Overexpression of UCP4 in astrocytic mitochondria prevents multilevel dysfunctions in a mouse model of Alzheimer s disease Alzheimers disease (AD) is becoming increasingly prevalent worldwide. It represents one of the greatest medical challenge as no pharmacologic treatments are available to prevent disease progression. Astrocytes play crucial functions within neuronal circuits by providing metabolic and functional support, regulating interstitial solute composition, and modulating synaptic transmission. In addition to these physiological functions, growing evidence points to an essential role of astrocytes in neurodegenerative diseases like AD. Early stage AD is associated with hypometabolism and oxidative stress. Contrary to neurons that are vulnerable to oxydative stress, astrocytes are particularly resistant to mitochondrial dysfunction and are therefore more resilient cells. In our study, we leveraged astrocytic mitochondrial uncoupling and examined neuronal function in the 3xTg AD mouse model. We overexpressed the mitochondrial uncoupling protein 4 (UCP4), which has been shown to improve neuronal survival in vitro. We found that this treatment efficiently prevented alterations of hippocampal metabolite levels observed in AD mice, along with hippocampal atrophy and reduction of basal dendrite arborization of subicular neurons. This approach also averted aberrant neuronal excitability observed in AD subicular neurons and preserved episodic-like memory in AD mice assessed in a spatial recognition task. These findings show that targeting astrocytes and their mitochondria is an effective strategy to prevent the decline of neurons facing AD-related stress at the early stages of the disease.
The Resting-State Causal Human Connectome is Characterized by Hub Connectivity of Executive and Attentional Networks We demonstrate a data-driven approach for calculating a "causal connectome" of directed connectivity from resting-state fMRI data using a greedy adjacency search and pairwise non-Gaussian edge orientations. We used this approach to construct n=442 causal connectomes. These connectomes were very sparse in comparison to typical Pearson correlation-based graphs (roughly 2.25% edge density) yet were fully connected in nearly all cases. Prominent highly connected hubs of the causal connectome were situated in attentional (dorsal attention) and executive (frontoparietal and cingulo-opercular) networks. These hub networks had distinctly different connectivity profiles: attentional networks shared incoming connections with sensory regions and outgoing connections with higher cognitive networks, while executive networks primarily connected to other higher cognitive networks and had a high degree of bidirected connectivity. Virtual lesion analyses accentuated these findings, demonstrating that attentional and executive hub networks are points of critical vulnerability in the human causal connectome. These data highlight the central role of attention and executive control networks in the human cortical connectome and set the stage for future applications of data-driven causal connectivity analysis in psychiatry.
Regulatory perturbations of ribosome allocation optimize the growth proteome with a trade-off in adaptation capacity Bacteria regulate their cellular resource allocation to enable fast growth-adaptation to a variety of environmental niches. We studied the ribosomal allocation, growth and expression profiles of two sets of fast-growing mutants of Escherichia coli K-12 MG1655 in glucose minimal medium. Mutants with only 3 of the seven copies of ribosomal RNA operons grew faster than the wild-type strain in minimal media and show similar phenotype to previously studied rpoB mutants. Higher growth rates due to increased ribosome content affected resource allocation. Expression profiles of fast-growing mutants shared downregulation of hedging functions and upregulated growth functions. Mutants showed longer diauxic shifts and reduced activity of gluconeogenic promoters during glucose-acetate shifts, suggesting reduced availability of the RNA Polymerase for expressing hedging proteome. These results show that the regulation of ribosomal allocation underlies the growth/hedging phenotypes obtained from laboratory evolution experiments. We show how two different regulatory perturbations (rRNA promoters or rpoB mutations) reshape the proteome for growth with a concomitant fitness cost

HighlightsMutants with only 3 ribosomal operons grow faster than wild-type in minimal medium

 {Delta}4 rrn and rpoB mutants share phenotypic traits

Faster growth of mutants is achieved by increased ribosome content

Fast-growing mutants display reduced hedging expression and adaptation trade-offs

