Community Evaluation of Glycoproteomics Informatics Solutions Reveals High-Performance Search Strategies of Serum N- and O-Glycopeptide Data Glycoproteome profiling (glycoproteomics) is a powerful yet analytically challenging research tool. The complex tandem mass spectra generated from glycopeptide mixtures require sophisticated analysis pipelines for structural determination. Diverse software aiding the process have appeared, but their relative performance remains untested. Conducted through the HUPO Human Proteome Project - Human Glycoproteomics Initiative, this community study, comprising both developers and users of glycoproteomics software, evaluates the performance of informatics solutions for system-wide glycopeptide analysis. Mass spectrometry-based glycoproteomics datasets from human serum were shared with all teams. The relative team performance for N- and O-glycopeptide data analysis was comprehensively established and validated through orthogonal performance tests. Excitingly, several high-performance glycoproteomics informatics solutions were identified. While the study illustrated that significant informatics challenges remain, as indicated by a high discordance between annotated glycopeptides, lists of high-confidence (consensus) glycopeptides were compiled from the standardised team reports. Deep analysis of the performance data revealed key performance-associated search variables and led to recommendations for improved "high coverage" and "high accuracy" glycoproteomics search strategies. This study concludes that diverse software for comprehensive glycopeptide data analysis exist, points to several high-performance search strategies, and specifies key variables that may guide future software developments and assist informatics decision-making in glycoproteomics.
In vitro synthesis and reconstitution using mammalian cell-free lysates enables the systematic study of the regulation of LINC complex assembly Membrane proteins perform numerous important functions in cells and tissues. Approximately 20% of the human genome encodes for membrane proteins, which represent the majority of targets for clinically relevant small molecules. Consequently, understanding their structure and structure-function relationships is a fundamental problem in biomedical research. Given the difficulties inherent to performing mechanistic biochemical and biophysical studies of membrane proteins in vitro, we previously developed a facile HeLa cell-based cell-free expression (CFE) system that enables the efficient reconstitution of full-length (FL) functional membrane proteins in supported lipid bilayers. Despite having shown the directional reconstitution of CFE-synthesized FL inner nuclear membrane SUN proteins (i.e. SUN1 and SUN2), which directly interact with outer nuclear membrane KASH proteins within the nuclear envelope lumen to form linker of nucleoskeleton and cytoskeleton (LINC) complexes that mechanically couple the cytoskeleton and nucleus, the mechanism underlying regulated LINC complex assembly remains unclear. Here, we provide evidence that suggests that the reconstitution of CFE-synthesized FL membrane proteins in supported lipid bilayers occurs primarily through the fusion of endoplasmic reticulum-derived microsomes present within our CFE reactions with our supported lipid bilayers. In addition, we demonstrate the ease with which our synthetic biology platform can be used to investigate the impact of the chemical environment (e.g. calcium ions and redox state) on the ability of CFE-synthesized FL SUN proteins reconstituted in supported lipid bilayers to interact with the luminal domain of the KASH protein nesprin-2. Moreover, we use our platform to study the molecular requirements for the homo- and hetero-typic interactions that can occur between SUN1 and SUN2. Finally, we show that our platform can be used to simultaneously reconstitute three different CFE-synthesized FL membrane proteins in a single supported lipid bilayer. Overall, these results establish our HeLa cell-based CFE and supported lipid bilayer reconstitution platform as a powerful tool for performing mechanistic dissections of the oligomerization and function of FL membrane proteins in vitro. While our platform is not a substitute for cell-based studies of membrane protein biochemistry and function, it does provide important mechanistic insights into the biology of difficult-to-study membrane proteins.

ABSTRACT GRAPHIC O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY
Citation needed? Wikipedia and the COVID-19 pandemic BackgroundWith the COVID-19 pandemics outbreak, millions flocked to Wikipedia for updated information. Amid growing concerns regarding an "infodemic", ensuring the quality of information is a crucial vector of public health. Investigating if and how Wikipedia remained up to date and in line with science is key to formulating strategies to counter misinformation. Using citation analyses, we asked: which sources informed Wikipedias COVID-19-related articles before and during the pandemics first wave (January-May 2020).

ResultsWe found that coronavirus-related articles referenced trusted media sources and high-quality academic research. Moreover, despite a surge in COVID-19 preprints, Wikipedia had a clear preference for open-access studies published in respected journals and made little use of preprints. Building a timeline of English COVID-19 articles from 2001-2020 revealed a nuanced trade-off between quality and timeliness. It further showed how preexisting articles on key topics related to the virus created a framework for integrating new knowledge. Supported by a rigid sourcing policy, this "scientific infrastructure" facilitated contextualization and regulated the influx of new information. Lastly, we constructed a network of DOI-Wikipedia articles, which showed the shifting landscape of pandemic-related knowledge on Wikipedia and how academic citations create a web of shared knowledge supporting topics like COVID-19 vaccine development.

