CRCM5484: A BET- BDII Selective Compound With Differential Anti-Leukemic Drug Modulation Differentially screening the Fr-PPIChem chemical library on the BET BRD4-BDII versus -BDI bromodomains led to the discovery of a BDII selective tetrahydropyridothienopyrimidinone (THPTP)-based compound. Structure-activity relationship (SAR) and hit-to-lead approaches allowed us to develop CRCM5484, a potent inhibitor of BET proteins with a preferential and 475-fold selectivity for the second bromodomain of the BRD3 protein (BRD3-BDII) over its first bromodomain (BRD3-BDI). Its very low activity was demonstrated in various cell-based assays, corresponding with recent data describing other selective BDII compounds. However, screening on a drug sensitivity and resistance-profiling platform revealed its ability to modulate the anti-leukemic activity in combination with various FDA-approved and/or in-development drugs in a cell- and context-dependent differential manner. Altogether, the results confirm the originality of the THPTP molecular mode of action in the BD cavity and its potential as starting scaffold for the development of potent and selective bromodomain inhibitors.
NPEPPS regulates intracellular import and sensitivity to cisplatin by interaction with volume regulated anion channels Despite routine use of platinum-based chemotherapeutics in cancer treatment, there remains a need to improve efficacy and patient selection. Multi-omic assessment of human bladder cancer cell lines and their cisplatin resistant derivatives and whole-genome CRISPR screens identified NPEPPS, the puromycin-sensitive aminopeptidase as a novel driver of cisplatin resistance. NPEPPS depletion increased cisplatin import and sensitization of resistant cells in vitro and in vivo. Pharmacologic inhibition of NPEPPS in cells and chemoresistant, patient-derived tumor organoids improved response to cisplatin. NPEPPS was found in complex with volume regulated anion channel (VRAC) subunits LRRC8A and LRRC8D, whose loss is known to enhance resistance to cisplatin. Depletion of LRRC8A the only obligate subunit for normal VRAC function abrogated the effect of NPEPPS-mediated cisplatin import. Our findings describe the first mechanism by which VRACs can be targeted for therapeutic benefit.

HIGHLIGHTSO_LIMulti-omic screening found NPEPPS is a driver of cisplatin resistance in cancer
C_LIO_LINPEPPS is in protein complex with LRRC8A and D, volume regulated anion channel (VRAC) subunits
C_LIO_LILRRC8A and D loss increases cisplatin resistance and reduce its intracellular levels
C_LIO_LINPEPPS regulates cisplatin import and sensitivity through VRACs
C_LIO_LINPEPPS function is inhibited by small molecules offering potential for clinical translation
C_LI
Comparison of hybrid learning and remote education in the implementation of the 'Adopt a Microorganism' methodology The Internet has changed the way teachers and students access information and build knowledge. The recent COVID-19 pandemic has created challenges for both teachers and students and a demand for new methodologies of remote learning. In the life sciences, mixing online content with practical activities represents an even greater challenge. In microbiology, the implementation of an active teaching methodology, the #Adopt project, based on the social network Facebook(R), represents an excellent option for connecting remote education with classroom activities. In 2020, the version applied in high school, "Adopt a Microorganism", was adapted to meet the demands of emergency remote education owing to the suppression of face-to-face activities caused by the pandemic. In the present study, we assessed how the change in methodology impacted the discourse richness of students from high school integrated with technical education in the Business Administration program of the Federal Institute of Sao Paulo, Sorocaba Campus. Three questionnaires related to the groups of microorganisms (Archaea, Bacteria, Virus, Fungi, and Protozoan) were applied. The students responses in the 2019 and 2020 classes were compared concerning content richness and multiplicity of concepts through the application of the Shannon diversity index, an approach that is generally used to assess biodiversity in different environments. The observed results suggest that remote learning provided students with a conceptual basis and richness of content equivalent to that achieved by students subjected to the hybrid teaching model. In conclusion, this study suggests that the #Adopt project methodology increases students discourse richness in microbiology even without face-to-face traditional classes. 