title: Developing all-in-one virus-like particles for Cas9 mRNA/single guide RNA co-delivery and aptamer-containing lentiviral vectors for improved gene expression

abstract: Lentiviral vectors (LVs) are widely used for delivering foreign genes for long-term expression. Normal LVs contain the RNA genome, which is reverse transcribed into DNA and integrates into the host genome to mediate long-term gene expression. Recently, virus-like particles (VLPs) were developed for mRNA delivery. In these VLPs, packaging mRNA into the particles is achieved via interactions between the aptamer and aptamer-binding protein (ABP), and mRNA is not reverse transcribed or integrated. These VLPs are useful for delivering Cas9 mRNA for short-term endonuclease expression. Generating high-titer normal integrating LVs is challenging. Until recently, VLPs were not efficient for co-delivering Cas9 mRNA and single guide RNA (sgRNA). By fusing ABP to the N-terminus of HIV Gag protein, hybrid particles were developed to co-deliver Cas9 mRNA and sgRNA. But the method for modifying Gag impaired particle assembly. Previously we found that adding an ABP after the second zinc finger domain of nucleocapsid (NC) protein had minimal effects on particle assembly. Based on this observation, we developed hybrid particles to co-deliver Cas9 mRNA and sgRNA. We further improved LVs for integrated gene expression by including an aptamer sequence in lentiviral transfer plasmids, which improved lentiviral particle production and enhanced LV genomic RNA packaging. In summary, we describe development of new all-in-one VLPs for co-delivery of Cas9 mRNA and sgRNA and new LVs for enhanced vector production and gene expression.


title: Reconstitution of an active human CENP-E motor

abstract: CENP-E is a large kinesin motor protein which plays pivotal roles in mitosis by facilitating chromosome capture, alignment and promoting microtubule flux in the spindle. So far, it has not been possible to obtain active human CENP-E to study its molecular properties. Xenopus CENP-E motor has been characterised in vitro and is used as a model motor, however its protein sequence differs significantly from human CENP-E. Here, we characterise human CENP-E motility in vitro. Full-length CENP-E exhibits an increase in run length and longer residency times on microtubules when compared to CENP-E motor truncations, indicating that the C-terminal microtubule binding site enhances the processivity when the full-length motor is active. In contrast to constitutively active human CENP-E truncations, full-length human CENP-E has a reduced microtubule landing rate in vitro, suggesting that the non-motor coiled coil regions self-regulate motor activity. Together, we demonstrate that human CENP-E is a processive motor, providing a useful tool to study the mechanistic basis for how human CENP-E drives chromosome congression and spindle organisation during human cell division.

category: biochemistry


title: PROTAC-mediated selective degradation of cytosolic soluble epoxide hydrolase enhances ER-stress reduction

abstract:
Soluble epoxide hydrolase (sEH) is a bifunctional enzyme responsible for lipid metabolism and is a promising drug target. Here, we report the first-in-class PROTACs small-molecule degraders of sEH. Our optimized PROTAC selectively targets the degradation of cytosolic but not peroxisomal sEH, resulting in exquisite spatiotemporal control. In comparison to the parent sEH inhibitor, sEH-directed PROTAC has higher potency in cellular assays, as measured by significantly reduced ER stress. Surprisingly, PROTAC directs degradation of cytosolic sEH via the lysosome, not through the proteasome, representing the first example of an intracellular lysosomal-dependent PROTAC. The molecules presented here are useful chemical probes to study the biology of sEH with the potential for therapeutic development. Broadly, our results represent a proof-of-concept for the subcellular compartment-selective modulation of a protein class by PROTACs, as well as assisting in the establishment of the cellular mechanism of PROTACs that preferentially direct target proteins for lysosomal degradation.

HighlightsO_LIFirst-in-class soluble epoxide hydrolase (sEH) small-molecule degraders.
C_LIO_LISelective degradation of cytosolic but not peroxisomal sEH.
C_LIO_LISignificant and stable reduction in sEH protein levels, leading to enhanced cellular efficacy in ER stress reduction relative to the parent inhibitor.
C_LIO_LIFirst example of a PROTAC that induces lysosomal-directed degradation of a soluble target protein.
C_LI


title: Global analysis of biosynthetic gene clusters reveals conserved and unique natural products in entomopathogenic nematode-symbiotic bacteria

