMV utilizes bat CD150 but not human CD150 as an entry receptor, we generated an infectious clone of MBaMV using reverse genetics. MBaMV exhibited features consistent with other morbilliviruses, including pleomorphic virions, P-editing and the rule-of-six. MBaMV replicated well in human epithelial cell lines in a nectin-4 dependent manner. Surprisingly, MBaMV was able to infect human macrophages in a CD150-independent manner. However, MBaMV was restricted by cross-neutralizing human sera and did not evade the human innate immune system, indicating that while zoonotic spillover into humans may be possible, MBaMV replication in humans would likely be restricted.
Pharmacological rescue of tumor intrinsic STING expression and immune response in LKB1-mutant lung cancer via the IAP-JAK regulatory axis
Authors: Shu Changfa , Jin Rui , Niu Qiankun , Cicka Danielle , Doyle Sean , Wahafu Alafate , Fan Dacheng , Zheng Xi , Du Yuhong , Ivanov Andrey A. , Doxie Deon B , Dhodapkar Kavita M , Carlisle Jennifer , Owonikoko Taofeek , Ramalingam Suresh , Sica Gabriel , Dhodapkar Madhav V , Zhou Wei , Mo Xiulei , Fu Haian
Published: September 2021

Harnessing the power of the immune system to treat cancer has become a core clinical approach. However, rewiring of intrinsic circuitry enables tumor cells to escape immune attacks, leading to therapeutic failure. Pharmacological strategies to reverse tumor genotype-dictated therapeutic resistance are urgently needed to advance precision immunotherapy. Here, we identify antagonists of Inhibitor of Apoptosis Protein (IAP) as potent sensitizers that restore immune-dependent killing of LKB1-mutant lung cancer cells. Mechanistic studies reveal an LKB1-IAP-JAK trimolecular complex that bridges the LKB1-mutant genotype with IAP-dependency and a STING-deficiency-mediated immune resistance phenotype. Ultimately, inhibition of IAP re-establishes JAK-regulated STING expression and DNA sensing pathway as well as enhanced cytotoxic immune cell infiltration and selective immune-dependent anti-tumor activity in an LKB1-mutant immune-competent mouse model. Thus, IAP-JAK-modulatory strategies, like IAP inhibitors, offer promising immunotherapy adjuvants to re-establish the responsiveness of “immunologically-cold” LKB1-mutant tumors to immune checkpoint inhibitors or STING-directed therapies.
Discovery of frog virus 3 microRNAs and their roles in evasion of host antiviral responses
Authors: Todd Lauren A. , Katzenback Barbara A.
Published: September 2021

Frog virus 3 (FV3, genus Ranavirus) causes devastating disease in amphibian populations and is capable of subverting host immune responses. Evidence suggests that virus-encoded microRNAs (v-miRNAs) are implicated in host immunoevasion tactics. Thus, we sought to discover FV3-encoded v-miRNAs and to uncover their putative roles in immunoevasion. Small RNA libraries were generated from FV3-infected Xela DS2, a Xenopus laevis dorsal skin epithelial-like cell line, at 24- and 72-hours post-infection (hpi). We discovered 43 FV3 v-miRNAs and identified that 15 are upregulated at 24 hpi, while 18 are upregulated at 72 hpi. Target prediction analyses revealed that FV3 v-miRNAs target host genes involved in key antiviral signaling pathways, while gene ontology analyses suggest that FV3 v-miRNAs may broadly impact host cell function. This is the first study to experimentally detect mature v-miRNAs produced by FV3. Our findings highlight the possibility that ranaviral v-miRNAs facilitate immunoevasion of frog antiviral responses.
Cofilin Regulates Filopodial Structure and Flexibility in Neuronal Growth Cones
Authors: Hylton Ryan , Heebner Jessica , Grillo Michael , Swulius Matthew
Published: September 2021