ConclusionsUnderstanding how scientific research interacts with the digital knowledge-sphere during the pandemic provides insight into how Wikipedia can facilitate access to science. It also reveals how, aided by what we term its "citizen encyclopedists", it successfully fended off COVID-19 disinformation and how this unique model may be deployed in other contexts. Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. Robust and Remote Measurement of Heart Rate Based on A Surveillance Camera Arrhythmia is a marked symptom of many cardiovascular diseases (CVDs). The accurate and timely detection of heart rate can greatly reduce the harm of arrhythmia to people. However, it is still a challenge to robustly and remotely measure heart rate in daily life due to the changing environmental conditions during measurement, such as the varying light intensity, the movement of people, and the uncertain distance between the sensor to people. In this study, we propose a method to accurately measure human heart rate within a distance of 4.5 meters under different light intensities by simply using a surveillance camera. After a 20-second color video of a persons hand is captured by the camera, a method based on the Fast Fourier Transform (FFT) algorithm is designed to extract the blood volume pulse wave to calculate the heart rate. According to the comparison between the real heart rate and results measured by electrocardiography (ECG), the proposed method achieves an accuracy of 95.8% when the measurement is performed within a distance of 4.5 meters and 90% when within 5.0 meters. Our experiments show that when the illuminance varies between 100-1000 lux (lighting level indoor), we still get the correct results. Our experiments also demonstrate that the proposed method accurately obtain heart rate even when the light intensity is below 32 lux (300-500 lux in a workplace environment). The methods strong adaptability to changing environmental conditions makes it applicable to many scenarios, such as homes of the elderly, classrooms, and other public spaces. Host Chitinase 3-like-1 is a Universal Therapeutic Target for SARS-CoV-2 Viral Variants in COVID 19 COVID 19 is the disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2; SC2) which has caused a world-wide pandemic with striking morbidity and mortality. Evaluation of SC2 strains demonstrated impressive genetic variability and many of these viral variants are now defined as variants of concern (VOC) that cause enhanced transmissibility, decreased susceptibility to antibody neutralization or therapeutics and or the ability to induce severe disease. Currently, the delta ({delta}) and omicron (o) variants are particularly problematic based on their impressive and unprecedented transmissibility and ability to cause break through infections. The delta variant also accumulates at high concentrations in host tissues and has caused waves of lethal disease. Because studies from our laboratory have demonstrated that chitinase 3-like-1 (CHI3L1) stimulates ACE2 and Spike (S) priming proteases that mediate SC2 infection, studies were undertaken to determine if interventions that target CHI3L1 are effective inhibitors of SC2 viral variant infection. Here we demonstrate that CHI3L1 augments epithelial cell infection by pseudoviruses that express the alpha, beta, gamma, delta or omicron S proteins and that the CHI3L1 inhibitors anti-CHI3L1 and kasugamycin inhibit epithelial cell infection by these VOC pseudovirus moieties. Thus, CHI3L1 is a universal, VOC-independent therapeutic target in COVID 19. First evidence of in vitro cytotoxic effects of marine microlitter on Merluccius merluccius and Mullus barbatus, two Mediterranean commercial fish species Marine litter is composed mainly of plastics and is recognized as a serious threats to marine ecosystems. Ecotoxicological approaches have started elucidating the potential severity of microplastics (MPs) in controlled laboratory studies with pristine materials but no information exist on marine environmental microlitter as a whole. Here, we characterized the litter in the coastal Northern Tyrrhenian sea and in the stomach of two fish species of socio-economic importance, and exposed primary cell cultures of mucosal and lymphoid organs to marine microlitter for evaluating possible cytotoxic effects. An average of 0.30 {+/-} 0.02 microlitter items m-3 was found in water samples. FT-IR analysis revealed that plastic particles, namely HDPE, polyamide and polypropylene were present in 100% and 83.3% of Merluccius merluccius and Mullus barbatus analyzed, which overall ingested 14.67 {+/-} 4.10 and 5.50 {+/-} 1.97 items/individual, respectively. Moreover, microlitter was confirmed as a vector of microorganisms. Lastly, the apical end-point of viability was found to be significantly reduced in splenic cells exposed in vitro to two microlitter conditions. Considering the role of the spleen in the mounting of adaptive immune responses, our results warrant more in-depth investigations for clarifying the actual susceptibility of these two species to anthropogenic microlitter.