abstract: Microorganisms contribute to the biology and physiology of eukaryotic hosts and affect other organisms through natural products. Xenorhabdus and Photorhabdus (XP) living in mutualistic symbiosis with entomopathogenic nematodes produce a myriad of natural products to mediate bacteria-nematode-insect interactions. However, a lack of systematic analysis of the biosynthetic gene clusters (BGCs) has limited the understanding of how natural products justify the bacterial niche specificity. Here we combine pangenome and sequence similarity networks to analyze BGCs from 45 XP species. The identified 1,000 BGCs belong to 176 families, over half of which are unknown. Eleven BGCs represent the most conserved families. We then homologously express the ubiquitous and unique BGCs and identify compounds featuring unusual architectures. The bioactivity evaluation demonstrates that the prevalent compounds are eukaryotic proteasome inhibitors, insect virulence factors, or insect immune suppressors. These findings account for the functional basis of bacterial natural products in this tripartite relationship.


title: Nanoformulated Remdesivir with Extremely Low Content of Poly(2-oxazoline) - Based Stabilizer for Aerosol Treatment of COVID-19

abstract: The rise of the novel virus SARS-CoV2 which causes the disease known as COVID-19 has led to a global pandemic claiming millions of lives. With no clinically approved treatment for COVID-19, physicians initially struggled to treat the disease and there is still need for improved anti-viral therapies in this area. We conceived early in the pandemic that an inhalable formulation of the drug Remdesivir which directly targets the virus at the initial site of infection could improve therapeutic outcomes in COVID-19. We developed a set of requirements that would be conducive to rapid drug approval: 1) try to use GRAS or GRAS similar reagents 2) minimize excipient concentration and 3) achieve a working concentration of 5 mg/mL Remdesivir to achieve a deliverable dose which is 5-10% of the IV dose. In this work, we discovered that Poly(2-oxazoline) block copolymers can stabilize drug nanocrystal suspensions and provide suitable formulation characteristics for aerosol delivery while maintaining anti-viral efficacy. We believe POx block copolymers can be used as a semi-ubiquitous stabilizer for the rapid development of nanocrystal formulations for new and existing diseases.

category: bioengineering


title: Deep learning based on multi-omics integration identifies potential therapeutic targets in breast cancer

abstract: Effective and precise classification of breast cancer patients for their disease risks is critical to improve early diagnosis and patient survival. In the recent past, a significant amount of multi-omics data derived from cancer patients has emerged. However, a robust framework for integrating multi-omics data to subgroup cancer patients and predict survival prognosis is still lacking. In addition, effective therapeutic targets for treating breast cancer patients with poor prognoses are in dire need. To begin to resolve this difficulty, we developed and optimized a sophisticated deep learning-based model in breast cancer that can accurately stratify patients based on their prognosis. We built a survival-associated predictive framework integrating transcription profile, miRNA expression, somatic mutations, copy number variation, DNA methylation and protein expression. This framework achieved promising performance in distinguishing high-risk breast cancer patients from those with good prognoses. Furthermore, we constructed multiple fully connected neural networks that are trained on prioritized multi-omics signatures or even only potential single-omics signatures, based on our customized scoring system. Together, the landmark multi-omics signatures we identified may serve as potential therapeutic targets in breast cancer.

category: bioinformatics


title: ReadItAndKeep: rapid decontamination of SARS-CoV-2 sequencing reads

abstract:
SummaryViral sequence data from clinical samples frequently contain human contamination, which must be removed prior to sharing for legal and ethical reasons. To enable host read removal for SARS-CoV-2 sequencing data on low-specification laptops, we developed ReadItAndKeep, a fast lightweight tool for Illumina and nanopore data that only keeps reads matching the SARS-CoV-2 genome. Peak RAM usage is typically below 10MB, and runtime less than one minute. We show that by excluding the polyA tail from the viral reference, ReadItAndKeep prevents bleed-through of human reads, whereas mapping to the human genome lets some reads escape. We believe our test approach (including all possible reads from the human genome, human samples from each of the 26 populations in the 1000 genomes data, and a diverse set of SARS-CoV-2 genomes) will also be useful for others.