Cofilin is best known for its ability to sever actin filaments, and facilitate cytoskeletal recycling inside of cells. At higher concentrations, in vitro, cofilin stabilizes a more flexible, hyper-twisted state of actin known as “cofilactin”, but a structural role for cofilactin, in situ, has not been observed. Combining cryo-electron tomography and live-cell imaging in neuronal growth cones, we show that filopodial actin bundles can switch between a fascin-linked and a cofilin-decorated state, composed of hyper-twisted cofilactin filaments. These cofilactin bundles contribute to the flexibility of filopodial actin networks, thus regulating growth cone searching dynamics. Our results provide mechanistic insight into the processes underlying proper brain development, as well as fundamentals of cytoskeletal mechanics inside confined cellular spaces.
Genetically diverse mouse models of SARS-CoV-2 infection reproduce clinical variation in type I interferon and cytokine responses in COVID-19
Authors: Robertson Shelly J. , Bedard Olivia , McNally Kristin L. , Shaia Carl , Clancy Chad , Lewis Matthew , Broeckel Rebecca M. , Chiramel Abhilash I. , Shannon Jeffrey G. , Sturdevant Gail L. , Rosenke Rebecca , Anzick Sarah L. , Forte Elvira , Preuss Christoph , Baker Candice N. , Harder Jeffrey , Brunton Catherine , Munger Steven , Bruno Daniel P. , Lack Justin B. , Leung Jacqueline M. , Shamsaddini Amirhossein , Gardina Paul , Sturdevant Daniel E. , Sun Jian , Martens Craig , Holland Steven M. , Rosenthal Nadia A. , Best Sonja M.
Published: September 2021

Inflammation in response to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection drives severity of coronavirus disease 2019 (COVID-19) and is influenced by host genetics. To understand mechanisms of inflammation, animal models that reflect genetic diversity and clinical outcomes observed in humans are needed. We report a mouse panel comprising the genetically diverse Collaborative Cross (CC) founder strains crossed to human ACE2 transgenic mice (K18-hACE2) that confers susceptibility to SARS-CoV-2. Infection of CC x K18- hACE2 resulted in a spectrum of survival, viral replication kinetics, and immune profiles. Importantly, in contrast to the K18-hACE2 model, early type I interferon (IFN-I) and regulated proinflammatory responses were required for control of SARS-CoV-2 replication in PWK x K18-hACE2 mice that were highly resistant to disease. Thus, virus dynamics and inflammation observed in COVID-19 can be modeled in diverse mouse strains that provide a genetically tractable platform for understanding anti-coronavirus immunity.

One Sentence Summary

Genetically diverse mice model a spectrum of clinically relevant innate immune responses to SARS-CoV-2 infection.
Quantifying the impact of gut microbiota on inflammation and hypertensive organ damage
Authors: Avery Ellen G. , Bartolomaeus Hendrik , Rauch Ariana , Chen Chia-Yu , N’Diaye Gabriele , Löber Ulrike , Bartolomaeus Theda U. P. , Fritsche-Guenther Raphaela , Rodrigues André F. , Tsvetkov Dmitry , Todiras Mihail , Park Joon-Keun , Markó Lajos , Maifeld András , Bader Michael , Kempa Stefan , Kirwan Jennifer A. , Forslund Sofia K. , Müller Dominik N. , Wilck Nicola
Published: September 2021

Aims

Hypertension (HTN) can lead to heart and kidney damage. The gut microbiota has been linked to HTN, although it is difficult to estimate its significance due to the variety of other features known to influence HTN. In the present study, we used germ-free (GF) and colonized (COL) littermate mice to quantify the impact of microbial colonization on organ damage in HTN.

