Proximity labeling reveals spatial regulation of the anaphase-promoting complex/cyclosome by a microtubule adaptor The anaphase-promoting complex/cyclosome (APC/C) coordinates advancement through mitosis via temporally controlled polyubiquitination of effector proteins. Despite the long-appreciated spatial organization of key events in mitosis mediated largely by cytoskeletal networks, the spatial regulation of APC/C, the major mitotic E3 ligase, is poorly understood. Here, we describe a microtubule-resident protein, PLEKHA5, as an interactor of APC/C and spatial regulator of its activity in mitosis. PLEKHA5 knockdown delayed mitotic progression, causing accumulation of APC/C substrates dependent upon the PLEKHA5-APC/C interaction. To assess the spatial importance of this interaction, we developed microtubule-localized proximity biotinylation tools, which revealed that depletion of PLEKHA5 decreased the extent of APC/C association with the microtubule cytoskeleton. This decreased APC/C microtubule-localization in turn prevented efficient loading of APC/C with its co-activator CDC20, leading to defects in E3 ligase catalytic activity. We propose that PLEKHA5 functions as an adaptor of APC/C that promotes its subcellular localization to microtubules and facilitates its activation by CDC20, thus ensuring the timely turnover of key mitotic APC/C substrates and proper progression through mitosis.
Mechanisms of A-type lamin targeting to nuclear ruptures are disrupted in LMNA- and BANF1-associated progerias Mutations in the genes LMNA and BANF1 can lead to accelerated aging syndromes called progeria. The protein products of these genes, A-type lamins and BAF, respectively, are nuclear envelope (NE) proteins that interact and participate in various cellular processes, including nuclear envelope rupture and repair. BAF localizes to sites of nuclear rupture and recruits NE-repair machinery, including the LEM-domain proteins, ESCRT-III complex, A-type lamins, and membranes. Here, we show that it is a mobile, nucleoplasmic population of A-type lamins that is rapidly recruited to ruptures in a BAF-dependent manner via BAFs association with the Ig-like {beta} fold domain of A-type lamins. These initially mobile lamins become progressively stabilized at the site of rupture. Farnesylated prelamin A and lamin B1 fail to localize to nuclear ruptures, unless that farnesylation is inhibited. Progeria-associated LMNA mutations inhibit the recruitment affected A-type lamin to nuclear ruptures, due to either permanent farnesylation or inhibition of BAF binding. A progeria-associated BAF mutant targets to nuclear ruptures but is unable to recruit A-type lamins. Together, these data reveal the mechanisms that determine how lamins respond to nuclear ruptures and how progeric mutations of LMNA and BANF1 impair recruitment of A-type lamins to nuclear ruptures.
Independent duplications of the Golgi phosphoprotein 3 oncogene in birds Golgi phosphoprotein 3 (GOLPH3) was the first reported oncoprotein of the Golgi apparatus. It was identified as an evolutionarily conserved protein upon its discovery about 20 years ago, but its function remains puzzling in normal and cancer cells. The GOLPH3 gene is part of a group of genes that also includes the GOLPH3L gene. Because cancer has deep roots in multicellular evolution, studying the evolution of the GOLPH3 gene family in non-model species represents an opportunity to identify new model systems that could help better understand the biology behind this group of genes. The main goal of this study is to explore the evolution of the GOLPH3 gene family in birds as a starting point to understand the evolutionary history of this oncoprotein. We identified a repertoire of three GOLPH3 genes in birds. We found duplicated copies of the GOLPH3 gene in all main groups of birds other than paleognaths, and a single copy of the GOLPH3L gene. We suggest there were at least three independent origins for GOLPH3 duplicates. Amino acid divergence estimates show that most of the variation is located in the N-terminal region of the protein. Our transcript abundance estimations show that one paralog is highly and ubiquitously expressed, and the others were variable. Our results are an example of the significance of understanding the evolution of the GOLPH3 gene family, especially for unraveling its structural and functional attributes. Abscisic Acid Drives Stomatal Closure through Increases in Hydrogen Peroxide in Distinct Subcellular Compartments Including Mitochondria Abscisic acid (ABA) closes stomata to minimize transpiration and gas exchange in response to drought. ABA signaling increases accumulation of reactive oxygen species (ROS) in guard cells to drive stomatal closure. We used chemical dyes and genetically-encoded biosensors to identify the subcellular localizations where ABA increases hydrogen peroxide (H2O2) in guard cells. The general redox sensor, dichlorofluorescein (DCF), revealed ABA-driven ROS increases in distinct subcellular locations including chloroplasts, cytosol, and nuclei. ABA also increased fluorescence of both DCF and an H2O2-selective probe, Peroxy Orange 1, in mitochondria, which colocalize with Mitotracker Red. Whole cell and mitochondrial localized ABA-increased ROS signals were lost in an ABA-insensitive quintuple ABA receptor mutant and accentuated in an ABA-hypersensitive mutant. The ABA-induced ROS accumulation in mitochondria was lost in guard cells of mutants with defects in genes encoding H2O2 elevating respiratory burst oxidase homolog (RBOH) enzymes and reduced by treatment with the RBOH inhibitor VAS2870. ABA treatment of guard cells transformed with the genetically-encoded H2O2 reporter targeted to the cytoplasm, roGFP2-Orp1, and the mitochondrial targeted, mt-roGFP2-Orp1, revealed significant ABA-induced H2O2 increases. Furthermore, H2O2 treatment is sufficient to close stomata. These results reveal that ABA elevates H2O2 accumulation in guard cell cytosol and mitochondria to drive stomatal closure.