Availability and implementationReadItAndKeep is implemented in C++, released under the MIT license, and available from https://github.com/GenomePathogenAnalysisService/read-it-and-keep.


title: Regulation of balanced root fungal community of Vanda falcata (Orchidaceae) by partitioning its mycorrhizal fungi and ascomycetous fungi across growth and development

abstract: Epiphytic orchids are commonly found in exposed environments, which plausibly lead to different root fungal community structures from terrestrial orchids. Until recently, no studies have been conducted to show the fungal community structure during the growth of a photosynthetic and epiphytic orchid in its natural growing state. In this study, the Vanda falcata (commonly known as Neofinetia falcata), one of Japans ornamental orchids, was used to characterize the fungal community structure at different developmental stages. Amplicon sequencing analysis showed that all development stages contain a similar fungal community: Ascomycota dominates half of the community while one-third of the community is Basidiomycota. Rhizoctonia-like fungi, a term to group the most common basidiomycetous fungi that form orchid mycorrhiza, exist even in a smaller portion (around a quarter) compared to other Basidiomycota members. While ascomycetous fungi exhibit pathogenicity, two Ceratobasidium strains isolated from young and adult plants could initiate seed germination in vitro. It was also found that the colonization of mycorrhizal fungi was concentrated in the lower part of the root where it directly attaches to the phorophyte bark, while ascomycetous fungi were distributed in the velamen but never colonized cortical cells. Additionally, lower root parts attached to the bark have denser exodermal passage cells, and these cells were colonized only by mycorrhizal fungi that further infiltrated the cortical area. Therefore, we propose that physical regulation of fungal entry to partition the ascomycetes and mycorrhizal fungi results in the balanced mycorrhizal symbiosis in this orchid.

category: plant biology


title: Engineered bacterial swarm patterns as spatial records of environmental inputs

abstract: A diverse array of bacteria species naturally self-organize into durable macroscale patterns on solid surfaces via swarming motility--a highly coordinated, rapid movement of bacteria powered by flagella1-5. Engineering swarming behaviors is an untapped opportunity to increase the scale and robustness of coordinated synthetic microbial systems. Here we engineer Proteus mirabilis, which natively forms centimeter-scale bullseye patterns on solid agar through swarming, to "write" external inputs into a visible spatial record. Specifically, we engineer tunable expression of swarming-related genes that accordingly modify pattern features, and develop quantitative approaches to decode input conditions. Next, we develop a two-input system that modulates two swarming-related genes simultaneously, and show the resulting patterns can be interpreted using a deep learning classification model. Lastly, we show a growing colony can record dynamic environmental changes, which can be decoded from endpoint images using a segmentation model. This work creates an approach for building a macroscale bacterial recorder and expands the framework for engineering emergent microbial behaviors.

category: synthetic biology


title: The effects of electroencephalogram feature-based transcranial alternating current stimulation on working memory and electrophysiology

abstract: Transcranial alternating current stimulation (tACS) can influence cognitive functions by modulating brain oscillations. However, results regarding the effectiveness of tACS in regulating cognitive performance have been inconsistent. In the present study, we aimed to find EEG characteristics associated with the improvements in working memory performance, to select tACS stimulus targets and frequency based on this feature, and to explore effects of selected stimulus on verbal working memory. To achieve this goal, we first investigated the EEG characteristics associated with improvements in working memory performance with the aid of EEG analyses and machine learning techniques. These analyses suggested that 8 Hz activity in the prefrontal region was related to accuracy in the verbal working memory task. The tACS stimulus target and pattern were then selected based on the EEG feature. Finally, the selected tACS frequency (8 Hz tACS in the prefrontal region) was applied to modulate working memory. The performance of working memory was improved significantly using the selected stimulation than using 40 Hz and sham stimulation (Especially for participants with low verbal working memory). In conclusion, using EEG features related to positive behavioral changes to select brain regions and stimulation patterns for tACS is an effective intervention for improving working memory. Our results contribute to the groundwork for future tACS closed-loop interventions for cognitive deterioration.

category: neuroscience


title: Tissue-intrinsic γδ T cells critically regulate Tissue-Resident Memory CD8 T cells

abstract: Because Tissue-Resident Memory T (TRM) cells contribute critically to body-surface immunoprotection and/or immunopathology in multiple settings, their regulation is biologically and clinically important. Interestingly, TRM commonly develop in epithelia part-shaped by innate-like lymphocytes that become tissue-intrinsic during development. Here we show that polyclonal TRM cells induced by allergic contact dermatitis (ACD) interact with signature intraepidermal {gamma}{delta} T cells, facilitating a feedback-loop wherein TRM-derived IFN{gamma} upregulates PD-L1 on {gamma}{delta} cells that can thereupon regulate PD1+ TRM. Thus, TRM induced by ACD in mice lacking either local {gamma}{delta} cells, or lacking a single gene (IFN{gamma}R) expressed by local {gamma}{delta} cells, displayed enhanced proliferative and effector potentials. Those phenotypes were associated with strikingly limited motility, reduced TRM quality. and an impaired capacity to restrain melanoma. Thus, inter-individual and tissue-specific variation in how tissue-intrinsic lymphocytes integrate with TRM may sit upstream of variation in responses to cancer, allergens and other challenges, and may likewise underpin inflammatory pathologies repeatedly observed in {gamma}{delta}-deficient animals.