HighlightsO_LI0.30 {+/-} 0.02 microlitter items m-3 were found at the surface of coastal Northern Tyrrhenian sea
C_LIO_LI14.67 {+/-} 4.10 and 5.50 {+/-} 1.97 items/individual were retrieved from the stomach of hakes and mullets
C_LIO_LIThe ingested microlitter contained plastic items
C_LIO_LIMicrolitter was validated as a carrier of bacteria, fungi and flagellates
C_LIO_LISplenic cells exposed to two microlitter conditions for 72 hours suffered cytotoxicity
C_LI Human inhalable antibody fragments neutralizing SARS-CoV-2 variants for COVID-19 therapy As of October 2021, coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains a global emergency, and novel therapeutics are urgently needed. Here we describe human single chain variable fragment (scFv) antibodies (76clAbs) that block an epitope of the SARS-CoV-2 spike protein essential for ACE2-mediated entry into cells. 76clAbs neutralize the delta variant and other variants being monitored (VBMs) and inhibit spike-mediated pulmonary cell-cell fusion, a critical feature of COVID-19 pathology. In two independent animal models, intranasal administration counteracted the infection. Due to high efficiency, remarkable stability, resilience to nebulization and low production cost, 76clAbs may become a relevant tool for rapid, self-administrable early intervention in SARS-CoV-2-infected subjects independently of their immune status. Smell-induced gamma oscillations in human olfactory cortex are required for accurate perception of odor identity Studies of neuronal oscillations have contributed substantial insight into the mechanisms of visual, auditory and somatosensory perception. However, progress in such research in the human olfactory system has lagged behind. As a result, the electrophysiological properties of the human olfactory system are poorly understood, and in particular, whether stimulus-driven high frequency oscillations play a role in odor processing is unknown. Here, we used direct intracranial recordings from human piriform cortex during an odor identification task to show that three key oscillatory rhythms are an integral part of the human olfactory cortical response to smell: odor induces theta, beta and gamma rhythms in human piriform cortex. We further show that these rhythms have distinct relationships with perceptual behavior. Odor-elicited gamma oscillations occur only during trials in which the odor is accurately perceived, and features of gamma oscillations predict odor identification accuracy, suggesting they are critical for odor identity perception in humans. We also found that the amplitude of high-frequency oscillations is organized by the phase of low frequency signals shortly following sniff onset, only when odor is present. Our findings reinforce previous work on theta oscillations, suggest that gamma oscillations in human piriform cortex are important for perception of odor identity, and constitute a robust identification of the characteristic electrophysiological response to smell in the human brain. Future work will determine whether the distinct oscillations we identified reflect distinct perceptual features of odor stimuli. Loci for prediction of penicillin and tetracycline susceptibility in Neisseria gonorrhoeae: a genome wide association study BackgroundWhile Neisseria gonorrhoeae poses an urgent public health threat because of increasing antimicrobial resistance, much of the circulating population remains susceptible to historical treatment regimens. Point-of-care diagnostics that report susceptibility could allow for reintroduction of these regimens, but development of such diagnostics has been limited to ciprofloxacin, for which susceptibility can be predicted from a single locus.

MethodsWe assembled a dataset of 12,045 N. gonorrhoeae genomes with phenotypic resistance data for tetracycline (n = 3,611) and penicillin (n = 6,935). Using conditional genome wide association studies (GWAS), we sought to define genetic variants associated with susceptibility to penicillin and tetracycline. We evaluated the sensitivity and specificity of these variants for predicting susceptibility and non-resistance in our collection of gonococcal genomes.

FindingsIn our conditional penicillin GWAS, the presence of a genetic variant defined by a non-mosaic penA allele without an insertion at codon 345 was significantly associated with penicillin susceptibility and had the highest negative effect size of significant variants (p = 5.0 x 10-14, {beta} = -2.5). In combination with the absence of blaTEM, this variant predicted penicillin susceptibility with high specificity (99.8%) and modest sensitivity (36.7%). For tetracycline, the wild type allele at rpsJ codon 57, encoding valine, was significantly associated with tetracycline susceptibility (p = 5.6 x 10-16, {beta} = -1.6) after conditioning on the presence of tetM. The combination of rpsJ codon 57 allele and tetM absence predicted tetracycline susceptibility with high specificity (97.2%) and sensitivity (88.7%).

InterpretationAs few as two genetic loci can predict susceptibility to penicillin and tetracycline in N. gonorrhoeae with high specificity. Molecular point-of-care diagnostics targeting these loci have the potential to increase available treatments for gonorrhea.

FundingNational Institute of Allergy and Infectious Diseases, the National Science Foundation, and the Smith Family Foundation

Research in ContextO_ST_ABSEvidence before this studyC_ST_ABSWe searched PubMed with the terms "Neisseria gonorrhoeae" and "diagnostic" or "assay" plus "penicillin" or "tetracycline" for reports in any language published up to July 1, 2021. We additionally searched for "Neisseria gonorrhoeae" and "genome wide association study". We found that previously proposed molecular diagnostics for penicillin and tetracycline susceptibility either exclusively focused on plasmid-mediated resistance (i.e., targeting blaTEM or tetM) or did not include variants in genes encoding antibiotic targets (e.g., did not include penA or rpsJ). Targets for molecular surveillance have focused on resistance-associated alleles rather than susceptibility-associated alleles. We did not find any previous penicillin or tetracycline conditional genome wide association studies (GWAS) in N. gonorrhoeae.