Despite similar ribosomal content in rich medium the mutants present growth defects 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. Synthetic periphyton as a model system to understand species dynamics in complex microbial freshwater communities Phototrophic biofilms, also known as periphyton, are microbial freshwater communities that drive crucial ecological processes in streams and lakes. Gaining a deep mechanistic understanding of the biological processes occurring in natural periphyton remains challenging due to the high complexity and variability of such communities. To address this challenge, we rationally developed a workflow to construct a synthetic community by co-culturing 26 phototrophic species (i.e., diatoms, green algae and cyanobacteria) that were inoculated in a successional sequence to create a periphytic biofilm on glass slides. We show that this community is diverse, stable and highly reproducible in terms of microbial composition, function and 3D spatial structure of the biofilm. We also demonstrate the ability to monitor microbial dynamics at the single species level during periphyton development and how their abundances are impacted by stressors such as increased temperature and a herbicide, singly and in combination. Overall, such a synthetic periphyton, grown under controlled conditions, can be used as a model system for theory testing through targeted manipulation. Population differences in the length and early-life dynamics of telomeres among European pied flycatchers Telomere length and shortening rate are increasingly used as biomarkers for long-term costs in ecological and evolutionary studies because of their relationships with survival and fitness. Both early-life conditions and growth, and later-life stressors can create variation in telomere shortening rate. Studies on between- population telomere length and dynamics are scarce, despite the expectation that populations exposed to varying environmental constraints would present divergent telomere length patterns. The pied flycatcher (Ficedula hypoleuca) is a passerine bird breeding across Eurasia (from Spain to western Siberia) and migrating through the Iberian Peninsula to spend the non-breeding period in sub-Saharan Africa. Thus, different populations show marked differences in migration distance. We studied the large-scale variation of telomere length and early-life dynamics in the pied flycatcher by comparing six European populations across a north- south gradient (Finland, Estonia, England, and Spain) predicting negative effect of migration distance on adult telomere length, and of nestling growth on nestling telomere dynamics. There were clear population differences in telomere length, with English birds from mid-latitudes having the longest telomeres. Telomere length did not thus show consistent latitudinal variation and was not linearly linked to differences in migration distance. Early-life telomere shortening rate tended to vary between populations. Fast growth was associated with shorter telomeres in the early life, but faster nestling growth affected telomeres more negatively in northern than southern populations. While the sources of between-population differences in telomere-related biology remain to be more intensively studied, our study illustrates the need to expand telomere studies at the between-population level. TeamTree analysis: a new approach to evaluate scientific production Advances in science and technology depend on the work of research teams and the publication of results through peer-reviewed articles representing a growing socio-economic resource. Current methods to mine the scientific literature regarding a field of interest focus on content, but the workforce credited by authorship remains largely unexplored, and appropriate measures of scientific production are debated. Here, a new bibliometric approach named TeamTree analysis is introduced that visualizes the development and composition of the workforce driving a field. A new citation-independent measure that scales with the H index estimates impact based on publication record, genealogical ties and collaborative connections. This author-centered approach complements existing tools to mine the scientific literature and to evaluate research across disciplines. A northern range extension of a Canadian species of Special Concern, Dielis pilipes (Hymenoptera: Scoliidae), in the Okanagan Valley of British Columbia The only known Canadian records of the yellow scarab hunter wasp, Dielis pilipes (Saussure) (Hymenoptera: Scoliidae), are from the southern Okanagan and Similkameen valleys of British Columbia. We report a 25-km northern range extension of the species into the Ponderosa Pine Biogeoclimatic Ecosystem Classification zone, collected in an unmanaged agricultural field in Summerland, British Columbia. This finding is of conservation importance and has implications for natural biological control of ten-lined June beetles, Polyphylla decemlineata (Say) and P. crinita LeConte (Coleoptera: Scarabaeidae), incidental agricultural pests in the Okanagan. Marine gregarine genomes reveal the breadth of apicomplexan diversity and provide new insights on gliding motility Our current view of the evolutionary history, coding and adaptive capacities of Apicomplexa, protozoan parasites of a wide range of metazoan, is currently strongly biased toward species infecting humans, as data on early diverging