category: immunology


title: Insights into the Human Gut Virome by Sampling a Population from the Indian Subcontinent

abstract:
Gut virome plays an important role in human physiology but remains poorly understood. This study reports an investigation of the human gut DNA-virome of a previously unexplored ethnic population through metagenomics of faecal samples collected from individuals residing in Northern India. Analysis shows that, similar to the populations investigated earlier, majority of the identified virome belongs to bacteriophages and a smaller fraction (< 20%) consists of viruses that infect animals, archaea, protists, multiple domains or plants. However, crAss-like phages, in this population, are dominated by the genera VII, VIII and VI. Interestingly, it also reveals the presence of a virus family, Sphaerolipoviridae, which has not been detected in the human gut earlier. Viral families, Siphoviridae, Myoviridae, Podoviridae, Microviridae, Herelleviridae and Phycodnaviridae are detected in all of the analyzed individuals, which supports the existence of a core virome. Lysogeny-associated genes were found in less than 10% of the assembled genomes and a negative correlation was observed in the richness of bacterial and free-viral species, suggesting that the dominant lifestyle of gut phage is not lysogenic. This is in contrast to some of the earlier studies. Further, several hundred high-quality viral genomes were recovered. Detailed characterization of these genomes would be useful for understanding the biology of these viruses and their significance in human physiology.

ImportanceViruses are important constituents of the human gut microbiome but it remains poorly understood. The Indian subcontinent is a unique biogeographic region and the Indian population is known to harbour a distinct bacterial microbiome. However, the gut virome in this population has not been investigated earlier. Therefore, in this study, we investigated fecal samples of 12 healthy individuals to analyze their gut virome, through metagenomics.

category: microbiology


title: The influence of resource overlap on bacterial reproductive success in the phyllosphere

abstract:
The phyllosphere is colonized by rich microbial communities, despite sparse and heterogeneously distributed resources. This resource limitation is expected to drive bacterial competition, resulting in either exclusion or coexistence based on the fitness differences and resource overlap between species. Here, we investigated the impact of competition in bacterial colonization and growth of the epiphyte Pantoea eucalypti 299R (Pe299R). To that end, pairwise competition experiments between Pe299R and diverse phyllosphere-colonizing bacteria were performed in vitro and in the Arabidopsis thaliana phyllosphere. Resource overlap was determined as the similarity in resource utilization in vitro. We found an effect of both resource overlap and phylogenetic relationships in the competition outcome between Pe299R and individual competitors in vitro. To account for bacterial individuality in the phyllosphere, we employed a single cell bioreporter to determine the number of divisions that individual cells from Pe299R populations underwent when challenged with individual bacterial competitors. We observed that at the single-cell level, resource utilization similarities and phylogenetic relationships were weakly correlated with Pe299R reproductive success. We observed contrasting results for two strains (Arthrobacter sp. Leaf145 and Sphingomonas melonis Fr1) that, despite sharing either a high or low resource overlap with Pe299R, both negatively affected the reproductive success of Pe299R. Interestingly, we also observed facilitative effects of Methylobacterium sp. Leaf85. This work furthers the understanding of bacterial assembly processes in heterogenous environments. Our high-resolution observations are important to build an ecological framework to predict competition outcomes in the phyllosphere and to design future bacterial biocontrol applications.