Methods and results

Four-week-old male GF C57BL/6J littermates were randomized to remain GF or receive microbial colonization. HTN was induced by subcutaneous infusion with angiotensin (Ang) II (1.44mg/kg/d) and 1% NaCl in the drinking water; sham-treated mice served as control. Renal damage was exacerbated in GF mice, whereas cardiac damage was more comparable between COL and GF, suggesting that the kidney is more sensitive to microbial influence. Multivariate analysis revealed a larger effect of HTN in GF mice. Serum metabolomics demonstrated that the colonization status influences circulating metabolites relevant to HTN. Importantly, GF mice were deficient in anti-inflammatory fecal short-chain fatty acids (SCFA). Flow cytometry showed that the microbiome has an impact on the induction of anti-hypertensive myeloid-derived suppressor cells and pro-inflammatory Th17 cells in HTN. In vitro inducibility of Th17 cells was significantly higher for cells isolated from GF than conventionally raised mice.

Conclusions

Microbial colonization status of mice had potent effects on their phenotypic response to a hypertensive stimulus, and the kidney is a highly microbiota-susceptible target organ in HTN. The magnitude of the pathogenic response in GF mice underscores the role of the microbiome in mediating inflammation in HTN.

Translation Perspective

To assess the potential of microbiota-targeted interventions to prevent organ damage in hypertension, an accurate quantification of microbial influence is necessary. We provide evidence that the development of hypertensive organ damage is dependent on colonization status and suggest that a healthy microbiota provides anti-hypertensive immune and metabolic signals to the host. In the absence of normal symbiotic host-microbiome interactions, hypertensive damage to the kidney in particular is exacerbated. We suggest that hypertensive patients experiencing perturbations to the microbiota, which are common in CVD, may be at a greater risk for target-organ damage than those with a healthy microbiome.
Kalium rhodopsins: Natural light-gated potassium channels
Authors: Govorunova Elena G. , Gou Yueyang , Sineshchekov Oleg A. , Li Hai , Wang Yumei , Brown Leonid S. , Xue Mingshan , Spudich John L.
Published: September 2021

We report a family of K+channels, kalium channelrhodopsins (KCRs) from a fungus-like protist. Previously known potassium channels, widespread and mainly ligand- or voltage-gated, share a conserved pore-forming domain and K+-selectivity filter. KCRs differ in that they are light-gated and they have independently evolved an alternative K+selectivity mechanism. The KCRs are potent, highly selective of K+over Na+, and open in less than 1 millisecond following photoactivation. Their permeability ratio PK/PNaof ∼ 20 make KCRs powerful hyperpolarizing tools that suppress excitable cell firing upon illumination, demonstrated here in mouse cortical neurons. KCRs enable specific optogenetic photocontrol of K+gradients promising for the study and potential treatment of potassium channelopathies such as epilepsy, Parkinson’s disease, and long-QT syndrome and other cardiac arrhythmias.

One-Sentence Summary

Potassium-selective channelrhodopsins long-sought for optogenetic research and therapy of neurological and cardiac diseases.
The isolation and characterization of Taphrina betulina and other yeasts residing in the Betula pendula phylloplane
Authors: Christita Margaretta , Sipilä Timo P. , Overmyer Kirk
Published: September 2021

The phylloplane is an important microbial habitat and a reservoir of organisms that affect plant health, both positively and negatively. Taphrina betulina is the causative agent of birch witches’ broom disease. Taphrina are dimorphic, invading theirs hosts in a filamentous form and residing in the host phyllosphere in their non-infectious yeast form. As such, they are widely accepted to be found a resident yeasts on their hosts, even on healthy tissues; however, there is little experimental data to support this. With the aim of exploring the local infection ecology of T. betulina, we had isolated yeasts from the phylloplane of birch, using three classes of samples; from infected symptom bearing leaves inside brooms, healthy leaves from branches away from brooms on broom bearing trees, and symptom-free leaves from symptom-free trees. Isolations yielded 224 yeast strains, representing 11 taxa, including T. betulina, which was the most common isolate and was found in all sample classes, including asymptomatic leaves. Genotyping with two genetic markers revealed genetic diversity among these T. betulina isolates, with seven distinct genotype differentiated by the markers used. Of the 57 T. betulina strains, 22 representative strains were selected for further studies and preliminarily characterized, revealing differences in size and the ability to produced compounds with activity to activate the signalling pathway for the plant hormone auxin.