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. High-quality Arabidopsis thaliana Genome Assembly with Nanopore and HiFi Long Reads Arabidopsis thaliana is an important and long-established model species for plant molecular biology, genetics, epigenetics, and genomics. However, the latest version of reference genome still contains significant number of missing segments. Here, we report a high-quality and almost complete Col-0 genome assembly with two gaps (Col-XJTU) using combination of Oxford Nanopore Technology ultra-long reads, PacBio high-fidelity long reads, and Hi-C data. The total genome assembly size is 133,725,193 bp, introducing 14.6 Mb of novel sequences compared to the TAIR10.1 reference genome. All five chromosomes of Col-XJTU assembly are highly accurate with consensus quality (QV) scores > 60 (ranging from 62 to 68), which are higher than those of TAIR10.1 reference (QV scores ranging from 45 to 52). We have completely resolved chromosome (Chr) 3 and Chr5 in a telomere-to-telomere manner. Chr4 has been completely resolved except the nucleolar organizing regions, which comprise long repetitive DNA fragments. The Chr1 centromere (CEN1), reportedly around 9 Mb in length, is particularly challenging to assemble due to the presence of tens of thousands of CEN180 satellite repeats. Using the cutting-edge sequencing data and novel computational approaches, we assembled about 4 Mb of sequence for CEN1 and a 3.5-Mb-long CEN2. We investigated the structure and epigenetics of centromeres. We detected four clusters of CEN180 monomers, and found that the centromere-specific histone H3-like protein (CENH3) exhibits a strong preference for CEN180 cluster 3. Moreover, we observed hypomethylation patterns in CENH3-enriched regions. We believe that this high-quality genome assembly, Col-XJTU, would serve as a valuable reference to better understand the global pattern of centromeric polymorphisms, as well as genetic and epigenetic features in plants. A novel Boolean network inference strategy to model early hematopoiesis aging Hematopoietic stem cell (HSC) aging is a multifactorial event that leads to changes in HSC properties and function. These changes are intrinsically coordinated and affect the early hematopoiesis, involving hematopoietic stem and progenitor cells (HSPCs). The objective of this work is to better understand the mechanisms and factors controlling these changes. We have therefore developed an original strategy to construct a Boolean network of genes explaining the priming and homeostasis of HSCs (graphical abstract). Based on our previous scRNA-seq data, we performed an exhaustive analysis of the transcriptional network and identified active transcription modules or regulons along the differentiation trajectory of selected HSPC states. This global view of transcriptional regulation led us to focus on 15 components, 13 selected TFs (Tal1, Fli1, Gata2, Gata1, Zfpm1, Egr1, Junb, Ikzf1, Myc, Cebpa, Bclaf1, Klf1, Spi1) and 2 complexes regulating the ability of HSC to cycle (CDK4/6 - Cyclin D and CIP/KIP). We then defined the connections controlling the differentiation dynamics of HSC states and constructed an influence graph between the TFs involved in the dynamics by mixing observations from our scRNA-seq data and knowledge from the literature. Then, using answer set programming (ASP) and in silico perturbation analysis, we obtained a Boolean model which is the solution of a Boolean satisfiability problem. Finally, perturbation of the model based on age-related changes revealed important regulations, such as the overactivation of Egr1 and Junb or the loss of Cebpa activation by Gata2, which were found to be relevant for the myeloid bias of aged HSC. Our work shows the efficiency of the combination of manual and systematic methods to elaborate a Boolean model. The developed strategy led to the proposal of new regulatory mechanisms underlying the differentiation bias of aged HSCs, explaining the decreased transcriptional priming of HSCs to all mature cell types except megakaryocytes. Ultra-deep through-skull mouse brain imaging via the combination of skull optical clearing and three-photon microscopy Optical microscopy has enabled in vivo monitoring of brain structures and functions with high spatial resolution. However, the strong optical scattering in turbid brain tissue and skull impedes the observation of microvasculature and neuronal structures at a large depth. Herein, we proposed a strategy to overcome the influence induced by the high scattering effect of both skull and brain tissue via the combination of skull optical clearing (SOC) technique and thee-photon fluorescence microscopy (3PM). The Visible-NIR-II compatible Skull Optical Clearing Agents (VNSOCA) we applied reduced the skull scattering and water absorption in long wavelength by refractive index matching and H2O replacement to D2O respectively. 