apicomplexan lineages infecting invertebrates is extremely limited. Here, we characterized the genome of the marine eugregarine Porospora gigantea, intestinal parasite of Lobsters, remarkable for the macroscopic size of its vegetative feeding forms (trophozoites) and its gliding speed, the fastest so far recorded for Apicomplexa. Two highly syntenic genomes named A and B were assembled. Similar in size (~9 Mb) and coding capacity (~5300 genes), A and B genomes are 10.8% divergent at the nucleotide level, corresponding to 16-38 My in divergent time. Orthogroup analysis across 25 (proto)Apicomplexa species, including Gregarina niphandrodes, showed that A and B are highly divergent from all other known apicomplexan species, revealing an unexpected breadth of diversity. Phylogenetically these two species branch sister to Cephaloidophoroidea, and thus expand the known crustacean gregarine superfamily. The genomes were mined for genes encoding proteins necessary for gliding, a key feature of apicomplexans parasites, currently studied through the molecular model called glideosome. Sequence analysis shows that actin-related proteins and regulatory factors are strongly conserved within apicomplexans. In contrast, the predicted protein sequences of core glideosome proteins and adhesion proteins are highly variable among apicomplexan lineages, especially in gregarines. These results confirm the importance of studying gregarines to widen our biological and evolutionary view of apicomplexan species diversity, and to deepen our understanding of the molecular bases of key functions enabling parasitism, such as the glideosome. SARS-CoV-2 variants of concern remain dependent on IFITM2 for efficient replication in human lung cells It has recently been shown that an early SARS-CoV-2 isolate (NL-02-2020) hijacks interferon-induced transmembrane proteins (IFITMs) for efficient replication in human cells. To date, several "Variants of Concern" (VOCs) showing increased infectivity and resistance to neutralization have emerged and globally replaced the early viral strains. Here, we determined whether the four SARS-CoV-2 VOCs (Alpha, Beta, Gamma and Delta) maintained the dependency on IFITM proteins for efficient replication. We found that depletion of IFITM2 strongly reduces viral RNA production by all four VOCs in the human epithelial lung cancer cell line Calu-3. Silencing of IFITM1 had little effect, while knock-down of IFITM3 resulted in an intermediate phenotype. Strikingly, depletion of IFITM2 generally reduced infectious virus production by more than four orders of magnitude. In addition, an antibody directed against the N-terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in iPSC-derived alveolar epithelial type II cells thought to represent major viral target cells in the lung. In conclusion, endogenously expressed IFITM proteins (especially IFITM2) are critical cofactors for efficient replication of genuine SARS-CoV-2 VOCs, including the currently dominating Delta variant.

IMPORTANCERecent results showed that an early SARS-CoV-2 isolate requires endogenously expressed IFITM proteins for efficient infection. However, whether IFITMs are also important cofactors for infection of emerging SARS-CoV-2 VOCs that out-competed the original strains and currently dominate the pandemic remained to be determined. Here, we demonstrate that depletion of endogenous IFITM2 expression almost entirely prevents the production of infectious Alpha, Beta, Gamma and Delta VOC SARS-CoV-2 virions in a human lung cell line. In comparison, silencing of IFITM1 had little impact, while knock-down of IFITM3 had intermediate effects on viral replication. Finally, an antibody targeting the N-terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in iPSC-derived alveolar epithelial type II cells. Our results show that SARS-CoV-2 VOCs including the currently dominant Delta variant are dependent on IFITM2 for efficient replication suggesting that IFITM proteins play a key role in viral transmission and pathogenicity. The pH Dependence of Niclosamide Solubility, Dissolution, and Morphology Motivates Potentially Universal Mucin-Penetrating Nasal and Throat Sprays for COVID19, its Contagious Variants, and Other Respiratory Viral Infections MotivationWith the coronavirus pandemic still raging, prophylactic nasal and early treatment throat sprays could help prevent infection and reduce viral load. Niclosamide has the potential to treat a broad range of viral infections if local bioavailability is optimized as mucin-penetrating solutions as opposed to more traditional microparticle-based sprays that cannot penetrate the mucin.

ExperimentalpH-dependence of supernatant concentrations and dissolution rates of niclosamide were measured in buffered solutions by Nanodrop-UV/Vis-spectroscopy for niclosamide from different suppliers, as precipitated material, and as cosolvates. Data was compared to predictions from Henderson Hasselbalch and precipitation pH models. Optimal microscopy was used to observe the morphologies of precipitated and converted niclosamide.