One sentence summaryGenetically encoded biosensors and chemical probes revealed ABA-dependent increases in hydrogen peroxide, a reactive oxygen species with signaling activity, in guard cell cytoplasm and mitochondria. The Nuclear Lamina binds the EBV genome during latency and regulates viral gene expression The Epstein Barr virus (EBV) infects almost 95% of the population worldwide. While typically asymptomatic, EBV latent infection is associated with several malignancies of epithelial and lymphoid origin in immunocompromised individuals. In latently infected cells, the EBV genome persists as a chromatinized episome that expresses a limited set of viral genes in different patterns, referred to as latency types, which coincide with varying stages of infection and various malignancies. We have previously demonstrated that latency types correlate with differences in the composition and structure of the EBV episome. Several cellular factors, including the nuclear lamina, regulate chromatin composition and architecture. While the interaction of the viral genome with the nuclear lamina has been studied in the context of EBV lytic reactivation, the role of the nuclear lamina in controlling EBV latency has not been investigated. Here, we report that the nuclear lamina is an essential epigenetic regulator of the EBV episome. We observed that in B cells, EBV infection affects the composition of the nuclear lamina by inducing the expression of lamin A/C, but only in EBV+ cells expressing the Type III latency program. Using ChIP-Seq, we determined that lamin B1 and lamin A/C bind the EBV genome, and their binding correlates with deposition of the histone repressive mark H3K9me2. By RNA-Seq, we observed that knock-out of lamin A/C in B cells alters EBV gene expression. Our data indicate that the interaction between lamins and the EBV episome contributes to the epigenetic control of viral gene expression during latency, suggesting a restrictive function of the nuclear lamina as part of the host response against viral DNA entry into the nucleus.

AUTHOR SUMMARYEpstein-Barr virus (EBV) is a common herpesvirus that establishes a lifelong latent infection in a small fraction of B cells of the infected individuals. In most cases, EBV infection is asymptomatic; however, especially in the context of immune suppression, EBV latent infection is associated with several malignancies. In EBV+ cancer cells, latent viral gene expression plays an essential role in sustaining the cancer phenotype. We and others have established that epigenetic modifications of the viral genome are critical to regulating EBV gene expression during latency. Understanding how the EBV genome is epigenetically regulated during latent infection may help identify new specific therapeutic targets for treating EBV+ malignancies. The nuclear lamina is involved in regulating the composition and structure of the cellular chromatin. In the present study, we determined that the nuclear lamina binds the EBV genome during latency, influencing viral gene expression. Depleting one component of the nuclear lamina, lamin A/C, increased the expression of latent EBV genes associated with cellular proliferation, indicating that the binding of the nuclear lamina with the viral genome is essential to control viral gene expression in infected cells. Our data show for the first time that the nuclear lamina may be involved in the cellular response against EBV infection by restricting viral gene expression. Latency structure of BOLD signals within white matter in resting-state fMRI PurposePrevious studies have demonstrated that BOLD signals in gray matter in resting-state functional MRI (RSfMRI) have variable time lags, representing apparent propagations of fMRI BOLD signals in gray matter. We complemented existing findings and explored the corresponding variations of signal latencies in white matter.

MethodsWe used data from the Brain Genomics Superstruct Project, consisting of 1412 subjects (both sexes included) and divided the dataset into ten equal groups to study both the patterns and reproducibility of latency estimates within white matter. We constructed latency matrices by computing cross-correlations between voxel pairs. We also applied a clustering analysis to identify functional networks within white matter, based on which latency analysis was also performed to investigate lead/lag relationship at network level. A dataset consisting of various sensory states (eyes closed, eyes open and eyes open with fixation) was also included to examine the relationship between latency structure and different states.