3PM with the excitation in the 1300-nm window reached 1.5 mm cerebrovascular imaging depth in cranial window. Combining the two advanced technologies together, we achieved so far the largest cerebrovascular imaging depth of 1.0 mm and neuronal imaging depth of >700 m through intact mouse skull. Dual-channel through-skull imaging of both brain vessels and neurons was also successfully realized, giving an opportunity of non-invasively monitoring the deep brain structures and functions at single-cell level simultaneously. Investigation of ongoing recombination through genealogical reconstruction for SARS-CoV-2 AO_SCPLOWBSTRACTC_SCPLOWThe evolutionary process of genetic recombination has the potential to rapidly change the properties of a viral pathogen, and its presence is a crucial factor to consider in the development of treatments and vaccines. It can also significantly affect the results of phylogenetic analyses and the inference of evolutionary rates. The detection of recombination from samples of sequencing data is a very challenging problem, and is further complicated for SARS-CoV-2 by its relatively slow accumulation of genetic diversity. The extent to which recombination is ongoing for SARS-CoV-2 is not yet resolved. To address this, we use a parsimony-based method to reconstruct possible genealogical histories for samples of SARS-CoV-2 sequences, which enables us to pinpoint specific recombination events that could have generated the data. We propose a statistical framework for disentangling the effects of recurrent mutation from recombination in the history of a sample, and hence provide a way of estimating the probability that ongoing recombination is present. We apply this to samples of sequencing data collected in England and South Africa, and find evidence of ongoing recombination. A novel two-generation approach to the population dynamics of gregarious parasitoids Parasitoids, as important natural enemies, help maintain balance in natural ecosystems. Their population dynamics is generally predicted from the number of individuals. Here, using gregarious parasitoids as models, we show that this traditional approach omits one important parameter: the mothers manipulation of offspring fertility due to the clutch size-body size- fertility correlation. As a result of this correlation, when females deliberately adjust the number of offspring laid in a host, they determine not only the number but also the body sizes and reproductive potentials of those offspring. For the first time in the model species Anaphes flavipes, we determined the parasitoids offspring fertility from clutch size. Using this, we experimentally clarified the advantage of specific clutch size combinations and we show that identical fertility in the F1 generation can lead to distinctly different fertility values in the F2 generation. Even with the same number of hosts, lower fertility in the F1 generation can cause higher fertility in the F2 generation. Based on these results, we propose a novel two-generation approach which includes the clutch size-body size-fertility correlation. Our novel approach provides a new perspective for determining the individual fitness levels of gregarious parasitoids with new options for the modelling of parasitoid population dynamics. An integrative approach leads to the discovery of a novel anti-leukemic peptide from human milk Chemotherapy in childhood leukemia is associated with late morbidity in leukemic survivors, while certain patient subsets are relatively resistant to standard chemotherapy. It is therefore important to identify new agents with sensitivity and selectivity towards leukemic cells, while having less systemic toxicity. Peptide-based therapeutics has gained much attention during the last few years. Here, we used an integrative workflow combining mass spectrometric peptide library construction, in silico anticancer peptide screening, and in vitro leukemic cell studies to discover a novel anti-leukemic peptide having 3+charges and alpha-helical structure, namely HMP-S7, from human breast milk. HMP-S7 showed cytotoxic activity against four distinct leukemic cell lines in a dose-dependent manner but had no effect on solid malignancies or representative normal cells. HMP-S7 induced leukemic cell death by penetrating the plasma membrane to enter the cytoplasm and cause leakage of lactate dehydrogenase, thus acting in a membranolytic manner. Importantly, HMP-S7 exhibited anti-leukemic effect against patient-derived leukemic cells ex vivo. In conclusion, HMP-S7 is a selective anti-leukemic peptide with promise which requires further validation in preclinical and clinical studies.