Significance StatementFor plant and consumer protection, plant microbiota manipulation has recently garnered much attention. To manipulate the plant microbiota while acknowledging its ecological context, understanding the mechanisms of microbial community assembly in the phyllosphere is essential. We explored the effect of resource overlap between competing species on the fitness of a focal species at the single-cell resolution using a bottom-up approach. Using a combination of in vitro and in planta pairwise co-inoculation experiments, we found that competition outcomes are associated with resource overlap and phylogenetic relationships. Although this effect is not strong enough to promote competitive exclusion, we provide examples of competitors that affect the growth of a focal species depending on their resource overlap.


title: Quantitative proteomics of the CDK9 interactome reveals a novel function of the HSP90-CDC37-P-TEFb complex for BETi-induced HIV-1 latency reactivation

abstract: Brd4 has been intensively investigated as a promising drug target because of its implicated functions in oncogenesis, inflammation and HIV-1 transcription. The formation of the Brd4-P-TEFb (CDK9/Cyclin T1) complex and its regulation of transcriptional elongation is critical for HIV latency reactivation and expression of many oncogenes. To further investigate the mechanism of the Brd4-P-TEFb complex in controlling elongation, mass spectrometry-based quantitative proteomics of the CDK9 interactome was performed. Upon treatment with the selective BET bromodomain inhibitor (BETi) JQ1, 535 proteins were successfully identified with high confidence as CDK9-interacting proteins. Among them, that increased bindings of HSP90 and CDC37 to CDK9 were particularly striking, and our data suggest that the HSP90-CDC37-P-TEFb complex is involved in controlling P-TEFbs dynamic equilibrium during BETi-induced HIV-1 latency reactivation. Furthermore, the HSP90-CDC37-P-TEFb complex directly regulates HIV-1 transcription and relies on the recruitment by heat shock factor 1 (HSF1) for binding to the HIV-1 promoter. These results advance the understanding of HSP90-CDC37-P-TEFb in HIV-1 latency reversal and enlighten the development of potential strategies to eradicate HIV-1 using a combination of targeted drugs.

category: molecular biology


title: Machine learning algorithms predict soil seed bank persistence from easily available traits

abstract:
QuestionSoil seed banks (SSB), i.e., pools of viable seeds in the soil and its surface, play a crucial role in plant biology and ecology. Information on seed persistence in soil is of great importance for fundamental and applied research, yet compiling datasets on this trait still requires enormous efforts. We asked whether the machine learning (ML) approach could be used to infer and predict SSB properties of a regional flora based on easily available data.

LocationEighteen calcareous grasslands located along an elevational gradient of almost 2000 m in the Bavarian Alps, Germany.

MethodsWe compared a commonly used ML model (random forest) with a conventional model (linear regression model) as to their ability to predict SSB presence/absence and density using empirical data on SSB characteristics (environmental, seed traits and phylogenetic predictors). Further, we identified the most important determinants of seed persistence in soil for predicting qualitative and quantitative SSB characteristics using the ML approach.

ResultsWe demonstrated that the ML model predicts SSB characteristics significantly better than the linear regression model. A single set of predictors (either environment, or seed traits, or phylogenetic eigenvectors) was sufficient for the ML model to achieve high performance in predicting SSB characteristics. Importantly, we established that a few widely available SSB predictors can achieve high predictive power in the ML approach, suggesting a high flexibility of the developed approach for use in various study systems.

ConclusionsOur study provides a novel methodological approach that combines empirical knowledge on the determinants of SSB characteristics with a modern, flexible statistical approach based on ML. It clearly demonstrates that ML can be developed into a key tool to facilitate labor-intensive, costly and time-consuming functional trait research.

category: ecology


title: Oxygen availability drives blood traits and the cell number-size tradeoff across Andean hummingbirds

abstract:
For endotherms with sustained high metabolism, like hummingbirds, blood-oxygen (O2) carrying capacity should be finely tuned to supply O2 to respiring tissues. Hematological adjustments are expected in response to changes in O2 availability that occur along elevational gradients; however, eco-physiological rules by which this variation occurs have remained elusive. If evolutionary divergence in elevational niches is accompanied by predictable shifts in blood trait reaction norms, we would expect different patterns of elevational variation for within-species versus among-species comparisons; but this has not been systematically tested. Here, we sought to elucidate these rules using hummingbirds as a model clade. We measured blood traits that affect O2-carrying functions: total hemoglobin concentration ([Hb]), hematocrit (Hct), erythrocyte count (TRBC), mean cell volume (MCV), mean cellular hemoglobin content (MCH), and mean cellular hemoglobin concentration (MCHC). For 1,217 Andean hummingbirds representing 77 species, spanning sea level to 4,578 meters in elevation, we modeled blood traits as responses to environmental conditions and individual and species traits. [Hb] and Hct increased with elevation similarly within species and across the phylogeny--for every 1,000-m ascent, [Hb] increased by one third of a gram per deciliter and Hct increased by one and a half percentage points, corresponding to about three-hundred thousand additional cells per microliter. Hummingbirds adjusted blood cell number and cell size by similar proportions to modulate blood-O2 carrying capacity; however, species at the lowest and highest elevations relied relatively more on adjustments to cell volume, revealing qualitatively different evolutionary responses elicited by moderate versus extreme reductions in O2 availability.