ResultsProjections of voxel latencies from the latency matrices were highly correlated (average Pearson correlation coefficient = 0.89) across the subgroups, confirming the reproducibility and structure of signal lags in white matter. Analysis of latencies within and between networks revealed a similar pattern of inter- and intra-network communication to that reported for gray matter. Moreover, a dominant direction, from inferior to superior regions, of BOLD signal propagation was revealed by higher resolution clustering. The variations of lag structure within white matter are associated with different sensory states.

ConclusionsThese findings provide additional insight into the character and roles of white matter BOLD signals in brain functions. Loss of SUMO-specific protease 2 causes isolated glucocorticoid deficiency by blocking adrenal cortex zonal transdifferentiation SUMOylation is a dynamic posttranslational modification, that provides fine-tuning of protein function involved in the cellular response to stress, differentiation, and tissue development. In the adrenal cortex, an emblematic endocrine organ that mediates adaptation to physiological demands, the SUMOylation gradient is inversely correlated with the gradient of cellular differentiation raising important questions about its role in functional zonation and the response to stress. Considering that SUMO-specific protease 2 (SENP2), a deSUMOylating enzyme, is upregulated by ACTH/PKA signalling within the zona Fasciculata (zF), we generated mice with adrenal-specific Senp2 loss to address these questions. Disruption of SENP2 activity in steroidogenic cells leads to specific hypoplasia of the zF, a blunted reponses to ACTH and isolated glucocorticoid deficiency. Mechanistically, overSUMOylation resulting from SENP2 loss shifts the balance between ACTH/PKA and WNT/{beta}-catenin signalling leading to repression of PKA activity and ectopic activation of {beta}-catenin. At the cellular level, this blocks transdifferentiation of {beta}-catenin-positive zona Glomerulosa cells into zF cells and sensitises them to premature apoptosis. Our findings indicate that the SUMO pathway is critical for adrenal homeostasis and stress responsiveness. A potent human monoclonal antibody with pan-neutralizing activities directly dislocates S trimer of SARS-CoV-2 through binding both up and down forms of RBD The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic of novel corona virus disease (COVID-19). The neutralizing monoclonal antibodies (mAbs) targeting the receptor binding domain (RBD) of SARS-CoV-2 are among the most promising strategies to prevent and treat COVID-19. However, SARS-CoV-2 variants of concern (VOCs) profoundly reduced the efficacies of most of mAbs and vaccines approved for clinical use. Herein, we demonstrated mAb 35B5 efficiently neutralizes both wild-type (WT) SARS-CoV-2 and VOCs, including B.1.617.2 (delta) variant, in vitro and in vivo. Cryo-electron microscopy (cryo-EM) revealed that 35B5 neutralizes SARS-CoV-2 by targeting a unique epitope that avoids the prevailing mutation sites on RBD identified in circulating VOCs, providing the molecular basis for its pan-neutralizing efficacy. The 35B5-binding epitope could also be exploited for the rational design of a universal SARS-CoV-2 vaccine. Scaling effects of temperature on parasitism from individuals to host-parasite systems Parasitism is expected to change in a warmer future, but whether warming leads to substantial increases in parasitism remains unclear. Understanding how warming effects on parasitism in individual hosts (e.g., parasite load) translate to effects on population-level parasitism (e.g., prevalence, R0) remains a major knowledge gap. We analyzed the temperature dependence of parasitism at both host and population levels in thirteen empirical vector-borne host-parasite systems and found a strong, significant positive correlation between the thermal optima of individual- and population-level parasitism. We also found a significant, positive correlation in eleven environmentally-transmitted parasite systems, though several of these systems exhibited thermal optima >5{o}C apart between individual and population levels. Similarly, parasitism thermal optima were close to host performance thermal optima in vector-borne systems but not in environmentally-transmitted systems. We then adapted and simulated simple models for both transmission modes and found a similar pattern to the empirical systems: thermal optima in vector-borne systems were more strongly correlated across scales compared to environmentally-transmitted systems. Generally, our results suggest that information on the temperature-dependence, and specifically the thermal optimum, at either the individual- or population-level should provide a useful--though not quantitatively exact--baseline for predicting temperature dependence at the other level, especially in vector-borne parasite systems. Environmentally-transmitted parasitism may operate by a different set of rules, in which temperature-dependence is decoupled in some systems, requiring the need for trait-based studies of temperature-dependence at individual and population levels. Betacoronavirus binding dynamics relevant to the functional evolution of the highly transmissible SARS-CoV-2 variant N501Y Comparative functional analysis of the dynamic interactions between various Betacoronavirus mutant strains and broadly utilized target proteins such as ACE2 and CD26, is crucial for a more complete understanding of zoonotic spillovers of viruses that cause diseases such as COVID-19. Here, we employ machine learning to replicated sets of nanosecond scale GPU accelerated molecular dynamics simulations to statistically compare and classify atom motions of these target proteins in both the presence and absence of different endemic and emergent strains of the viral receptor binding domain (RBD) of the S spike glycoprotein. Machine learning was used to identify functional binding dynamics that are evolutionarily conserved from bat CoV-HKU4 to human endemic/emergent strains. Conserved dynamics regions of ACE2 involve both the N-terminal helices, as well as a region of more transient dynamics encompassing K353, Q325 and a novel motif AAQPFLL 386-92 that appears to coordinate their dynamic interactions with the viral RBD at N501. We also demonstrate that the functional evolution of Betacoronavirus zoonotic spillovers involving ACE2 interaction dynamics are likely pre-adapted from two precise and stable binding sites involving the viral bat progenitor strains interaction with CD26 at SAMLI 291-5 and SS 333-334. Our analyses further indicate that the human endemic strains hCoV-HKU1 and hCoV-OC43 have evolved more stable N-terminal helix interactions through enhancement of an interfacing loop region on the viral RBD, whereas the highly transmissible SARS-CoV-2 variants (B.1.1.7, B.1.351 and P.1) have evolved more stable viral binding via more focused interactions between the viral N501 and ACE2 K353 alone. Reference genome-independent taxonomic profiling of microbiomes with mOTUs3 BackgroundTaxonomic profiling is a fundamental task in microbiome research that aims to detect and quantify the relative abundance of microorganisms in biological samples. Available methods using shotgun metagenomic data generally depend on the availability of sequenced and taxonomically annotated reference genomes. However, the majority of microorganisms have not been cultured yet and lack such reference genomes. Thus, a substantial fraction of microbial community members remains unaccounted for during taxonomic profiling of metagenomes, particularly in samples from underexplored environments. To address this issue, we have developed the mOTU profiler, a tool that enables reference genome-independent species-level profiling of metagenomes. As such, it supports the identification and quantification of both "known" and "unknown" species based on a set of select marker genes.