TeaserIn silico screening of naturally occurring human milk peptides discovers a new anticancer peptide that kills leukemic cells in vitro and ex vivo. Between two walls: Modeling the adsorption behavior of Beta-glucosidase A on bare and SAM-functionalised gold surfaces The efficient immobilization of enzymes on surfaces remains a complex but central issue in the biomaterials field, which requires us to understand this process at the atomic level. Using a multi-scale approach combining all-atom molecular dynamics and coarse-grain Brownian dynamics simulations, we investigated the adsorption behavior of {beta}-glucosidase A ({beta}GA) on bare and SAM-functionalized gold surfaces. We monitored the enzyme position and orientation during the MD trajectories, and measured the contacts it forms with both surfaces. While the adsorption process has little impact on the protein conformation, it can nonetheless perturb its mechanical properties and catalytic activity. Our results show that compared to the SAM-functionalized surface, the adsorption of {beta}GA on bare gold is more stable, but also less specific, and more likely to disrupt the enzymes function. This observation emphasizes the fact that the structural organization of proteins at the solid interface is a keypoint when designing devices based on enzyme immobilization, as one must find an acceptable stability-activity trade-off.

TOC image

O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=94 SRC="FIGDIR/small/450859v2_ufig1.gif" ALT="Figure 1">
View larger version (26K):
org.highwire.dtl.DTLVardef@d3d0d7org.highwire.dtl.DTLVardef@3403e4org.highwire.dtl.DTLVardef@eb804dorg.highwire.dtl.DTLVardef@13469c2_HPS_FORMAT_FIGEXP  M_FIG C_FIG Glucose inhibits haemostasis and accelerates diet-induced hyperlipidaemia in zebrafish larvae Hyperglycaemia damages the microvasculature in part through the reduced recruitment of immune cells and interference with platelet signalling, leading to poor wound healing and accelerated lipid deposition in mammals. We investigated the utility of zebrafish larvae to model the effect of glucose on neutrophil and macrophage recruitment to a tail wound, wound-induced haemostasis, and chicken egg yolk feed challenge-induced hyperlipidaemia by supplementing larvae with exogenous glucose by immersion or injection. Neither method of glucose supplementation affected the recruitment of neutrophils and macrophages following tail transection. Glucose injection reduced thrombocyte retention and fibrin plug formation while only thrombocyte retention was reduced by glucose immersion following tail transection. We observed accelerated lipid accumulation in glucose-injected larvae challenged with high fat chicken egg yolk feeding. Our study identifies conserved and divergent effects of high glucose on inflammation, haemostasis, and hyperlipidaemia in zebrafish larvae compared to mammals. Loss of histone methyltransferase SETD1B in oogenesis results in the redistribution of genomic histone 3 lysine 4 trimethylation Histone 3 lysine 4 trimethylation (H3K4me3) is an epigenetic mark found at gene promoters and CpG islands. H3K4me3 is essential for mammalian development, yet mechanisms underlying its genomic targeting are poorly understood. H3K4me3 methyltransferases SETD1B and MLL2 are essential for oogenesis. We investigated changes in H3K4me3 in Setd1b conditional knockout (cKO) oocytes using ultra-low input ChIP-seq, with comparisons to DNA methylation and gene expression analyses. H3K4me3 was redistributed in Setd1b cKO oocytes showing losses at active gene promoters associated with downregulated gene expression. Remarkably, many regions also gained H3K4me3, in particular those that were DNA hypomethylated, transcriptionally inactive and CpG-rich, which are hallmarks of MLL2 targets. Consequently, loss of SETD1B disrupts the balance between MLL2 and de novo DNA methyltransferases in determining the epigenetic landscape during oogenesis. Our work reveals two distinct, complementary mechanisms of genomic targeting of H3K4me3 in oogenesis, with SETD1B linked to gene expression and MLL2 to CpG content.

Graphical AbstractIn oogenesis, SETD1B and CXXC1 target H3K4me3 to actively transcribed gene promoters, while MLL2 targets transcriptionally inactive regions based on underlying CpG composition (upper panel). When SETD1B is ablated, H3K4me3 is lost at a subset of active promoters, resulting in downregulation of transcription (lower panel). Loss of SETD1B alters the activity of MLL2, permitting MLL2 to deposit H3K4me3 at CpG-rich regions, many of which should otherwise be DNA methylated. Thus, it is evident that MLL2 and de novo DNMTs compete for genomic occupancy late in oogenesis, and loss of SETD1B disrupts the balance of these mechanisms.