ResultsSupernatant-concentrations of niclosamide increased with increasing pH, from 1.77uM at pH 3.66 to 30uM at pH 8, and more rapidly from 90uM at pH8.5 to 300uM at pH 9.1, reaching 641uM at pH 9.5. Logarithmic rates for dissolution increased by [~]3x for pHs 8.62 to 9.44. However, when precipitated from supersaturated solution, niclosamide equilibrated to much lower final supernatant concentrations, reflective of more stable polymorphs at each pH that were also apparent for niclosamide from other suppliers and cosolvates.

ConclusionsGiven niclosamides activity against COVID19, its more contagious variants, and other respiratory viral infections, these niclosamide solutions, that put the virus in lockdown, could represent universal prophylactic nasal and early treatment throat sprays. As solutions they would be the simplest and potentially most effective formulations from both an efficacy standpoint as well as manufacturing and distribution, with no cold chain. They now just need testing. Once-daily feeding is associated with better health in companion dogs: Results from the Dog Aging Project A variety of diets have been studied for possible anti-aging effects. In particular, studies of intermittent fasting and time-restricted feeding in laboratory rodents have found evidence of beneficial health outcomes. Companion dogs represent a unique opportunity to study diet in a large mammal that shares human environments. The Dog Aging Project has been collecting data on thousands of companion dogs of all different ages, sizes, and breeds since 2019. We leveraged this diverse cross-sectional dataset to investigate associations between feeding frequency and cognitive function (n = 10,474) as well as nine broad categories of health conditions (n = 24,238). Controlling for sex, age, breed, and other potential confounders, we found that dogs fed once daily rather than more frequently had lower mean scores on a cognitive dysfunction scale, and lower odds of having gastrointestinal, dental, orthopedic, kidney/urinary, and liver/pancreas disorders. Therefore, we find that once-daily feeding is associated with better health in multiple domains. Future research with longitudinal data can provide stronger evidence for a possible causal effect of feeding frequency on health in companion dogs. Proton-driven alternating access in a spinster transporter, an emerging family of broad-specificity efflux pumps Spinster (Spns) lipid transporters are critical for transporting sphingosine-1-phosphate (S1P) across cellular membranes. In humans, Spns2 functions as the main S1P transporter in endothelial cells, making it a potential drug target for modulating S1P signaling. We employed an integrated approach in lipid membranes to identify unknown conformational states of a bacterial Spns from Hyphomonas neptunium (HnSpns) and to define its proton- and substrate-coupled conformational dynamics. Our systematic study reveals conserved residues critical for protonation steps and their regulation, and how sequential protonation of these proton switches coordinates the conformational transitions in the context of a noncanonical ligand-dependent alternating access. A conserved periplasmic salt bridge (Asp60TM2:Arg289TM7) keeps the transporter in a closed conformation, while proton-dependent conformational dynamics are significantly enhanced on the periplasmic side, providing a pathway for ligand exchange. Furthermore, our resistance assays reveal substrate polyspecificity and HnSpns multidrug resistance (MDR) activity that underscore the previously unknown role of Spns proteins in MDR, beyond their activity in sphingolipid transport and signaling. Embedded 3D printing in self-healing annealable composites for precise patterning of functionally mature human neural constructs Human in vitro models of neural tissue with controllable cellular identity, tunable microenvironment, and defined spatial arrangement are needed to facilitate studies of brain development and disease. Towards this end, embedded printing in jammed microgel supports (i.e., granular gels) holds great promise as it allows precise and programmable patterning of extremely soft and compliant tissue constructs. However, in contrast to the vast material landscape available for bulk hydrogels, granular printing support formulations are restricted to a handful of materials without the ability for facile adjustment of biofunctional properties of the cellular microenvironment. Therefore, there has been a need for novel materials that take advantage of versatile biomimicry of bulk hydrogels while providing high-fidelity support for embedded printing akin to granular gels. To address this need, we present a modular platform for bioengineering of neuronal networks via direct embedded 3D printing of human stem cells inside Self-Healing Annealable Particle-Extracellular matrix (SHAPE) composites. SHAPE composites consist of soft microgels immersed in viscous extracellular-matrix solution to enable precise freeform patterning of human stem