bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sensitive protein detection using site-specifically oligonucleotide-conjugated nanobodies
Authors: Al-Amin Rasel A. , Muthelo Phathutshedzo M. , Abdurakhmanov Eldar , Vincke Cécile , Amin Shahnaz P. , Muyldermans Serge , Danielson U. Helena , Landegren Ulf 
Published: January 2022

ABSTRACTHigh-quality affinity probes are critical for sensitive and specific protein detection, in particular for detection of protein biomarkers at early phases of disease development. Proximity extension assays (PEA) have been used for high-throughput multiplexed protein detection of up to a few thousand different proteins in one or a few microliters of plasma. Clonal affinity reagents can offer advantages over the commonly used polyclonal antibodies (pAbs) in terms of reproducibility and standardization of such assays. Here we explore nanobodies (Nb) as an alternative to pAbs as affinity reagents for PEA. We describe an efficient site-specific approach for preparing high-quality oligo-conjugated Nb probes via enzyme coupling using Sortase A (SrtA). The procedure allows convenient removal of unconjugated affinity reagents after conjugation. The purified high-grade Nb probes were used in PEA and the reactions provided an efficient means to select optimal pairs of binding reagents from a group of affinity reagents. We demonstrate that Nb-based PEA (nano-PEA) for interleukin-6 (IL6) detection can augment assay performance, compared to the use of pAb probes. We identify and validate Nb combinations capable of binding in pairs without competition for IL6 antigen detection by PEA.
Altered retinal structure and function in Spinocerebellar ataxia type 3
Authors: Toulis Vasileios , Casaroli-Marano Ricardo , Camós-Carreras Anna , Figueras-Roca Marc , Sánchez-Dalmau Bernardo , Muñoz Esteban , Ashraf Naila S. , Ferreira Ana F. , Khan Naheed , Marfany Gemma , do Carmo Costa Maria 
Published: January 2022

ABSTRACTSpinocerebellar ataxia type 3 is an autosomal dominant neurodegenerative disorder caused by expansion of a polyglutamine (polyQ)-encoding CAG repeat in the ATXN3 gene. Because the ATXN3 protein regulates photoreceptor ciliogenesis and phagocytosis, we aimed to explore whether expanded polyQ ATXN3 impacts retinal function and integrity in SCA3 patients and transgenic mice.We evaluated the retinal structure and function in five patients with Spinocerebellar ataxia type 3 and in a transgenic mouse model of this disease (YACMJD84.2, Q84) using, respectively, optical coherence tomography (OCT) and electroretinogram (ERG). We further determined in the transgenic mice: a) the retinal expression pattern of ATXN3 and assessed the distribution of cones and rods by immunofluorescence (IF); and b) the retinal ultrastructure by transmission electron microscopy (TEM).Some patients with Spinocerebellar ataxia type 3 in our cohort revealed: i) reduced central macular thickness indirectly correlated with disease duration; ii) decreased thickness of the macula and the ganglion cell layer, and reduced macula volume inversely correlated with disease severity (SARA score); and iii) electrophysiological dysfunction of cones, rods, and inner retinal cells. Transgenic mice replicated the human OCT and ERG findings with aged homozygous Q84/Q84 mice showing a stronger phenotype accompanied by further thinning of the outer nuclear layer and photoreceptor layer and highly reduced cone and rod activities, thus supporting severe retinal dysfunction in these mice. In addition, Q84 mice showed progressive accumulation of ATXN3-positive aggregates throughout several retinal layers and depletion of cones alongside the disease course. TEM analysis of aged Q84/Q84 mouse retinas supported the IF ATXN3 aggregation findings by revealing the presence of high number of negative electron dense puncta in ganglion cells, inner plexiform and inner nuclear layers, and further showed thinning of the outer plexiform layer, thickening of the retinal pigment epithelium and elongation of apical microvilli.Our results indicate that retinal alterations detected by non-invasive eye examination using OCT and ERG could represent a biological marker of disease progression and severity in patients with Spinocerebellar ataxia type 3.
Intracellular flow cytometry staining of antibody-secreting cells using phycoerythrin-conjugated antibodies: pitfalls and solutions
Authors: Renner Patrick , Crone Michael , Kornas Matthew , Pioli KimAnh T. , Pioli Peter D. 
Published: January 2022