O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY Design, immunogenicity and efficacy of a Pan-SARS-CoV-2 synthetic DNA vaccine Here we have employed SynCon(R) design technology to construct a DNA vaccine expressing a pan-Spike immunogen (INO-4802) to induce broad immunity across SARS-CoV-2 variants of concern (VOC). Compared to WT and VOC-matched vaccines which showed reduced cross-neutralizing activity, INO-4802 induced potent neutralizing antibodies and T cell responses against WT as well as B.1.1.7, P.1, and B.1.351 VOCs in a murine model. In addition, a hamster challenge model demonstrated that INO-4802 conferred superior protection following intranasal B.1.351 challenge. Protection against weight loss associated with WT, B.1.1.7, P.1 and B.1.617.2 challenge was also demonstrated. Vaccinated hamsters showed enhanced humoral responses against VOC in a heterologous WT vaccine prime and INO-4802 boost setting. These results demonstrate the potential of the pan-SARS-CoV-2 vaccine, INO-4802 to induce cross-reactive immune responses against emerging VOC as either a standalone vaccine, or as a potential boost for individuals previously immunized with WT-matched vaccines. Muscle activation strategies of the vastus lateralis according to sex AimDespite men exhibiting greater muscle strength and fatigibility than women, it remains unclear if there are sex-based differences in muscle recruitment strategies e.g. motor unit (MU) recruitment and modulation of firing rate (FR) at normalised forces and during progressive increases in force.

MethodsTwenty-nine healthy male and thirty-one healthy female participants (18-35 years) were studied. Intramuscular electromyography was used to record individual motor unit potentials (MUPs) and near fibre MUPs from the vastus lateralis (VL) during 10% and 25% maximum isometric voluntary contractions (MVC), and spike-triggered averaging was used to obtain motor unit number estimates (MUNE) of the VL. Multilevel mixed-effects linear regression models were used to investigate the effects of sex at each contraction level.

ResultsMen exhibited greater muscle strength (p<0.001) and size (p<0.001) than women, with no difference in force steadiness at 10% or 25% MVC. Women had smaller MUs and higher FR at 10% MVC (both p<0.02), similar to that at 25% MVC in MU size (p=0.062) and FR (p=0.031). However, both sexes showed similar increases in MU size and FR when moving from low-to mid-level contractions. There were no sex differences in any near fibre MUP parameters or in MUNE.

ConclusionIn the vastus lateralis, women produce muscle force via different neuromuscular recruitment strategies to men which is characterised by smaller MUs discharging at higher rates. However, similar strategies are employed to increase force production from low to moderate contractions. These findings of similar proportional increases between sexes support the use of mixed sex cohorts in studies of this nature.

Key pointsO_LIIncreases in muscle force production are mediated by motor unit (MU) recruitment, and MU firing rate (FR).
C_LIO_LIWomen are underrepresented in studies of human neuromuscular research and markedly differ to men in a number of aspects of neuromuscular function, yet little is known of the recruitment strategies of each.
C_LIO_LIHere we demonstrate men and women have similar vastus lateralis MU number estimates, yet women recruit smaller MUs with higher FR than men at normalised contraction levels. However, increases in force are achieved via similar trajectories of MU recruitment and MU FR in men and women.
C_LIO_LIAlthough men and women exhibit divergent neuromuscular recruitment strategies to achieve normalised forces, increases in force are achived similarly and support the inclusion of mixed sex cohorts in studies of this nature.