ResultsHere, we present mOTUs3, a command line tool that enables the profiling of metagenomes for >33,000 species-level operational taxonomic units. To achieve this, we leveraged the reconstruction and analysis of >600,000 draft genomes, most of which are metagenome assembled genomes (MAGs), from diverse microbiomes, including soil, freshwater systems, and the gastrointestinal tract of ruminants and other animals, which we found to be greatly underrepresented by reference genomes. Overall, two-thirds of all species-level taxa lacked a reference genome. The cumulative relative abundance of these newly included taxa was low in well-studied microbiomes, such as the human body sites (6-11%). By contrast, they accounted for substantial proportions (ocean, freshwater, soil: 43-63%) or even the vast majority (pig, fish, cattle: 60-80%) of the relative abundance across diverse non-human-associated microbiomes. Using community-developed benchmarks and datasets, we found mOTUs3 to be more accurate than other methods and to be more congruent with 16S rRNA gene-based methods for taxonomic profiling. Furthermore, we demonstrate that mOTUs3 greatly increases the resolution of well-known microbial groups into species-level taxa and helps identify new differentially abundant taxa in comparative metagenomic studies.

ConclusionsWe developed mOTUs3 to enable accurate species-level profiling of metagenomes. Compared to other methods, it provides a more comprehensive view of prokaryotic community diversity, in particular for currently underexplored microbiomes. To facilitate comparative analyses by the research community, it is released with >11,000 precomputed profiles for publicly available metagenomes and is freely available at: https://github.com/motu-tool/mOTUs. Co-option of immune effectors by the hormonal signalling system triggering metamorphosis in Drosophila melanogaster Insect metamorphosis is regulated by the production, secretion and degradation of two peripheral hormones: 20-hydroxyecdysone (ecdysone) and juvenile hormone (JH). In addition to their roles in developmental regulation, increasing evidence suggests that these hormones are involved in innate immunity processes, such as phagocytosis and the induction of antimicrobial peptide (AMP) production. AMP regulation includes systemic responses as well as local responses at surface epithelia that contact with the external environment. At pupariation, Drosophila melanogaster increases dramatically the expression of three AMP genes, drosomycin (drs), drosomycin-like 2 (drsl2) and drosomycin-like 5 (drsl5). We show that the systemic action of drs at pupariation is dependent on ecdysone signalling in the fat body and operates via the ecdysone downstream target, Broad-Z4. In parallel, ecdysone also regulates local responses, specifically through the activation of drsl2 expression in the gut. Finally, we confirm the relevance of this ecdysone dependent AMP expression for the control of bacterial persistence by showing that flies lacking drs expression in the fat body have higher bacterial persistence over metamorphosis. Together, our data emphasize the importance of the association between ecdysone signalling and immunity using in vivo studies and establish a new role for ecdysone at pupariation, which impacts developmental success by regulating the immune system in a stage-dependent manner. We speculate that this co-option of immune effectors by the hormonal system may constitute a pre-emptive mechanism to control bacterial numbers in the pupa at the core of metamorphosis evolution. Visualizing cancer-originated acetate uptake through MCT1 in reactive astrocytes demarcates tumor border and extends survival in glioblastoma patients Glioblastoma multiforme (GBM) is a devastating brain tumor with dismal prognosis of only 15-month survival regardless of surgical resection. Here, we report an advanced