AbstractAntibody-secreting cells are terminally differentiated B cells that play a critical role in humoral immunity through immunoglobulin secretion along with possessing the potential to be long-lived. It is now appreciated that antibody-secreting cells regulate multiple aspects of biology through the secretion of various cytokines. In this regard, intracellular flow cytometry is a key tool used to assess the presence of intracellular proteins such as cytokines and transcription factors. Here, we showed that the use of phycoerythrin-containing antibody conjugates led to a false interpretation of antibody-secreting cell intracellular protein expression compared to other cell types. This was mainly due to the inappropriate retention of these antibodies specifically within antibody-secreting cells. Furthermore, we demonstrated how to reduce this retention which allowed for a more accurate comparison of intracellular protein expression between antibody-secreting cells and other cell types such as B lymphocytes. Using this methodology, our data revealed that spleen antibody-secreting cells expressed Toll-like receptor 7 as well as the pro-form of the inflammatory cytokine interleukin-1β.
Multi-GBS: A massively multiplexed GBS-based protocol optimized for large, repetitive conifer genomes
Authors: Vallebueno-Estrada Miguel , Steindl Sonja , Akulova Vasilina , Riefler Julia , Slusarz Lucyna , Arizpe Alexis , Swarts Kelly 
Published: January 2022

AbstractReduced representation library approaches are still a valuable tool for breeding and population and ecological genomics, even with impressive increases in sequencing capacity in recent years. Unfortunately, current approaches only allow for multiplexing up to 384 samples. To take advantage of increased sequencing capacity, we present Multi-GBS, a massively multiplexable extension to Genotyping-by-Sequencing that is also optimized for large conifer genomes. In Norway Spruce, a highly repetitive 20Gbp diploid genome with high population genetic variation, we call over a million variants in 32 genotypes from three populations, two natural forest in the Alps and Bohemian Alps, and a managed population from southeastern Austria using the existing TASSEL GBSv2 pipeline. Metric MDS analysis of replicated genotypes shows that technical bias in resulting genotype calling is minimal and that populations cluster in biologically meaningful ways.
Vascular topology is acutely impacted by experimental febrile status epilepticus
Authors: Salehi Arjang , Salari Sirus , Daglian Jennifer , Chen Kevin , Baram Tallie Z. , Obenaus Andre 
Published: January 2022

AbstractFebrile status epilepticus (FSE) is an important risk factor for temporal lobe epilepsy and early identification is vital. In a rat model of FSE, we identified an acute novel MRI signal in the basolateral amygdala (BLA) at 2 hours post FSE that predicted epilepsy in adulthood. This signal remains incompletely understood and hypothesized that it might derive from changes to vascular topology. Experimental FSE was induced in rat pups and compared to normothermic littermate controls. We examined cerebral vascular topology at 2 hours, using a novel vessel painting and analysis protocol. Blood vessel density of the cortical vasculature was significantly reduced in FSE rats, and this effect was lateralized, as reported for the MRI signal. The middle cerebral artery (MCA) exhibited abnormal topology in FSE pups but not in controls. In the BLA, significant vessel junction reductions and decreased vessel diameter were observed, together with a strong trend for reduced vessel length. In summary, FSE results in acute vascular topological changes in the cortex and BLA that may underlie the acute MRI signal that predicts progression to future epilepsy. The altered vasculature may be amenable to intervention treatments to potentially reduce the probability of progression to epilepsy following FSE.
How many bits of information can be transferred between residues in a protein and how fast?
Authors: Hacisuleyman Aysima , Erman Burak 
Published: January 2022

AbstractTime resolved Raman and infrared spectroscopy experiments show the basic features of information transfer between residues in proteins. Here, we present the theoretical basis of information transfer using a simple elastic net model and recently developed entropy transfer concept in proteins. Mutual information between two residues is a measure of communication in proteins which shows the maximum amount of information that may be transferred between two residues. However, it does not explain the actual amount of transfer nor the transfer rate of information between residues. For this, dynamic equations of the system are needed. We used the Schreiber theory of information transfer and the Gaussian network Model of proteins, together with the solution of the Langevin equation, to quantify allosteric information transfer. Results of the model are in perfect agreement with ultraviolet resonance Raman measurements. Analysis of the allosteric protein Human NAD-dependent isocitrate dehydrogenase shows that a multitude of paths contribute collectively to information transfer. While the peak values of information transferred are small relative to information content of residues, considering the estimated transfer rates, which are in the order of megabits per second, sustained transfer during the activity time-span of proteins may be significant.
Membrane-active Polymers: NCMNP13-x, NCMNP21-x and NCMNP21b-x for Membrane Protein Structural Biology
Authors: Trinh Thi Kim Hoang , Catalano Claudio , Guo Youzhong 
Published: January 2022