C_LI A general framework to learn tertiary structure for protein sequence characterization During the past five years, deep-learning algorithms have enabled ground-breaking progress towards the prediction of tertiary structure from a protein sequence. Very recently, we developed SAdLSA, a new computational algorithm for protein sequence comparison via deep-learning of protein structural alignments. SAdLSA shows significant improvement over established sequence alignment methods. In this contribution, we show that SAdLSA provides a general machine-learning framework for structurally characterizing protein sequences. By aligning a protein sequence against itself, SAdLSA generates a fold distogram for the input sequence, including challenging cases whose structural folds were not present in the training set. About 70% of the predicted distograms are statistically significant. Although at present the accuracy of the intra-sequence distogram predicted by SAdLSA self-alignment is not as good as deep-learning algorithms specifically trained for distogram prediction, it is remarkable that the prediction of single protein structures is encoded by an algorithm that learns ensembles of pairwise structural comparisons, without being explicitly trained to recognize individual structural folds. As such, SAdLSA can not only predict protein folds for individual sequences, but also detects subtle, yet significant, structural relationships between multiple protein sequences using the same deep-learning neural network. The former reduces to a special case in this general framework for protein sequence annotation. A nuclear import pathway exploited by pathogenic noncoding RNAs The prevailing view regarding intracellular RNA trafficking in eukaryotic cells describes that RNAs transcribed in the nucleus either stay in the nucleus or cross the nuclear envelope entering the cytoplasm for function. Interestingly, emerging evidence illustrates numerous functional RNAs trafficking in the reverse direction from the cytoplasm to the nucleus. However, the mechanism underlying the RNA nuclear import has not been well elucidated. Viroids are single-stranded circular noncoding RNAs that infect plants. Using nuclear-replicating viroids as a model, we showed that cellular Importin alpha-4 is likely involved in viroid RNA nuclear import, empirically supporting the involvement of Importin-based cellular pathway in RNA nuclear import. We also confirmed the involvement of a cellular protein (Virp1) that binds both Importin alpha-4 and viroids. Furthermore, a conserved C-loop in nuclear-replicating viroids is critical for Virp1 binding. Disrupting C-loop impairs Virp1 binding, viroid nuclear accumulation and infectivity. Further, C-loop exists in a subviral satellite noncoding RNA that relies on Virp1 for nuclear import. These results have significant implications for understanding the infection process of subviral agents. In addition, our data outline a cellular pathway responsible for the nuclear import of RNAs and uncover a 3-dimensional RNA motif-based regulation over RNA nuclear import. A unified view of low complexity regions (LCRs) across species Low-complexity regions (LCRs) in proteins are important for higher-order assemblies of organisms, yet we lack a unified view of their sequences, features, relationships, and functions. Here, we use dotplots and dimensionality reduction to systematically define LCR type/copy relationships and create a map of LCR sequence space capable of integrating LCR features and functions. By defining LCR relationships across the proteome, we provide insight into how LCR type and copy number contribute to higher-order assemblies, such as the importance of K-rich LCR copy number for assembly of the nucleolar protein RPA43 in vivo and in vitro. With LCR maps, we reveal the underlying structure of LCR sequence space, and relate differential occupancy in this space to the conservation and emergence of higher-order assemblies, including the metazoan extracellular matrix and plant cell wall. Together, LCR relationships and maps uncovered the distribution and prevalence of E-rich LCRs in the nucleolus, and revealed previously undescribed regions of LCR sequence space with properties of higher order assemblies, including a teleost-specific T/H-rich sequence space. Thus, this unified view of LCRs enables discovery of how LCRs encode higher-order assemblies of organisms. Identifying and correcting for misspecifications in GWAS summary statistics and polygenic scores Publicly available genome-wide association studies (GWAS) summary statistics exhibit uneven quality, which can impact the validity of follow-up analyses. First, we present an overview of possible misspecifications that come with GWAS summary statistics. Then, in both simulations and real data analyses, we show that additional information such as imputation INFO scores, allele frequencies, and per-variant sample sizes in GWAS summary statistics can be used to detect possible issues and correct for misspecifications in the GWAS summary statistics. One important motivation for us is to improve the predictive performance of polygenic scores built from these summary statistics. Unfortunately, due to the lack of reporting standards for GWAS summary statistics, this additional information is not systematically reported. We also show that using well-matched LD references can improve model fit and translate into more accurate prediction. Finally, we discuss how to make polygenic score methods such as lassosum and LDpred2 more robust to these misspecifications to improve their predictive power. 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. Recovery of high-quality assembled genomes via metagenome binning guided with single-cell amplified genomes BackgroundObtaining high-quality (HQ) reference genomes from microbial communities is crucial for understanding the phylogeny and function of uncultured microbes in complex microbial ecosystems. Despite the improved bioinformatic approaches to generate curated metagenome-assembled genomes (MAGs), existing metagenomic binners often fail to obtain reliable MAGs, and thus, they are nowhere comparable to genomes sequenced from isolates in terms of strain level resolution. Here, we present a single-cell genome-guided metagenome binning (MetaSAG) to reconstruct the strain-resolved genomes from microbial communities at once.