SIGNIFICANCE STATEMENTTrait variation along environmental gradients reveals how individuals adjust or populations evolve to improve performance, foreshadowing climate change effects. For blood traits such as hemoglobin concentration ([Hb]), rules of elevational variation have remained elusive; it is unknown whether species adaptation affects reaction of blood phenotypes to local oxygen availability. We tested this in a model clade, hummingbirds, that have undergone many elevational shifts. Elevational variation was predictable, but similar within and among species, indicating no predictable evolution of reaction norms. However, for the cell number-size tradeoff that underpins [Hb], species varied by elevational range. Cell number explained [Hb] at intermediate elevations, while cell size was more important at extremes, indicating qualitative differences in respiratory adaptation to moderate versus high elevations.

category: evolutionary biology


title: Xeno-monitoring of molecular drivers of artemisinin and partner drug resistance in P. falciparum populations in malaria vectors across Cameroon

abstract:
BackgroundMonitoring of drug resistance in Plasmodium populations is crucial for malaria control. This has primarily been performed in humans and rarely in mosquitoes where parasites genetic recombination occurs. Here, we characterized the Plasmodium spp populations in wild Anopheles vectors by analyzing the genetic diversity of the P. falciparum kelch13 and mdr1 gene fragments implicated in artemisinin and partner drug resistance across Cameroon in three major malaria vectors.

MethodsAnopheles mosquitoes were collected across nine localities in Cameroon and dissected into the head/thorax (H/T) and abdomen (Abd) after species identification. A TaqMan assay was performed to detect Plasmodium infection. Fragments of the Kelch 13 and mdr1 genes were amplified in P. falciparum positive samples and directly sequenced to assess their drug resistance polymorphisms and genetic diversity profile.

ResultsThe study revealed a high Plasmodium infection rate in the major Anopheles vectors across Cameroon. Notably, An. funestus vector recorded the highest sporozoite (8.02%) and oocyst (14.41%) infection rates. A high P. falciparum sporozoite rate (80.08%) alongside epidemiological signatures of significant P. malariae (15.94%) circulation were recorded in these vectors. Low genetic diversity with six (A578S, R575I, G450R, L663L, G453D, N458D) and eight (H53H, V62L, V77E, N86Y, G102G, L132I, H143H, Y184F) point mutations were observed in the k13 and mdr1 backbones respectively. Remarkably, the R575I (4.44%) k13 and Y184F (64.2%) mdr1 mutations were the predominant variants in the P. falciparum populations.

ConclusionThe emerging signal of the R575I polymorphism in the Pfk13 propeller backbone entails the regular surveillance of molecular markers to inform evidence-based policy decisions. Moreover, the high frequency of the 86N184F allele highlights concerns on the plausible decline in efficacy of artemisinin-combination therapies (ACTs); further implying that parasite genotyping from mosquitoes can provide a more relevant scale for quantifying resistance epidemiology in the field.

category: genomics


title: A novel, NADH-dependent acrylate reductase in Vibrio harveyi

abstract:
Bacteria coping with oxygen deficiency use alternative terminal electron acceptors for NADH regeneration, particularly fumarate. Fumarate is reduced by the FAD_binding_2 domain of cytoplasmic fumarate reductase in many bacteria. The variability of the primary structure of this domain in homologous proteins suggests the existence of reducing activities with different specificities. Here we produced and characterized one such protein, Vibrio harveyi ARD, and found it to be a specific NADH:acrylate oxidoreductase. This previously unknown enzyme contains covalently bound FMN and non-covalently bound FAD and FMN in a ratio of 1:1:1. The covalently bound FMN is absolutely required for activity and is attached by the specific flavin transferase, ApbE, to a threonine residue in the auxiliary FMN_bind domain. RT-qPCR and activity measurements indicated dramatic stimulation of ARD biosynthesis by acrylate in the V. harveyi cells grown aerobically. In contrast, the ard gene expression in the cells grown anaerobically was high without acrylate and increased only twofold in its presence. These findings suggest that the principal role of ARD in Vibrio is energy-saving detoxification of acrylate coming from the environment.