AbstractMembrane proteins are a ubiquitous group of bio-macromolecules responsible for many crucial biological processes and serve as drug targets for a wide range of modern drugs. Detergent-free technologies such as styrene-maleic acid lipid particles (SMALP), diisobutylene-maleic acid lipid particles (DIBMALP), and native cell membrane nanoparticles (NCMN) systems have recently emerged as revolutionary alternatives to the traditional detergent-based approaches for membrane protein research. NCMN systems aim to create a membrane-active polymer library suitable for high-resolution structure determination. Herein, we report our design, synthesis, characterization and comparative application analyses of three novel classes of NCMN polymers, NCMNP13-x, NCMNP21-x and NCMNP21b-x. Although each NCMN polymer can solubilize various model membrane proteins and conserve native lipids into NCMN particles, only the NCMNP21b-x series reveals lipid-protein particles with good buffer compatibility and high homogeneity suitable for single-particle cryo-EM analysis. Consequently, the NCMNP21b-x polymers that bring out high-quality NCMN particles are particularly attractive for membrane protein structural biology.Graphical abstractHighlightsDemonstrate straightforward strategies for tailoring styrene-maleic co-polymer (SMA) that revealed novel buffer compatible polymers, NCMNP13-50, NCMNP21-20 and NCMNP21b-x.Elucidate how modification factors alter the membrane-active properties of these polymers, i.e., membrane protein extraction efficiency, morphology, etc.Provide valuable insights into the rational design of membrane-active polymers for membrane protein structural biology.NCMNP21b-x polymers are highly compatible with high-resolution structure determination using an emerging technique, cryo-EM.
Evidence for quinol oxidase activity of ImoA, a novel NapC/NirT family protein from the neutrophilic Fe(II) oxidizing bacterium Sideroxydans lithotrophicus ES-1
Authors: Jain Abhiney , Coelho Anaísa , Madjarov Joana , Todorovic Smilja , Louro Ricardo O. , Gralnick Jeffrey A. , Paquete Catarina M. 
Published: January 2022

AbstractThe freshwater chemolithoautotrophic Gram-negative bacterium Sideroxydans lithotrophicus ES-1 oxidizes Fe(II) at the cell surface. In this organism, it is proposed that the monoheme cytochrome MtoD from the Mto pathway transfer electrons across the periplasm to an inner membrane NapC/NirT family tetraheme cytochrome encoded by Slit_2495, for which we propose the name ImoA (inner membrane oxidoreductase). ImoA has been proposed to function as the quinone reductase, receiving electrons from iron oxidizing extracellular electron uptake pathway to reduce the quinone pool. In this study, ImoA was cloned on a pBAD plasmid vector and overexpressed in Escherichia coli. Biochemical and spectroscopic characterization of the purified ImoA reveals that this 26.5 kDa cytochrome contains one high-spin and three low-spin hemes. Our data show that ImoA can function as a quinol oxidase and is able to functionally replace CymA, a related NapC/NirT family tetraheme cytochrome required for anaerobic respiration of a wide range of substrates by Shewanella oneidensis. We demonstrate that ImoA can transfer electrons to different periplasmic proteins from S. oneidensis including STC and FccA, but in a manner that is distinct from that of CymA. Phylogenetic analysis shows that ImoA is clustered closer to NirT sequences than to CymA. This study suggests that ImoA functions as a quinol oxidase in S. oneidensis and raises questions about the directionality and/or reversibility of electron flow through the Mto pathway in S. lithotrophicus ES-1.ImportanceFe(II)-oxidizing bacteria play an important role in the biogeochemical cycling of iron, representing a promising class of organisms for the development of novel biotechnological processes, including bioelectrosynthesis. These organisms perform extracellular electron transfer, taking up electrons from Fe(II) outside of the cell, possibly through a porin-cytochrome complex in the outer membrane. The electrons are then transferred to the quinone pool in the inner membrane via periplasmic and inner membrane electron transfer proteins. In this paper, we produced and characterized the NapC/NirT family tetraheme cytochrome ImoA, encoded by Slit_2495, an inner membrane protein from the Gram-negative Fe(II)-oxidizing bacterium Sideroxydans lithotrophicus ES-1, proposed to be involved in extracellular electron transfer to the quinone pool. We show that ImoA may function instead as a quinol oxidase. The obtained insights represent the first step in understanding mechanisms of electron flow in S. lithotrophicus ES-1 and may lead towards practical biotechnological applications of Fe(II)-oxidizing bacteria.
Responses to song playback differ in sleeping versus anesthetized songbirds
Authors: Bottjer Sarah W. , Le Moing Chloé , Li Ellysia , Yuan Rachel 
Published: January 2022