Added value of this studyTo identify targets for molecular diagnostics that predict penicillin and tetracycline susceptibility, we conducted GWAS conditioning on the presence of plasmid-mediated resistance determinants to detect chromosomal loci with the highest association with susceptibility. We discovered a sequence (penA_01) that differentiates susceptible isolates from those with a resistance-associated insertion at codon 345 and from those with mosaic penA alleles, which is associated with penicillin susceptibility. We also found that rpsJ codon 57 was the chromosomal locus contributing the most to tetracycline susceptibility. The combination of these chromosomal loci and the absence of plasmid encoded determinants predicts penicillin and tetracycline susceptibility with high specificity in both a large global collection of N. gonorrhoeae and a validation dataset consisting of recently published genomes from CDCs Gonococcal Isolate Surveillance Program (GISP) surveillance collected in 2018.

Implications of all the available evidenceThe chromosomal loci penA_01 and rpsJ codon 57 in combination with plasmid loci blaTEM and tetM are candidates for the development of point-of-care molecular diagnostics for penicillin and tetracycline susceptibility. The loci may be combined with the currently available ciprofloxacin susceptibility diagnostics to predict susceptibility to multiple antibiotics. Additionally, our study suggests that conditional GWAS focused on variants associated with susceptibility may be a promising approach to identify minimal sets of loci for molecular diagnostics and surveillance. Genome editing in the unicellular holozoan Capsaspora owczarzaki suggests a premetazoan role for the Hippo pathway in multicellular morphogenesis Animal development is mediated by a surprisingly small set of canonical signaling pathways such as Wnt, Hedgehog, TGF-beta, Notch and Hippo pathways. Although once thought to be present only in animals, recent genome sequencing has revealed components of these pathways in the closest unicellular relatives of animals. These findings raise questions about the ancestral functions of these developmental pathways and their potential role in the emergence of animal multicellularity. Here, we provide the first functional characterization of any of these developmental pathways in unicellular organisms by developing techniques for genetic manipulation in Capsaspora owczarzaki, a close unicellular relative of animals that displays aggregative multicellularity. We then use these tools to characterize the Capsaspora ortholog of the Hippo signaling nuclear effector YAP/TAZ/Yorkie (coYki), a key regulator of tissue size in animals. In contrast to what might be expected based on studies in animals, we show that coYki is dispensable for cell proliferation but regulates cytoskeletal dynamics and the three-dimensional (3D) shape of multicellular structures. We further demonstrate that the cytoskeletal abnormalities of individual coYki mutant cells underlie the abnormal 3D shape of coYki mutant aggregates. Taken together, these findings implicate an ancestral role for the Hippo pathway in cytoskeletal dynamics and multicellular morphogenesis predating the origin of animal multicellularity, which was co-opted during evolution to regulate cell proliferation. Superinfection exclusion: a viral strategy with short-term benefits and long-term drawbacks Viral superinfection occurs when multiple viral particles subsequently infect the same host. In nature, several viral species are found to have evolved diverse mechanisms to prevent superinfection (superinfection exclusion) but how this strategic choice impacts the fate of mutations in the viral population remains unclear. Using stochastic simulations, we find that genetic drift is suppressed when superinfection occurs, thus facilitating the fixation of beneficial mutations and the removal of deleterious ones. Interestingly, we also find that the competitive (dis)advantage associated with variations in life history parameters is not necessarily captured by the viral growth rate for either infection strategy. Putting these together, we then show that a mutant with superinfection exclusion will easily overtake a superinfecting population even if the latter has a much higher growth rate. Our findings suggest that while superinfection exclusion can negatively impact the long-term adaptation of a viral population, in the short-term it is ultimately a winning strategy.

Author summaryViral social behaviour has recently been receiving increasing attention in the context of ecological and evolutionary dynamics of viral populations. One fascinating and still relatively poorly understood example is superinfection or co-infection, which occur when multiple viruses infect the same host. Among bacteriophages, a wide range of mechanisms have been discovered that enable phage to prevent superinfection (superinfection exclusion) even at the cost of using precious resources for this purpose. What is the evolutionary impact of this strategic choice and why do so many phages exhibit this behaviour? Here, we conduct an extensive simulation study of a phage population to address this question. In particular, we investigate the fate of viral mutations arising in an environment with a constant supply of bacterial hosts designed to mimic a "turbidostat," as these are increasingly being used in laboratory evolution experiments. Our results show that allowing superinfection in the long-term yields a population which is more capable of adapting to changes in the environment. However, when in direct competition, mutants capable of preventing superinfection experience a very large advantage over their superinfecting counterparts, even if this ability comes at a significant cost to their growth rate. This indicates that while preventing superinfection can negatively impact the long-term prospects of a viral population, in the short-term it is ultimately a winning strategy. How much does it take to be old? Modelling the time since the last harvesting to infer the distribution of overmature forests in France AimThe distribution of overmature forests in metropolitan France is poorly known, with only a few well-studied prominent sites, and has never been evaluated countrywide. Here, we modelled French forest reserves time since the last harvesting operation - a proxy for forest maturity - then inferred the current statistical distribution of overmature forests (i.e. forests over 50 years without harvesting) in France.

LocationMetropolitan France

MethodsWe used inventories from forest reserves and managed forests to calibrate a generalised linear mixed model explaining the time since the last harvesting with selected structural attributes and environmental variables. We then projected this model on the independent National Forest Inventory dataset. We thus obtained an updated estimation of the proportion and a rough distribution of overmature forest stands in metropolitan France.