cells and consequent generation and long-term maintenance of mature subtype-specific neurons that extend projections within the volume of the annealed support. The developed approach further allows multi-ink deposition, live spatial and temporal monitoring of oxygen levels, as well as creation of vascular channels. Due to its modularity, SHAPE biomanufacturing toolbox not only offers a solution for functional modeling of mechanically sensitive neural constructs, but also has potential to be applied to a wide range of biomaterials with different crosslinking mechanisms to model tissues and diseases where recapitulation of complex architectural features and topological cues is essential. SUFI: An automated approach to spectral unmixing of fluorescent multiplex images captured in mouse and postmortem human brain tissues Multispectral fluorescence imaging coupled with linear unmixing is a form of image data collection and analysis that uses multiple fluorescent dyes - each measuring a specific biological signal - that are simultaneously measured and subsequently "unmixed" to provide a read-out for each individual signal. This strategy allows for measuring multiple signals in a single data capture session - for example, multiple proteins or RNAs in tissue slices or cultured cells, but can often result in mixed signals and bleed-through problems across dyes. Existing spectral unmixing algorithms are not optimized for challenging biological specimens such as postmortem human brain tissue, and often require manual intervention to extract spectral signatures. We therefore developed an intuitive, automated, and flexible package called SUFI: spectral unmixing of fluorescent images (https://github.com/LieberInstitute/SUFI). This package unmixes multispectral fluorescence images by automating the extraction of spectral signatures using Vertex Component Analysis, and then performs one of three unmixing algorithms derived from remote sensing. We demonstrate these remote sensing algorithms performance on four unique biological datasets and compare the results to unmixing results obtained using ZEN Black software (Zeiss). We lastly integrate our unmixing pipeline into the computational tool dotdotdot that is used to quantify individual RNA transcripts at single cell resolution in intact tissues and perform differential expression analysis of smFISH data, and thereby provide a one-stop solution for multispectral fluorescence image analysis and quantification. In summary, we provide a robust, automated pipeline to assist biologists with improved spectral unmixing of multispectral fluorescence images. A flexible method for estimating tip diversification rates across a range of speciation and extinction scenarios Estimates of diversification rates at the tips of a phylogeny provide a flexible approach for correlation analyses with multiple traits and to map diversification rates in space, while also avoiding the uncertainty of deep time rate reconstructions. Available methods for tip rate estimation make different assumptions, and thus their accuracy usually depends on characteristics of the underlying model generating the tree. Here we introduce MiSSE, a trait-free, state-dependent speciation and extinction approach that can be used to estimate varying speciation, extinction, net-diversification, turnover rates, and extinction fraction at the tips of the tree. We compare the accuracy of tip rates inferred by MiSSE against similar methods and demonstrate that, due to certain characteristics of the model, the error is generally low across a broad range of speciation and extinction scenarios. MiSSE can be used alongside regular phylogenetic comparative methods in trait related diversification hypotheses, and we also describe a simple correction to avoid pseudoreplication from sister tips in analyses of independent contrasts. Finally, we demonstrate the capabilities of MiSSE, with a renewed focus on classic comparative methods, to examine correlation between plant height and turnover rates in eucalypts, a species-rich lineage of flowering plants. The DC1 domain protein BINUCLEATE POLLEN is required for pollen development in Arabidopsis thaliana Development of the male gametophyte is a tightly regulated process that requires precise control of cell division and gene expression. A relevant aspect to understand the events underlying pollen development regulation constitutes the identification and characterization of the genes required for this process. In this work we showed that the DC1 domain protein BINUCLEATE POLLEN (BNP) is essential for pollen development and germination. Pollen grains carrying the defective BNP allele failed to complete mitosis II and are impaired in pollen germination. By yeast two hybrid screening and bimolecular fluorescence complementation assays, we identified a set of BNP-interacting proteins. Among confirmed interactors we found NAC family transcriptional regulators Vascular plant One-Zinc finger 1 (VOZ1) and VOZ2. VOZ1 localization changes during