AbstractVocal learning in songbirds is mediated by a highly localized system of interconnected forebrain regions, including recurrent loops that traverse the cortex, basal ganglia, and thalamus. This brain-behavior system provides a powerful model for elucidating mechanisms of vocal learning, with implications for learning speech in human infants, as well as for advancing our understanding of skill learning in general. A long history of experiments in this area has tested neural responses to playback of different song stimuli in anesthetized birds at different stages of vocal development. These studies have demonstrated selectivity for different song types that provide neural signatures of learning. In contrast to the ease of obtaining responses to song playback in anesthetized birds, song-evoked responses in awake birds are greatly reduced or absent, indicating that behavioral state is an important determinant of neural responsivity. Song-evoked responses can be elicited in sleeping as well as anesthetized zebra finches, and the selectivity of responses to song playback in adult birds tends to be highly similar between anesthetized and sleeping states, encouraging the idea that anesthesia and sleep are highly similar. In contrast to that idea, we report evidence that cortical responses to song playback in juvenile zebra finches (Taeniopygia guttata) differ greatly between sleep and urethane anesthesia. This finding indicates that behavioral states differ in sleep versus anesthesia and raises questions about relationships between developmental changes in sleep activity, selectivity for different song types, and the neural substrate for vocal learning.SignificancePatterns of spiking activity based on electrophysiological recordings in many different taxa are known to be heavily dependent on behavioral state. Neural activity patterns are frequently similar between sleep and anesthesia, which has encouraged the idea that similar states characterize sleep and anesthesia. Based on comparisons across studies, we report that activity patterns are highly dissimilar between sleep and urethane anesthesia in a cortical region of juvenile songbirds. These data argue against the idea that similar behavioral states are achieved in sleep versus anesthesia.
Effects of larval diet on the life-history traits and phenotypic expression of pyrethroid resistance in the major malaria vectorAnopheles gambiae s.s.
Authors: Sovegnon Pierre Marie , Fanou Marie Joelle , Akoton Romaric , Djihinto Oswald Yédjinnavênan , Lagnika Hamirath Odée , Agonhossou Romuald , Djogbénou Luc Salako 
Published: January 2022

AbstractThe success achieved in reducing malaria transmission by vector control is threatened by insecticide resistance. To strengthen the current vector control programmes, the non-genetic factors underlying the emergence of insecticide resistance inAnophelesvectors and its widespread need to be explored. This study aimed to assess the effects of larval diet on some life-history traits and pyrethroid-insecticide susceptibility ofAnopheles gambiae s.s.Three (3)An. gambiaestrains, namely Kisumu (insecticide susceptible), AcerKis (homozygousace-1RG119S resistant) and KisKdr (homozygouskdrRL1014F resistant) were fed with three different diets (low, medium, and high) of TetraMin®Baby fish food. Pre-imaginal developmental time, larval mortality, adult emergence rate and female wing length were measured. Mosquito females were exposed to insecticide-treated net (ITN) PermaNet 2.0 and PermaNet 3.0.In the threeAn. gambiaestrains, significant differences in adult emergence rates (F= 1054.2;df= 2;p<0.01), mosquito wing length (F= 970.5;df= 2;p<0.01) and adult survival post insecticide exposure (χ2= 173;df= 2;p<0.01), were noticed among the three larval diets. Larvae fed with the low food diets took more time to develop, were smaller at emergence and displayed a short lifespan, while the specimens fed with a high regime developed faster and into big adults. Although being fed with a high diet, none ofAn. gambiaestrain harbouring thekdrRandace-1Rallele survived 24 hours after exposure against PermaNet 3.0.This study showed that variation in the larval diet significantly impactsAn. gambiaelife-history traits such as larval mortality and developmental time, adult wing length, and female susceptibility to pyrethroid insecticides. Further investigations through field-based studies would allow an in-depth understanding of the implications of these non-genetic parameters on the physiological traits of malaria vectors and consequently improve resistance management.
Parasitized or non-parasitized, why? A study of factors influencing tick burden in roe deer neonates
Authors: Bariod Léa , Saïd Sonia , Calenge Clément , Chabot Stéphane , Badeau Vincent , Bourgoin Gilles 
Published: January 2022