ResultsWe found that high basal area of very large trees, high volumes of standing and downed deadwood, high diversity of tree-related microhabitats and more marginally diversity of decay stages best characterized the time since the last harvesting. Volumes of stumps and high density of coppices translating legacy of past forest management also distinguished more overmature plots. Our projection yielded an estimated 3% of French forests over 50 years without harvesting mostly located in more inaccessible areas (i.e. mountainous areas) and a promising proportion of future overmature forests if left unharvested.

Main conclusionsOur study showed that the time since the last harvesting is a good proxy for a combination of stand structure attributes key in characterising overmature temperate forests. It gives the first robust statistical estimate of the proportion of overmature forests and may serve to report on their status in metropolitan France. Our method could be implemented at a larger spatial scale, notably in countries with accessible National Forest Inventory and calibration data, to produce indicators at international level. Genome-wide association study suggests an independent genetic basis of zinc and cadmium concentrations in fresh sweet corn kernels Despite being one of the most consumed vegetables in the United States, the elemental profile of sweet corn (Zea mays L.) is limited in its dietary contributions. To address this through genetic improvement, a genome-wide association study was conducted for the concentrations of 15 elements in fresh kernels of a sweet corn association panel. In concordance with mapping results from mature maize kernels, we detected a probable pleiotropic association of zinc and iron concentrations with nicotianamine synthase5 (nas5), which purportedly encodes an enzyme involved in synthesis of the metal chelator nicotianamine. Additionally, a pervasive association signal was identified for cadmium concentration within a recombination suppressed region on chromosome 2. The likely causal gene underlying this signal was heavy metal ATPase3 (hma3), whose counterpart in rice, OsHMA3, mediates vacuolar sequestration of cadmium and zinc in roots, whereby regulating zinc homeostasis and cadmium accumulation in grains. In our association panel, hma3 associated with cadmium but not zinc accumulation in fresh kernels. This finding implies that selection for low cadmium will not affect zinc levels in fresh kernels. Although less resolved association signals were detected for boron, nickel, and calcium, all 15 elements were shown to have moderate predictive abilities via whole-genome prediction. Collectively, these results help enhance our genomics-assisted breeding efforts centered on improving the elemental profile of fresh sweet corn kernels. Mathematical Modeling and Analysis of Mitochondrial Retrograde Signaling Dynamics Mitochondria, semi-autonomous eukaryotic organelles, participate in energy production and metabolism, making mitochondrial quality control crucial. As most mitochondrial proteins are encoded by nuclear genes, quality control depends on proper mitochondria-nucleus communication, designated mitochondrial retrograde signaling. Early studies focused on retrograde signaling participants and specific gene knockouts. However, mitochondrial signal modulation remains elusive. Using yeast, we simulated signal propagation following mitochondrial damage and proposed a mathematical model based ordinary differential equations. Mitochondrial retrograde signaling decisions were described using a Boolean model. Dynamics were analyzed through an ordinary differential equation-based model and extended to evaluate the model response to noisy damage signals. Simulation revealed localized protein concentration dynamics, including waveforms, frequency response, and robustness under noise. Retrograde signaling is bistable with three localized steady states, and increased damage compromises robustness. We elucidated mitochondrial retrograde signaling, thus providing a basis for drug design against yeast and fungi.