pollen development, moving to the vegetative nucleus at the tricellular stage. We observed that this relocalization requires BNP, as in their absence in pollen from bnp/BNP plants, VOZ1 nuclear localization resulted impaired. As voz1voz2 double mutants showed the same developmental defect observed in bnp pollen grains, we propose that BNP requirement to complete microgametogenesis could be linked to its interaction with VOZ1/2 proteins. BNP could have a role of scaffold protein, recruiting VOZ1/2 to the endosomal system into assemblies that are required for their further translocation to the nucleus, where they act as transcriptional regulators. Regulatory T cells suppress the formation of potent KLRK1 and IL-7R expressing effector CD8 T cells by limiting IL-2 Regulatory T cells (Tregs) are indispensable for maintaining self-tolerance by suppressing conventional T cells. On the other hand, Tregs may promote tumor growth by inhibiting anti-cancer immunity. In this study, we identified that Tregs increase the quorum of self-reactive CD8+ T cells required for the induction of experimental autoimmune diabetes. Their major suppression mechanism is limiting available IL-2, an essential T-cell cytokine. Specifically, Tregs inhibit the formation of a previously uncharacterized subset of antigen-stimulated KLRK1+ IL7R+ (KILR) CD8+ effector T cells, which are distinct from conventional effector CD8+ T cells. KILR CD8+ T cells show a superior cell killing abilities in vivo. The administration of agonistic IL-2 immunocomplexes phenocopies the absence of Tregs, i.e., it induces KILR CD8+ T cells, promotes autoimmunity, and enhances anti-tumor responses. Counterparts of KILR CD8+ T cells were found in the human blood, revealing them as a potential target for immunotherapy. Enhancers regulate 3' end processing activityto control expression of alternative 3'UTR isoforms Multi-UTR genes are widely transcribed but show cell type-specific 3'UTR isoform expression. As transcriptional enhancers regulate mRNA expression, we investigated if they also regulate mRNA isoform expression. Deletion of an endogenous enhancer of a multi-UTR gene did not impair transcript production but prevented a switch in 3'UTR isoform expression. Also, the same enhancers are able to increase transcript production in the context of single-UTR gene promoters, but they increase 3' end processing activity when paired with multi-UTR gene promoters. We show that transcription factors regulate processing activity of weak polyadenylation sites to control cell type-specific alternative 3'UTR isoform expression of widely expressed genes. SufB intein splicing in Mycobacterium tuberculosis is influenced by two remote conserved N-extein Histidines Inteins are auto-processing domains that implement a multi-step biochemical reaction termed protein splicing, marked by cleavage and formation of peptide bonds. They excise from a precursor protein, generating a functional protein via covalent bonding of flanking exteins. We report the kinetic study of splicing and cleavage reaction in [Fe-S] cluster assembly protein SufB from Mycobacterium tuberculosis. Although it follows a canonical intein splicing pathway, distinct features are added by extein residues present in the active site. Sequence analysis identified two conserved histidines in the N-extein region; His-5 and His-38. Kinetic analyses of His-5Ala and His-38Ala SufB mutants exhibited significant reductions in splicing and cleavage rates relative to the SufB wild-type precursor protein. Structural analysis and molecular dynamics simulations suggested that Mtu SufB displays a unique mechanism where two remote histidines work concurrently to facilitate N-terminal cleavage reaction. His-38 is stabilized by the solvent-exposed His-5, and can impact N-S acyl shift by direct interaction with the catalytic Cys1. Development of inteins as biotechnological tools or as pathogen specific novel antimicrobial targets requires a more complete understanding of such unexpected roles of conserved extein residues in protein splicing. 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. RASopathy Mutations Demonstrate the Critical Function of the Cysteine-rich Domain in Maintaining BRAF in an Autoinhibited State to Prevent Aberrant Signaling The RAF kinases are essential effectors of the RAS-GTPases, and germline mutations in BRAF or CRAF are disease drivers in the RASopathy developmental syndromes, with mutations in the cysteine-rich domain (CRD) often observed in BRAF but not CRAF. Although the CRD is known for roles in phosphatidylserine (PS) binding, the RAS-RAF interaction, and RAF autoinhibition, the interplay of these activities and their impact on BRAF function in normal and disease states is unclear. Here, we analyze a panel of BRAF-CRD RASopathy mutations and show that they all increase the biological activity of BRAF by relieving autoinhibition and/or enhancing PS/membrane binding. Strikingly, we find that relief of autoinhibition is the predominant factor determining