neuroimaging technique combining 11C-acetate PET and MRI (AcePET), visualizing the boundary beyond the MRI-defined tumor. Targeted biopsy of the regions with increased 11C-acetate uptake revealed the presence of reactive astrocytes with enhanced acetate-transporter MCT1, along with cancer stem cells. Reactive astrogliosis and MCT1-dependent 11C-acetate-uptake were recapitulated in U87MG-orthotopic models. Mechanistically, glycolytic tumor cells release excessive acetate causing reactive astrogliosis, leading to the release of aberrant astrocytic GABA and H2O2, which further down-regulate the neuronal glucose uptake through GLUT3. Clincally, AcePET-guided surgery allows complete tumor resection of infiltrating cancer stem cells and extends the overall survival of patients by 5.25 months compared to conventional MRI-guided surgery. We established a new concept of the metabolic interactions between GBM cells and neighboring neurons through reactive astrocytes and developed AcePET-guided surgery to fight against GBM. Genome archaeology of two laboratory Salmonella enterica enterica sv Typhimurium The Salmonella research community has used strains and bacteriophages over decades, exchanging useful new isolates among laboratories for study of cell surface antigens, metabolic pathways and restriction-modification studies. Here we present the sequences of two laboratory Salmonella strains (STK005, an isolate of LB5000; and its descendant ER3625). In the ancestry of LB5000, segments of ~15 and ~42 kb were introduced from Salmonella enterica sv Abony 103 into Salmonella enterica sv Typhimurium LT2, forming strain SD14; this strain is thus a hybrid of S. enterica isolates. Strains in the SD14 lineage were used to define flagellar antigens from the 1950s to the 1970s, and to define three restriction-modification systems from the 1960s to the 1980s. LB5000 was also used as host in phage typing systems used by epidemiologists. In the age of cheaper and easier sequencing, this resource will provide access to the sequence that underlies the extensive literature. A Eukaryotic Circuit for Secrete-and-Sense Autonomy Cancers represent complex autonomous robust systems, displaying interconnectivity with feedback control. Autonomous growth is fueled by a cancer cells ability to  secrete-and-sense growth factors: a poorly understood phenomenon. Using an integrated systems and experimental approach, here we dissect the impact of a feedback-coupled GTPase circuit within the secretory pathway that imparts secretion-coupled autonomy. The circuit is assembled when the Ras-superfamily monomeric GTPase Arf1, and the heterotrimeric GTPase Giabg and their corresponding GAPs and GEFs are coupled by GIV/Girdin, a protein that is known to fuel aggressive traits in diverse cancers. One forward and two key negative feedback loops within the circuit create closed-loop control (CLC), allow the two GTPases to coregulate each other, and convert the expected switch-like behavior of Arf1-dependent secretion into an unexpected dose response alignment behavior of sensing and secretion. Such behavior translates into cell survival that is self-sustained by stimuli-proportionate secretion. Findings highlight how enhanced coupling of two biological switches in cancer cells is critical for multiscale feedback control to achieve secretion-coupled autonomy.

GRAPHIC ABSTRACT

O_FIG O_LINKSMALLFIG WIDTH=163 HEIGHT=200 SRC="FIGDIR/small/436048v3_ufig1.gif" ALT="Figure 1">
View larger version (51K):
org.highwire.dtl.DTLVardef@183ff5corg.highwire.dtl.DTLVardef@f21de7org.highwire.dtl.DTLVardef@1893683org.highwire.dtl.DTLVardef@159d966_HPS_FORMAT_FIGEXP  M_FIG C_FIG In BriefThis work delivers an experimentally validated dynamical systems model of the cooperativity between two distinct classes of biological switches in eukaryotic cells and reveals the basis of secretion-coupled cellular autonomy in cancer.