Author SummaryThe yeast RTG pathway regulates mitochondrial metabolism and mitochondrial quality through passing mitochondrial signal to the nucleus to modulate gene expressions. Using microscopic data of RTG proteins translocation from cytosol to nucleus, the parameters were found by fitting 16 knockout conditions simulated from the proposed differential equation-based model. Further, through dose response, ultrasensitivity, frequency response and noise interference, we demonstrated a switch-like property of RTG pathway activation, a capacity charging property of RTG protein translocation, a low pass filter property of signals in different frequencies, and noise amplification due to mitochondrial signal. The proposed mathematical model enables us to understand the dynamics and mechanisms in mitochondrial retrograde signaling in yeast and provides potential antifungal treatment strategies. A Hierarchy of Biomolecular Proportional-Integral-Derivative Feedback Controllers for Robust Perfect Adaptation and Dynamic Performance Proportional-Integral-Derivative (PID) feedback controllers have been the most widely used controllers in industry for almost a century due to their good performance, simplicity, and ease of tuning. Motivated by their success in various engineering disciplines, PID controllers recently found their way into synthetic biology, where the design of feedback molecular control systems has been identified as an important goal. In this paper, we consider the mathematical realization of PID controllers via biomolecular interactions. We propose an array of topologies that offer a compromise between simplicity and high performance. We first demonstrate that different Proportional-Integral (PI) controllers exhibit different capabilities for enhancing the dynamics and reducing variance (cell-to-cell variability). Next, we introduce several derivative controllers that are realized based on incoherent feedforward loops acting in a feedback configuration. Alternatively, we show that differentiators can be realized by placing molecular integrators in a negative feedback loop--an arrangement that can then be augmented by PI components to yield PID feedback controllers. We demonstrate that the derivative component can be exploited for enhancing system stability, dramatically increasing the molecular control systems dynamic performance, and for reducing the noise effect on the output of interest. The PID controller features are established through various deterministic and stochastic analyses as well as numerical simulations. Finally, we provide an experimental demonstration using a recently developed hybrid setup, the cyberloop, where the controller is implemented in silico to control a biological genetic circuit in vivo. The large array of novel biomolecular PID controllers introduced here forms a basis for the design and construction of advanced high-performance biomolecular control systems that robustly regulate the dynamics of living systems. Fatal neuroinvasion and SARS-CoV-2 tropism in K18-hACE2 mice is partially independent on hACE2 expression Animal models recapitulating distinctive features of severe COVID-19 are critical to enhance our understanding of SARS-CoV-2 pathogenesis. Transgenic mice expressing human angiotensin-converting enzyme 2 (hACE2) under the cytokeratin 18 promoter (K18-hACE2) represent a lethal model of SARS-CoV-2 infection. The precise mechanisms of lethality in this mouse model remain unclear. Here, we evaluated the spatiotemporal dynamics of SARS-CoV-2 infection for up to 14 days post-infection. Despite infection and moderate pneumonia, rapid clinical decline or death of mice was invariably associated with viral neuroinvasion and direct neuronal injury (including brain and spinal neurons). Neuroinvasion was observed as early as 4 dpi, with virus initially restricted to the olfactory bulb supporting axonal transport via the olfactory neuroepithelium as the earliest portal of entry. No evidence of viremia was detected suggesting neuroinvasion occurs independently of entry across the blood brain barrier. SARS-CoV-2 tropism was not restricted to ACE2-expressing cells (e.g., AT1 pneumocytes), and some ACE2-positive lineages were not associated with the presence of viral antigen (e.g., bronchiolar epithelium and brain capillaries). Detectable ACE2 expression was not observed in neurons, supporting overexpression of ACE2 in the nasal passages and neuroepithelium as more likely determinants of neuroinvasion in the K18-hACE2 model. Although our work incites caution in the utility of the K18-hACE2 model to study global aspects of SARS-CoV-2 pathogenesis, it underscores this model as a unique platform for exploring the mechanisms of SARS-CoV-2 neuropathogenesis that may have clinical relevance acknowledging the growing body of evidence that suggests COVID-19 may result in long-standing neurologic consequences.

IMPORTANCECOVID-19 is predominantly a respiratory disease caused by SARS-CoV-2 that has infected more than 191 million people with over 4 million fatalities (2021-07-20). The development of animal models recapitulating distinctive features of severe COVID-19 is critical to enhancing our understanding of SARS-CoV-2 pathogenesis and in the evaluation of vaccine and therapeutic efficacy. Transgenic mice expressing human angiotensin-converting enzyme 2 (hACE2) under the cytokeratin 18 promoter (K18-hACE2) represent a lethal model of SARS-CoV-2 infection. Here, we show lethality of this model is invariably associated with viral neuroinvasion linked with viral replication and assembly. Importantly, pneumonia albeit invariably present was generally moderate with the absence of culturable infectious virus at peak neuroinvasion. The dynamics of viral neuroinvasion and pneumonia were only partially dependent on hACE2. Overall, this study provides an in-depth sequential characterization of the K18-hACE2 model following SARS-CoV-2 infection, highlighting its significance to further study the mechanisms of SARS-CoV-2 neuropathogenesis. Do males know? Evidence-driven rainmaking mating strategy by male bonobos to meet the fertile window of females Female bonobos exhibit prolonged receptivity, one suggested function of which is to lower male mating competition. However, it is questionable whether easier access to receptive females can reduce male-male competition, given the exclusive nature of male reproductive success. We tested whether males could determine the fertile phase of females. We found that ovulation probability predicted male mating effort. High-ranking males copulated with females with higher fertility, and male-male agonistic interactions increased when there were fertile females in the party. When there were multiple females with maximal swelling, males concentrated their mating effort on females with an older infant whose maximal swelling started earlier, and they continued mating efforts until detumescence (rainmaking). These findings suggest that male bonobos distinguish between fertile and non-fertile phases of females and that having more receptive females in the party does not reduce male-male competition for fertile females.