ResultsMetaSAG employs single-cell amplified genomes (SAGs) generated with microfluidic technology as binning guides to recover improved draft genomes with the metagenomic data. To assess the performance of reconstructing genomes from various microbial communities, we compared MetaSAG with four conventional metagenomic binners using a cell mock community, human gut microbiota, and skin microbiota samples. MetaSAG showed precise contig binning and higher recovery rates (>97%) of rRNA and plasmids compared to conventional binners in genome reconstruction from the cell mock community. In human microbiota samples, MetaSAG recovered the largest number of genomes with a total of 103 gut microbial genomes (21 HQ and 65 showed >90% completeness) and 45 skin microbial genomes (10 HQ and 40 showed >90% completeness), respectively. Conventional binners recovered one Staphylococcus hominis genome, whereas MetaSAG recovered two S. hominis genomes from the identical skin microbiota sample. Single-cell sequencing indicated that these S. hominis genomes clearly derived from two distinct strains harboring specifically different plasmids. We found that all conventional S. hominis MAGs had substantial lack or excess of the genome sequences and contamination of other Staphylococcus bacteria (S. epidermidis).

ConclusionsMetaSAG enabled us to obtain the strain-resolved genomes in the mock community and human microbiota samples by assigning metagenomic sequences correctly and covering both highly conserved genes such as rRNA genes and unique extrachromosomal elements, including plasmids. MetaSAG will provide HQ genomes that are difficult to obtain with metagenomic analyses alone and will facilitate the understanding of microbial ecosystems by elucidating detailed metabolic pathways and horizontal gene transfer networks through microbial genomes. MetaSAG is available at https://github.com/kojiari/metasag. An epigenomic shift in amygdala marks the transition to maternal behaviors in alloparenting virgin female mice In many species, adults care for young offspring that are not their own, a phenomenon called alloparenting. However, most nonparental adults must be sensitized by repeated or extended exposures to newborns before they will robustly display parental-like behaviors. To capture neurogenomic events underlying the transition to active parental caring behaviors, we analyzed brain gene expression and chromatin profiles of virgin female mice co-housed with mothers during pregnancy and after birth. After an initial display of antagonistic behaviors and a surge of defense-related gene expression, we observed a dramatic shift in the chromatin landscape specifically in amygdala of the pup-exposed virgin females, accompanied by a dampening of anxiety-related gene expression. This epigenetic shift coincided with hypothalamic expression of the oxytocin gene and the emergence of behaviors and gene expression patterns classically associated with maternal care. The results outline a neurogenomic program associated with dramatic behavioral changes and suggest molecular networks relevant to human postpartum mental health. Computational Simulations of the 4-D Micro-Circulatory Network in Zebrafish Tail Amputation and Regeneration Wall shear stress (WSS) in the micro-vasculature contributes to biomechanical cues that regulate mechanotransduction underlying vascular development, regeneration, and homeostasis. We hereby elucidate the interplay between hemodynamic shear forces and luminal remodeling in response to vascular injury and regeneration in the zebrafish model of tail amputation. Using the transgenic Tg(fli1:eGFP; Gata1:ds-red) line, we were able to track the enhanced green-fluorescent protein (eGFP)-labeled endothelial lining of the 3-D microvasculature for post-image segmentation and reconstruction of fluid domain for computational fluid dynamics (CFD) simulation. At 1 day post amputation (dpa), dorsal aorta (DA) and posterior cardinal vein (PCV) were severed, and vasoconstriction developed in the dorsal longitudinal anastomotic vessel (DLAV) with a concomitant increase in WSS in the segmental vessels (SV) proximal to the amputation site and a decrease in WSS in SVs distal to amputation. Simultaneously, we observed angiogenesis commencing at the tips of the amputated DLAV and PCV where WSS was minimal in the absence of blood flow. At 2 dpa, vasodilation occurred in a pair of SVs proximal to amputation, resulting in increased flow rate and WSS, whereas in the SVs distal to amputation, WSS normalized to the baseline. At 3 dpa, the flow rate in the arterial SV proximal to amputation continued to rise and merged with DLAV that formed a new loop with PCV. Thus, our CFD modeling uncovered a well-coordinated micro-vascular adaptation process following tail amputation, accompanied by