HIGHLIGHTSO_LIModeling and experimental approaches were used to dissect a coupled GTPase circuit
C_LIO_LICoupling enables closed loop feedback and mutual control of GTPases
C_LIO_LICoupling generates dose response alignment behavior of sensing and secretion.
C_LIO_LICoupling is critical for multiscale feedback control to achieve secretion-coupled autonomy.
C_LI 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. RNF213 loss of function reshapes vascular transcriptome and spliceosome leading to disrupted angiogenesis and aggravated vascular inflammatory responses. RationaleMoyamoya disease (MMD) is a rare cerebrovascular occlusive disease that affects Asian population more often. The pathogenesis of MMD is related to mutation in RNF213 gene. However, why, and how RNF213 mutation leads to MMD is still not fully understood.

ObjectiveAnalyze the impact of RNF213 loss of function on vascular cells and the observed changes correlate with MMD.

Methods and resultsRNF213 KD was conducted in HUVEC and vascular smooth muscle cells (vSMCs). First, HUVEC cells showed alteration of angiogenesis, migration under LPS stimulation, Leukocyte endothelial transmigration, and endothelial-to-vSMCs communication.

Transcriptome analysis revealed downregulation of genes regulating cell division and mitosis. Interestingly, Alternative splicing (AS) analysis revealed hundreds of AS events to occur after RNF213 KD in various types of AS. The alternatively spliced genes showed minimal overlap with transcriptome profiling. Pathway analysis revealed many processes and pathways to be regulated by AS events observed. Transcriptome profiling was also performed after LPS treatment and revealed the basis for increased sensitivity to LPS observed in our analysis. AS changes were also observed after LPS treatment in RNF213 KD HUVEC. Different types of AS showed different patterns of convergence or divergence in terms of the regulated pathways after LPS treatment. Finally, transcriptome and AS analysis was performed in vSMCs and showed various processes that impact vSMCs function and phenotype in RNF213 loss of function.

ConclusionOur data provide a wealth of information on RNF213 gene function in vascular tissues and shed important light on different processes that contribute to MMD pathogenesis. Our results signify an immune-driven and hemodynamically linked process of MMD initiation and progression. Frequency modulated timer regulates mammalian hibernation Mammalian hibernators decrease basal metabolism and body temperature (Tb) to minimize energy expenditure in harsh seasons. During hibernation, Tb drops to low temperature (<10 {degrees}C) and remains constant for days, known as deep torpor in small mammalian hibernators. Spontaneous interbout arousals interrupt torpor bouts, when Tb recovers to euthermic state ~37 {degrees}C. Torpor-interbout arousal event repeats during several months of hibernation. However, little is known about mechanisms governing Tb fluctuation across torpor-interbout arousal cycles during hibernation. Recent improvement in data-logging techniques enables us to monitor Tb for more than hundred days with high precision, opening up new avenues for quantitative analysis to address the principle governing Tb fluctuation. Here, we analyzed Tb fluctuation across torpor-interbout arousal cycle of Syrian hamster, which can hibernate in response to chronic cold and short photoperiod under a laboratory condition, using generalized harmonic analysis and discovered a model with frequency modulation quantitatively reproducing Tb fluctuation. This analysis also identified an unexpectedly longer period of 120-430 days as the period that modulates another period of several days, generating Tb fluctuation for Syrian hamster. We propose that concerted action of two endogenous periods governs torpor-interbout arousal cycles during hibernation. Preprints in motion: tracking changes between preprint posting and journal publication during a pandemic Amidst the COVID-19 pandemic, preprints in the biomedical sciences are being posted and accessed at unprecedented rates, drawing widespread attention from the general public, press and policymakers for the first time. This phenomenon has sharpened longstanding questions about the reliability of information shared prior to journal peer review. Does the information