TeaserMales use the rainmakers rule to meet the periovulatory phase of the female bonobo for better reproductive success. Molecular Dynamics Simulations of Phytopthora sojae Avirulence Factor 5, PsAvh5, Define Membrane Binding, Inositolphosphate-3'-phosphate Interactions and Protein Mechanics Phytopthora Avirulence proteins are a primary target for development of rational chemical and biological control of some of the most devastating plant pathogens. Despite the sequencing of entire genomes, and characterization of many of these proteins at the chemical level, many questions remain regarding actual chemical and biological interactions involved. In addition, disputed roles of ligands, such as Inositolphosphate-3-phosphate and amino acids of important function remain unclear. To address some of these issues, we developed molecular models from structural elements and published data for Phytopthora sojae avirulence protein 5. Molecular dynamics simulations are used to study protein function, interactions involved primarily with lipids and membranes, and inositol derivatives. Our findings indicate that the protein is stable as a monomer, and in a dimeric form. Also, that these proteins interact with Inositolphosphate-3phosphate as a necessary membrane element, in binding. We identified several amino acids of importance, additional to defining the mechanical features of the protein within the binding process to different membranes. A high affinity, comparable to other membrane surface binding molecules of -219.54 Kcal for the dimer, and -176.61 for the monomer were determined. With either form, we found the inositolphosphate-3-phosphate to be essential in the membrane binding process. Our findings answer some of the debated questions while creating a point to further test avirulence proteins in general for functional aspects. Additionally, the structures and data can be utilized to provide a better starting point for rational design approaches to control this pathogen. GPI-anchored FGF directs cytoneme-mediated bidirectional signaling to self-regulate tissue-specific dispersion How signaling proteins generate a multitude of information to organize tissue patterns is critical to understanding morphogenesis. In Drosophila, FGF produced in wing-disc cells regulates the development of the disc-associated air-sac-primordium (ASP). Here, we show that FGF is Glycosylphosphatidylinositol-anchored to the producing cell surface and that this modification both inhibits free FGF secretion and activates target-specific bidirectional FGF-FGFR signaling through cytonemes. FGF-source and recipient ASP cells extend cytonemes that present FGF and FGFR on their surfaces and reciprocally recognize each other over distance by contacting through CAM-like FGF-FGFR binding. Contact-mediated FGF-FGFR binding induces bidirectional signaling, which, in turn, promotes ASP and source cells to polarize cytonemes toward each other and reinforce signaling contacts. Subsequent un-anchoring of FGFR-bound-FGF from the source cell membrane dissociates cytoneme contacts and delivers FGF target-specifically to ASP cytonemes for paracrine functions. Thus, GPI-anchored FGF organizes both source and recipient cells and self-regulates its cytoneme-mediated tissue-specific dispersion and signaling. Protein-protein docking with large-scale backbone flexibility using coarse-grained Monte-Carlo simulations Most of the protein-protein docking methods treat proteins as almost rigid objects. Only the side-chains flexibility is usually taken into account. The few approaches enabling docking with a flexible backbone typically work in two steps, in which the search for protein-protein orientations and structure flexibility are simulated separately. In this work, we propose a new straightforward approach for docking sampling. It consists of a single simulation step during which a protein undergoes large-scale backbone rearrangements, rotations, and translations. Simultaneously, the other protein exhibits small backbone fluctuations. Such extensive sampling was possible using the CABS coarse-grained protein model and Replica Exchange Monte Carlo dynamics at a reasonable computational cost. In our proof-of-concept simulations of 62 protein-protein complexes, we obtained acceptable quality models for a significant number of cases. Action at a distance: Defects in division plane positioning in the root meristematic zone affect cell organization in the differentiation zone Cell division plane orientation is critical for plant and animal development and growth. TANGLED1 (TAN1) and AUXIN-INDUCED-IN-ROOT-CULTURES9 (AIR9) are division-site localized microtubule-binding proteins required for division plane positioning. tan1 and air9 Arabidopsis thaliana single mutants have minor or no noticeable phenotypes but the tan1 air9 double mutant has synthetic phenotypes including stunted growth, misoriented divisions, and aberrant cell-file rotation in the root differentiation zone. These data suggest that TAN1 plays a role in nondividing cells. To determine whether TAN1 is required in elongating and differentiating cells in the tan1 air9 double mutant, we limited its expression to actively dividing cells using the G2/M-specific promoter of the syntaxin KNOLLE (pKN:TAN1-YFP). Unexpectedly, in addition to rescuing division plane defects, pKN:TAN1-YFP rescued root growth and the root differentiation zone cell file rotation defects in the tan1 air9 double mutant. This suggests that defects that occur in the meristematic zone later affect the organization of elongating and differentiating cells.

Summary StatementExpression of TAN1 in the root meristematic zone rescues cell file rotation defects in tan1 air9 mutants, suggesting defects that occur in mitosis may influence organization of nondividing cells. Genomic hallmarks and therapeutic implications of cancer cell quiescence Therapy resistance in cancer is often driven by a subpopulation of cells that are temporarily arrested in a non-proliferative, quiescent state, which is difficult to capture and whose mutational drivers remain largely unknown. We developed methodology to uniquely identify this state from transcriptomic signals and characterised its prevalence and genomic constraints in solid primary tumours. We show quiescence preferentially emerges in the context of more stable, less mutated genomes which maintain TP53 integrity and lack the hallmarks of DNA damage repair deficiency, while presenting increased APOBEC mutagenesis. We employ machine learning to uncover novel genomic dependencies of this process, and validate the role of the centrosomal gene CEP89 as a modulator of proliferation/quiescence capacity. Lastly, we demonstrate that quiescence underlies unfavourable responses to various therapies exploiting cell cycle, kinase signalling and epigenetic mechanisms in single cell data, and propose a signature of quiescence-linked therapeutic resistance to further study and clinically track this state.