the rise and fall of WSS and dynamic changes in flow rate during vascular regeneration. Nuclear NAD+-biosynthetic enzyme NMNAT1 facilitates survival of developing retinal neurons Despite mounting evidence that the mammalian retina is exceptionally reliant on proper NAD+ homeostasis for health and function, the specific roles of subcellular NAD+ pools in retinal development, maintenance, and disease remain obscure. Here, we show that deletion of the nuclear-localized NAD+ synthase nicotinamide mononucleotide adenylyltransferase-1 (NMNAT1) in the developing murine retina causes early and severe degeneration of photoreceptors and select inner retinal neurons via multiple distinct cell death pathways. This severe phenotype is associated with disruptions to retinal central carbon metabolism, purine nucleotide synthesis, and amino acid pathways. Furthermore, large-scale transcriptomics reveals dysregulation of a collection of photoreceptor and synapse-specific genes in NMNAT1 knockout retinas prior to detectable morphological or metabolic alterations. Collectively, our study reveals previously unrecognized complexity in NMNAT1-associated retinal degeneration and suggests a yet-undescribed role for NMNAT1 in gene regulation during photoreceptor terminal differentiation. Inhibitory proteins block substrate access by occupying the active site cleft of Bacillus subtilis intramembrane metalloprotease SpoIVFB Intramembrane proteases function in numerous signaling pathways that impact health, but knowledge about regulation of intramembrane proteolysis is limited. We examined inhibition of intramembrane metalloprotease SpoIVFB by proteins BofA and SpoIVFA. We found that BofA residues in and near a predicted transmembrane segment are required for SpoIVFB inhibition, and cross-linking experiments indicated that this transmembrane segment occupies the SpoIVFB active site cleft. SpoIVFA is also required for SpoIVFB inhibition. The inhibitory proteins block access of the substrate N-terminal Proregion to the membrane-embedded SpoIVFB active site, based on additional cross-linking experiments; however, the inhibitory proteins did not prevent interaction between the substrate C-terminal region and the SpoIVFB soluble domain. A structural model was built of SpoIVFB in complex with BofA and parts of SpoIVFA and substrate, using partial homology and constraints from cross-linking and co-evolutionary analyses. The model predicts that conserved BofA residues interact to stabilize a transmembrane segment and a membrane-embedded C-terminal region. SpoIVFA is predicted to bridge the BofA C-terminal region and SpoIVFB, forming a membrane-embedded inhibition complex. Our results reveal a novel mechanism of intramembrane protease inhibition with clear implications for relief from inhibition in vivo and design of inhibitors as potential therapeutics. Confronting population models with experimental microcosm data: from trajectory matching to state-space models O_LIPopulation and community ecology traditionally has a very strong theoretical foundation with well-known models, such as the logistic and its many variations, and many modification of the classical Lotka-Volterra predator-prey and interspecific competition models. More and more, these classical models are confronted to data via fitting to empirical time-series for purposes of projections or for estimating model parameters of interest. However, the interface between mathematical population or community models and data, provided by a statistical model, is far from trivial.
C_LIO_LIIn order to help empiricists, especially researchers working with experimental laboratory populations in micro- and mesocosms, make informed decisions, we here ask which error structure one should use when fitting classical deterministic ODE models to empirical data, from single species to community dynamics and trophic interactions. We use both realistically simulated data and empirical data from microcosms to answer this question in a Bayesian framework.
C_LIO_LIWe find that many model parameters can be estimated precisely with an appropriate choice of error structure using pure observation error or state-space models, if observation errors are not too high. However, Allee effect models are typically hard to identify and state-space models should be preferred with when model complexity increases.
C_LIO_LIOur work shows that, at least in the context of experimental laboratory populations, deterministic models can be used to describe stochastic population dynamics that include process variability and observation error. Also, many scenarios do not require a complex state-space model formulation and simpler trajectory matching is sufficient for accurate parameter estimates. Finally, we provide a comprehensive tutorial for fitting these models in R.
C_LI