shared in preprints typically withstand the scrutiny of peer review, or are conclusions likely to change in the version of record? We assessed preprints from bioRxiv and medRxiv that had been posted and subsequently published in a journal through 30th April 2020, representing the initial phase of the pandemic response. We utilised a combination of automatic and manual annotations to quantify how an article changed between the preprinted and published version. We found that the total number of figure panels and tables changed little between preprint and published articles. Moreover, the conclusions of 7.2% of non-COVID-19-related and 17.2% of COVID-19-related abstracts undergo a discrete change by the time of publication, but the majority of these changes do not qualitatively change the conclusions of the paper. Design, Mutate, Screen: High-throughput creation of genetic clocks with different period-amplitude characteristics Directed evolution has become an invaluable tool in protein engineering and has also greatly influenced the construction of synthetic gene circuits. The ability to generate diversity at precise targets for directed evolution approaches has improved vastly, allowing researchers to create large, specific mutant libraries with relative ease. Screening approaches for large mutant libraries have similarly come a long way, especially when the desired behavior can easily be tested for with static, single time-point assays. For more complex gene circuits with dynamic phenotypes that change over time, directed evolution approaches to controlling and tuning circuit behavior have been hindered by the lack of sufficiently high-throughput screening methods to isolate variants with desired characteristics. Here we utilize directed mutagenesis and multiplexed microfluidics to develop a workflow for creating, screening and tuning dynamic gene circuits that operate at the population level. Specifically, we create a mutant library of an existing oscillator, the synchronized lysis circuit, and tune its dynamics while uncovering principles regarding its behavior. Lastly, we utilize this directed evolution workflow to construct a new synchronized genetic oscillator that exhibits robust dynamics over long time scales. The ribose methylation enzyme FTSJ1 has a conserved role in neuron morphology and learning performance FTSJ1 is a conserved human 2-O-methyltransferase (Nm-MTase) that modifies several transfer RNAs (tRNAs) at position 32 and the wobble position 34 in the AntiCodon Loop (ACL). Its loss of function has been linked to Non-Syndromic X-Linked Intellectual Disability (NSXLID), and more recently to cancers. However, the molecular mechanisms underlying these pathologies are currently unclear. Here we report a novel FTSJ1 pathogenic variant from a NSXLID patient. Using blood cells derived from this patient and other affected individuals carrying FTSJ1 mutations, we performed an unbiased and comprehensive RiboMethSeq analysis to map the ribose methylation (Nm) on all tRNAs and identify novel targets. In addition, we performed a transcriptome analysis in these cells and found that several genes previously associated with intellectual disability and cancers were deregulated. We also found changes in the miRNA population that suggest potential cross-regulation of some miRNAs with these key mRNA targets. Finally, we show that differentiation of FTSJ1-depleted human neuronal progenitor cells (NPC) into neurons displays long and thin spine neurites compared to control cells. These defects are also observed in Drosophila and are associated with long term memory deficit in this organism. Altogether, our study adds insight into FTSJ1 pathologies in human by the identification of novel FTSJ1 targets and the defect in neuron morphology. IRGM/Irgm1 Aggravates Progression of Atherosclerosis by Inducing Macrophage Apoptosis through the MAPK Signaling Pathway AimsAtherosclerosis underlies most cardiovascular diseases, among which acute coronary syndrome (ACS) caused by plaque rupture (PR) often leads to death. Immune-related GTPases (IRGM/Irgm1) have been extensively studied in inflammatory diseases, but their role in atherosclerosis is unclear. Determining how IRGM/Irgm1 promotes atherosclerotic plaque vulnerability will provide information for new biomarkers and/or therapeutic targets.