AbstractIxodes ricinus, the most common species of tick in Europe, is known to transmit major pathogens to animals and humans such asBabesiaspp. orBorreliaspp.. Its abundance and distribution have been steadily increasing in Europe during recent decades, due to global environmental changes. Indeed, as ticks spend most of their life in the environment, their activity and life cycle are highly dependent on environmental conditions, and therefore on climate or habitat changes. Simultaneously, wild ungulates have expanded their range and increased dramatically in abundance worldwide, in particular roe deer (Capreolus capreolus), have allowed tick populations to grow and spread. Currently, tick infestation on newborn wild ungulates is poorly documented. However, newborn ungulates are considered more sensitive to tick bites and pathogen transmission because of their immature immune system. Thus, improving knowledge about the factors influencing tick infestation on newborns is essential to better understand their health risks. This study was conducted at Trois-Fontaines forest, Champagne-Ardenne, France (1992-2018). Based on a long-term monitoring of roe deer fawns, we used a novel Bayesian model of the infestation of fawns to identify which biotic or abiotic factors are likely to modify the level of infestation by ticks of 965 fawns over time. We show that tick burden increased faster during the first days of life of the fawns and became constant when fawns were 5 days old and more, which could be explained by the depletion of questing ticks or the turnover of ticks feeding on fawns. Moreover, the humidity, which favors tick activity, was weakly positively related to the tick burden. Our results demonstrate that tick infestation was highly variable among years, particularly between 2000 and 2009. We hypothesize that this results from a modification of habitat caused by hurricane Lothar.
Viscoelastic properties of suspended cells measured with shear flow deformation cytometry
Authors: Gerum Richard , Mirzahossein Elham , Eroles Mar , Elsterer Jennifer , Mainka Astrid , Bauer Andreas , Sonntag Selina , Winterl Alexander , Bartl Johannes , Fischer Lena , Abuhattum Shada , Goswami Ruchi , Girardo Salvatore , Guck Jochen , Schrüfer Stefan , Ströhlein Nadine , Nosratlo Mojtaba , Herrmann Harald , Schultheis Dorothea , Rico Felix , Müller Sebastian , Gekle Stephan , Fabry Ben 
Published: January 2022

ABSTRACTNumerous cell functions are accompanied by phenotypic changes in viscoelastic properties, and measuring them can help elucidate higher-level cellular functions in health and disease. We present a high-throughput, simple and low-cost microfluidic method for quantitatively measuring the elastic (storage) and viscous (loss) modulus of individual cells. Cells are suspended in a high-viscosity fluid and are pumped with high pressure through a 5.8 cm long and 200 µm wide microfluidic channel. The fluid shear stress induces large, near ellipsoidal cell deformations. In addition, the flow profile in the channel causes the cells to rotate in a tank-treading manner. From the cell deformation and tank treading frequency, we extract the frequency-dependent viscoelastic cell properties based on a theoretical framework developed by R. Roscoe1 that describes the deformation of a viscoelastic sphere in a viscous fluid under steady laminar flow. We confirm the accuracy of the method using atomic force microscopy-calibrated polyacrylamide beads and cells. Our measurements demonstrate that suspended cells exhibit power-law, soft glassy rheological behavior that is cell cycle-dependent and mediated by the physical interplay between the actin filament and intermediate filament networks.