the severity of the mutations. Comparison of the BRAF and CRAF CRDs further indicates that the BRAF-CRD is a stronger mediator of both autoinhibition and PS binding, and we identify residues contributing to these differences. Moreover, given the increased catalytic activity of BRAF versus CRAF, our studies reveal a more critical role for the CRD and autoinhibition in BRAF regulation and demonstrate that CRD-mediated autoinhibition is required to prevent aberrant BRAF signaling, a finding consistent with the high frequency of mutations that disrupt this function in the RASopathies. Key features of the genetic architecture and evolution of host-microbe interactions revealed by high-resolution genetic mapping of the mucosa-associated gut microbiome in hybrid mice Determining the forces that shape diversity in host-associated bacterial communities is critical to understanding the evolution and maintenance of metaorganisms. To gain deeper understanding of the role of host genetics in shaping gut microbial traits, we employed a powerful genetic mapping approach using inbred lines derived from the hybrid zone of two incipient house mouse species. Further, we uniquely performed our analysis on microbial traits measured at the gut mucosal interface, which is in more direct contact with host cells and the immune system. A high number of mucosa-associated bacterial taxa have significant heritability estimates; heritabilities are greater for 16S rRNA transcript-compared to gene copy-based traits, and interestingly, are positively correlated with cospeciation rate estimates. Genomewide association mapping identifies 443 loci influencing 123 taxa, with narrow genomic intervals pinpointing promising candidate genes and pathways. Importantly, we identified an enrichment of candidate genes associated with several human diseases, including inflammatory bowel disease, and functional categories including innate immunity and G-protein-coupled receptors. These results highlight key features of the genetic architecture of mammalian host-microbe interactions and how they diverge as new species form. The Temporal Statistics of Musical Rhythm across Western Genres: An Amplitude Modulation Phase Hierarchy Model Statistical learning by the human brain plays a core role in the development of cognitive systems like language and music. Both music and speech have structured inherent rhythms, however the acoustic sources of these rhythms are debated. Theoretically, rhythm structures in both systems may be related to a novel set of acoustic statistics embedded in the amplitude envelope, statistics originally revealed by modelling childrens nursery rhymes. Here we apply similar modelling to explore whether the amplitude modulation (AM) timescales underlying rhythm in music match those in child-directed speech (CDS). Utilising AM-driven phase hierarchy modelling previously applied to infant-directed speech (IDS), adult-directed speech (ADS) and CDS, we test whether the physical stimulus characteristics that yield speech rhythm in IDS and CDS describe rhythm in music. Two models were applied. One utilized a low-dimensional representation of the auditory signal adjusted for known mechanisms of the human cochlear, and the second utilized probabilistic amplitude demodulation, estimating the modulator (envelope) and carriers using Bayesian inference. Both models revealed a similar hierarchically-nested temporal modulation structure across Western musical genres and instruments. Core bands of AM and spectral patterning matched prior analyses of IDS and CDS, and music showed strong phase dependence between slower bands of AMs, again matching IDS and CDS. This phase dependence is critical to the perception of rhythm. Control analyses modelling other natural sounds (wind, rain, storms, rivers) did not show similar temporal modulation structures and phase dependencies. We conclude that acoustic rhythm in language and music has a shared statistical basis. Spontaneous differentiation leads to emergence of hybrid cell states relate to poor prognosis in oral cancer PurposeCellular dynamics between phenotypically heterogeneous subpopulations of cancer cells within individual tumor is shown to be responsible for drug tolerance and overall poor prognosis; however, evidences were largely missing in oral cancer. Therefore, this study was undertaken to describe the dynamic phenotypic states among oral cancer cells, its influence on transcriptomic heterogeneity as well as its clinical significance.

Experimental DesignWe multiplexed phenotypic markers of putative oral-stem-like cancer cells (SLCCs) and characterized diversity among CD44-positive oral cancer cell subpopulations with respect to distinct expression of CD24 and aldehyde dehydrogenase (ALDH)-activity in multiple cell lines. Population trajectories were characterized by Markov model and cell states were defined based on the population specific RNA sequencing (RNAseq). ssGSEA based gene expression signatures were explore for prognostic significance.