Methods and resultsWe identified ruptured and unruptured plaques by optical coherence tomography, and found that serum IRGM was highly expressed in patients with ST-segment elevation myocardial infarction in PR. We used ApoE-/-Irgm1+/+, ApoE-/-Irgm1+/- mice and chimeric mice to establish a model of advanced atherosclerosis. The results of pathological experiments showed that Irgm1 caused plaque necrosis. The ratio of neutral lipids and cholesterol crystals increases, while the content of collagen fibers decreases, aggravating the destabilization of atherosclerotic plaques. In vitro, we used multiple approaches to confirm that Irgm1 promotes macrophage apoptosis by promoting the production of reactive oxygen species and activating the MAPK signaling pathway.

ConclusionsIRGM may be a potential risk factor for PR. Mechanistic studies have shown that IRGM/Irgm1 contributes to the formation and rupture of fragile plaques. This is partly mediated by the induction of macrophage apoptosis via the MAPK signaling pathway. IRGM may offer new strategies for early treatment of ACS.

Translation viewOur findings indicate that IRGM/Irgm1 contributes to formation and rupture of vulnerable plaques. It suggests that IRGM may provide a new target for the early treatment of ACS. Differences in gait stability and acceleration characteristics between healthy young and older females Our aim was to evaluate differences in gait acceleration intensity, variability and stability of feet and trunk between older females and young females using inertial sensors. Twenty older females (OF; mean age 68.4, SD 4.1 years) and eighteen young females (YF; mean age 22.3, SD 1.7 years) were asked to walk straight for 100 meters at their preferred speed, while wearing inertial sensors on heels and lower back. We calculated spatiotemporal measures, foot and trunk acceleration characteristics and their variability, as well as trunk stability using the local divergence exponent (LDE). Two-way analysis of variance (including the factors foot and age), Students t-test, and Mann-Whitney U test were used to compare statistical differences of measures between groups. Cohens d effects were calculated for each variable. Foot maximum vertical acceleration and amplitude, trunk-foot vertical acceleration attenuation, as well as their variability were significantly smaller in OF than in YF. In contrast, trunk mediolateral acceleration amplitude, maximum vertical acceleration, and amplitude, as well as their variability were significantly larger in OF than in YF. Moreover, OF showed lower stability (i.e., higher LDE values) in mediolateral acceleration, mediolateral and vertical angular velocity of the trunk. Even though we measured healthy older females, these participants showed lower vertical foot accelerations with higher vertical trunk acceleration, lower trunk-foot vertical acceleration attenuation, less gait stability, and more variability of the trunk, and hence, were more likely to fall. These findings suggest that instrumented gait measurements may help for early detection of changes or impairments in gait performance, even before this can be observed by clinical eye or gait speed. An Extended Motif in the SARS-CoV-2 Spike Modulates Binding and Release of Host Coatomer in Retrograde Trafficking {beta}-Coronaviruses such as SARS-CoV-2 hijack coatomer protein-I (COPI) for spike protein retrograde trafficking to the progeny assembly site in endoplasmic reticulum-Golgi intermediate compartment (ERGIC). However, limited residue-level details are available into how the spike interacts with COPI. Here we identify an extended COPI binding motif in the spike that encompasses the canonical K-x-H dibasic sequence. This motif demonstrates selectivity for COPI subunit. Guided by an in silico analysis of dibasic motifs in the human proteome, we employ mutagenesis and binding assays to show that the spike motif terminal residues are critical modulators of complex dissociation, which is essential for spike release in ERGIC. COPI residues critical for spike motif binding are elucidated by mutagenesis and crystallography and found to be conserved in the zoonotic reservoirs, bats, pangolins, camels, and in humans. Collectively, our investigation on the spike motif identifies key COPI binding determinants with implications for retrograde trafficking. Neurological mechanism of sensory deficits after developmental exposure to non-dioxin-like polychlorinated biphenyls (PCBs) in zebrafish larvae The most abundant polychlorinated biphenyl (PCB) congeners found in the environment and in humans are neurotoxic. This is of particular concern for early life stages because the exposure of the more vulnerable developing nervous system to chemicals can result in neurobehavioral disorders. To uncover currently unknown links between PCB target mechanisms and neurobehavioral deficits, we investigated the effects of the non-dioxin-like (NDL) congener PCB153 on neuronal morphology and synaptic transmission linked to the proper execution of a sensorimotor response using zebrafish as a vertebrate model. Zebrafish that were exposed during development to concentrations similar to those found in human cord blood and PCB contaminated sites showed a delay in startle response. A similar delay was observed for other NDL congeners but not for the potent dioxin-like congener PCB126. Morphological and biochemical data demonstrate that while exposure to PCB153 induced swelling of afferent sensory neurons, the disruption of dopaminergic and GABAergic signaling is the dominant mechanism associated with motor movement. The effects on important and broadly conserved signaling mechanisms in vertebrates suggest that NDL PCBs may contribute to neurodevelopmental abnormalities in humans and, with the startle response being critical for the survival of fish, to evolutionary adaptation in wildlife. The initial response of females towards congeneric males matches the propensity to hybridise in Ophthalmotilapia. AO_SCPLOWBSTRACTC_SCPLOWCichlid radiations often harbour closely related species with overlapping niches and distribution ranges. Such species sometimes hybridise in nature, which raises the question how can they coexist. This also holds for the Tanganyika mouthbrooders Ophthalmotilapia ventralis and O. nasuta. Earlier studies found indications of asymmetrical hybridisation with females of O. ventralis accepting males of O. nasuta, but not the other way around. We hypothesised that this was due to differences in the capacity for species recognition. Given the higher propensity of O. ventralis females towards hybridisation, we expect a reduced ability for species recognition in O. ventralis females, compared to O. nasuta females. We staged two experiments, one focusing on 22 female O. nasuta and one on 21 female O. ventralis. These fish were placed in one half of a tank and briefly exposed to a conspecific or a heterospecific male, a conspecific female, or nothing (control). Female response was evaluated by scoring six tracking parameters and by noting the occurrence of ten discrete behaviours before and during the encounter. Females always responded to the presence of another fish by approaching it. Remarkably, for both O. nasuta and O. ventralis, we did not find a different response between encounters with conspecific males and females. However, in agreement with our hypothesis, O. nasuta females behaved differently towards conspecific or heterospecific males, whereas O. ventralis females did not. When presented with a heterospecific male, O. nasuta females performed a lower number of  ram behaviours. Additionally, they never displayed the  flee behaviour, a component of the species mating repertoire that was seen in all but one of the presentations with a conspecific male. Our findings show that differences in species recognition at first encounter predict to a large degree the outcome of the mating process, even in the absence of mating behaviour.