STATEMENT OF SIGNIFICANCEWe developed a robust transcriptomic signature of cellular quiescence, and employed it to systematically characterise proliferation/quiescence decisions in solid primary cancers and the genomic events influencing them. We propose CEP89 as a novel target whose suppression increases quiescence. Our expression signature of quiescence could be employed to track resistance to multiple anti-cancer compounds in a drug-tolerant persister cell setting. Host Variation in Type I Interferon Signaling Genes (MX1), CCR5{triangleup}32, and MHC Class I Alleles in Treated HIV+ Non-Controllers Predict Viral Reservoir Size Prior genomewide association studies have identified variation in MHC Class I alleles and CCR5{Delta}32 as genetic predictors of viral control, especially in "elite" controllers, individuals who remain virally suppressed in the absence of therapy. We analyzed custom host whole exome sequencing and direct HLA typing from 202 ART-suppressed HIV+ non-controllers in relation to four measures of the peripheral CD4+ T cell reservoir: HIV intact DNA, total (t)DNA, unspliced (us)RNA, and RNA/DNA. Linear mixed models were adjusted for potential covariates including age, sex, nadir CD4+ T cell count, pre-ART HIV RNA, timing of ART initiation from date of detected HIV infection, and duration of ART suppression. Previously reported HLA "protective" B*57:01 (in untreated elite controllers) was significantly associated with lower usRNA (p=4.2x10-4, Benjamini-Hochberg false discovery rate [FDR] q=3.3x10-3). The favorable CCR5{Delta}32 mutation was also associated with lower tDNA and usRNA, primarily in Europeans (p=4.3x10-3 and 8.7x10-3). Several SNPs in interferon stimulated gene, MX1, were associated with HIV total DNA (p=1.3x10-7; q=0.02), and the direction of these associations paralleled predicted eQTL (expression quantitative trait) MX1 gene expression. Among Europeans, gene set enrichment analysis identified genes involved in "negative regulation of host cell viral entry" to be significantly associated with usRNA (q=0.03). These findings suggest a potential role for interferon signaling, MHC class I alleles, and CCR5{Delta}32 in maintaining the reservoir size in HIV non-controllers. These data need to be further validated in functional studies and replication cohorts. Cellular feedback to organotelluranes displays modulation of antioxidant proteins gene expression Organotelluranes RT3 and RT4 are thiol reagents that induce mitochondrial transition pore (MTP) opening in a sensitive and insensitive manner to cyclosporin A. Although RT3 and RT4 promote glutathione depletion, paradoxically, they are also an efficient antioxidant for membrane lipids. These compounds antagonistic effects elicited the challenging question of how the gene expression of antioxidant enzymes would respond to treatment with these compounds. The influence of RT3 and RT4 on antioxidant enzyme expression was investigated in cultured aortic smooth muscle cells (ASMC). RT3 and RT4 promoted disruption of ionic calcium homeostasis, mitochondrial transmembrane potential ({Delta}{Psi}), and cell death in a dose-dependent manner. The cell death mechanisms responded qualitatively to the increase of the organotellurane concentration and changed from apoptosis to necrosis. RT3 and RT4 increased the expression of thioredoxin significantly. RT3 also increased the expression of glutaredoxin and glutathione peroxidase, slightly the catalase expression without significant effects on SOD expression. The results are consistent with GSH and protein thiol depletion and discussed based on the cell toxicity mechanism exhibited by these compounds.



O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/425411v4_ufig1.gif" ALT="Figure 1">
View larger version (73K):
org.highwire.dtl.DTLVardef@52bb82org.highwire.dtl.DTLVardef@19476d9org.highwire.dtl.DTLVardef@6aa99org.highwire.dtl.DTLVardef@1aa605e_HPS_FORMAT_FIGEXP  M_FIG C_FIG Lifespan prolonging mechanisms and insulin upregulation without fat accumulation in long-lived reproductives of a higher termite Kings and queens of eusocial termites can live for decades, while queens sustain a nearly maximal fertility. To investigate the molecular mechanisms underlying their long lifespan, we carried out transcriptomics, lipidomics and metabolomics in Macrotermes natalensis on sterile short-lived workers, long-lived kings and five stages spanning twenty years of adult queen maturation. Reproductives share gene expression differences from workers in agreement with a reduction of several aging-related processes, involving upregulation of DNA damage repair and mitochondrial functions. Anti-oxidant gene expression is downregulated, while peroxidability of membranes in queens decreases. Against expectations, we observed an upregulated gene expression in fat bodies of reproductives of several components of the IIS pathway, including an insulin-like peptide, Ilp9. This pattern does not lead to deleterious fat storage in physogastric queens, while simple sugars dominate in their hemolymph and large amounts of resources are allocated towards oogenesis. Our findings support the notion that all processes causing aging need to be addressed simultaneously in order to prevent it.