ResultsOral cancer cells followed two distinct patterns of spontaneous repopulation dynamics with stochastic inter-conversions on  ALDH-axis, however a strict non-interconvertible transition on  CD24-axis. Interestingly, plastic  ALDH-axis was harnessed to enrich ALDHHigh subpopulations in response to Cisplatin treatment, to adapt a drug tolerant state. Phenotype-specific RNAseq results suggested the organization of subpopulations into hierarchical structure with possible maintenance of intermediate states of stemness within the differentiating oral cancer cells. Further, survival analysis with each subpopulation-specific gene signature strongly suggested that the cell-state dynamics may act as possible mechanism to drive ITH, resulting in poor prognosis in patient.

ConclusionsOur results emphasized the prognostic power of the population dynamics in oral cancer. Importantly, we have described the phenotypic-composition of heterogeneous subpopulations critical for global tumor behaviour in oral cancer; which is a prerequisite knowledge important for precision treatment, however largely lacking for most solid tumors.

Graphical AbstractWe have characterized diversity among CD44-positive oral cancer cells lines with respect to distinct expression of CD24 and ALDH-activity. Subpopulations showed stochastic inter-conversions on ALDH-axis but a strict non-interconvertible transition of CD24Low to CD24High phenotype, even in response to chemotherapy-induced stress. RNAseq study suggested the organization of subpopulations into hierarchical structure with possible maintenance of intermediate alternate states of stemness within the differentiating oral cancer cells. The described population dynamics demonstrtaed influence tumor behaviour possibly by increasing intratumoral heterogeneity in aggressive oral tumors.



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org.highwire.dtl.DTLVardef@1dc6a7org.highwire.dtl.DTLVardef@dd0948org.highwire.dtl.DTLVardef@18c9603org.highwire.dtl.DTLVardef@cca8fb_HPS_FORMAT_FIGEXP  M_FIG C_FIG Translational relevanceIntratumoral heterogeneity (ITH) has been the clinically important factor, impacting aggressive cancer behaviour, drug tolerance and overall poor prognosis. Recent high-throughput studies have provided better cellular and molecular resolution of ITH; however, the prerequisite knowledge which defines the composition of subpopulations critical for global tumor behaviour is majorly lacking for most of the solid tumors. By combining phenotypic markers, we have defined four subpopulations of oral cancer cells. These subpopulations showed stochastic inter-conversions as well as a strict non-interconvertible transition among them to acheive heterogeneity. Importantly, transcriptional states of each subpopulations indicated a clinically relevant signatures for patient prognosis. Also, we observed interconversions of these subpopulations in response to Cisplatin to accumulate drug-tolerant cell state, as rapid and reversible strategy to respond to chemotherapy induced stress. Thus, the characteristics of described phenotypic subgroups may be translated to the clinic for estimating the extent of intratumoral heterogeneity in oral cancer patients. Characterization of methylation profiles in spontaneous preterm birth placental villous tissue Preterm birth is a global public health crisis which results in significant neonatal and maternal mortality. Yet little is known regarding the molecular mechanisms of idiopathic spontaneous preterm birth, and we have few diagnostic markers for adequate assessment of placental development and function. Previous studies of placental pathology and our transcriptomics studies suggest a role for placental maturity in idiopathic spontaneous preterm birth. It is known that placental DNA methylation changes over gestation. We hypothesized that if placental hypermaturity is present in our samples, we would observe a unique idiopathic spontaneous preterm birth DNA methylation profile potentially driving the gene expression differences we previously identified in our placental samples. Our results indicate the idiopathic spontaneous preterm birth DNA methylation pattern mimics the term birth methylation pattern suggesting hypermaturity. Only seven significant differentially methylated regions fitting the idiopathic spontaneous preterm birth specific (relative to the controls) profile were identified, indicating unusually high similarity in DNA methylation between idiopathic spontaneous preterm birth and term birth samples. We identified an additional 1,718 significantly methylated regions in our gestational age matched controls were the idiopathic spontaneous preterm birth DNA methylation pattern mimics the term birth methylation pattern, again indicating a striking level of similarity between the idiopathic spontaneous preterm birth and term birth samples. Pathway analysis of these regions revealed differences in genes within the WNT and Cadherin signaling pathways, both of which are essential in placental development and maturation. Taken together, these data demonstrate that the idiopathic spontaneous preterm birth samples are molecularly more mature than expected given their respective gestational age which likely impacts birth timing.