ImportanceThe benefits of the massive genomic information accumulated in recent years for biological sciences have been limited by the lack of data on the function of most gene products. Approximately half of the known prokaryotic genes are annotated as "proteins with unknown functions," and many other genes are annotated incorrectly. Thus, the functional and structural characterization of the products of such genes, including identification of all existing enzymatic activities, is a pressing issue in modern biochemistry. In this work, we have shown that the ard gene product of V. harveyi (GenBank ID: AIV07243) exhibits a yet undescribed NADH:acrylate oxidoreductase activity. This activity may allow acrylate detoxification and its use as a terminal electron acceptor in anaerobic or substrate in aerobic respiration of marine and other bacteria.


title: Exploring genomic data coupled with 3D chromatin structures using the WashU Epigenome Browser

abstract: Biological functions are not only encoded by the genomes sequence but also regulated by its three-dimensional (3D) structure. More and more studies have revealed the importance of 3D chromatin structures in development and diseases; therefore, visualizing the connections between genome sequence, epigenomic dynamics (1D) and the 3D genome becomes a pressing need. The WashU Epigenome Browser introduces a new 3D visualization module to integrate visualization of 1D (such as sequence features, epigenomic data) and 2D data (such as chromosome conformation capture data) with 3D genome structure. Genomic coordinates are encoded in 3D models of the chromosomes; thus, all genomic information displayed on a 1D genome browser can be visualized on a 3D model, supported by genome browser utilities and facilitating interpretation of genomic data. Biological information that is difficult to illustrate in 1D becomes more intuitive when displayed in 3D, providing novel and powerful tools for investigators to hypothesize and understand the connections between biological functions and 3D genome structures.


title: Lira: Rotational Invariant Shape and ElectrostaticDescriptors for Small Molecules and ProteinPockets based on Real Spherical Harmonics

abstract:
AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSMotivationC_ST_ABSModern AI-based tools are increasing the number of protein structures available, creating an opportunity and a challenge for automated high-throughput drug discovery pipelines. The amount of data is overwhelming for the current methods, thus demanding new high-performance approaches for Machine Learning-based rational drug design. As shape and electrostatics are the main components for understanding protein-ligand interaction; they are the primary targets for efficient AI-compatible descriptors and their associated comparison methods.

ResultsThe Lira toolbox is a set of components devised for describing, comparing and analysing shape and electrostatics for small ligands, peptides and protein pockets. It can generate databases with descriptors for tens of millions of shapes in a few hours, which can then be queried in seconds. The Lira design, focused on performance and reliability, makes its integration into AI-driven rational drug design pipelines simple.

Availability and implementationLira packages, available for download at https://pinheirolab.com/, are free to use for research and educational purposes.


title: Single-Cell Transcriptomic Analyses of Tumor Ecosystems and Spatial Architectures in Human Small Cell Lung Cancers

abstract: Small cell lung cancer (SCLC) is a highly heterogenous disease characterized by aggressive phenotypes and poor prognosis. In order to dissect the cellular components and their spatial distribution in SCLC ecosystems, here we performed single-cell RNA sequencing and spatial transcriptomics analyses of 12 resected human primary SCLCs. Our analysis revealed extensive heterogeneity of both tumor and non-tumor cells, and a predominant immunosuppressive microenvironment. Importantly, multiple hybrid tumor cell states were unmasked, including hybrid SCLC and lung adenocarcinoma cells as well as hybrid tumor and immune cells, indicating high phenotypic plasticity of tumor cells. Moreover, we revealed the cellular origin and evolution of these hybrid tumor cells, and critical roles of tumor-stromal cellular crosstalk in driving the development of hybrid cells. In conclusion, this study deepens our understanding of human SCLC ecosystems and provides rationales to target the hybrid tumor cells and immunosuppressive microenvironment for SCLC therapies.

category: cancer biology


title: Immunotherapy of glioblastoma explants induces interferon-γ responses and immune cell rearrangements in tumor center, but not periphery

abstract:
Recent therapeutic strategies for glioblastoma (GBM) aim at targeting immune tumor microenvironment (iTME) components to induce antitumoral immunity. A patient-tailored, ex vivo drug testing and response analysis platform for GBM would facilitate personalized therapy planning, provide insights into treatment-induced immune mechanisms in the iTME, and enable the discovery of biomarkers of therapy response and resistance.

We cultured 47 GBM explants from tumor center and periphery from 7 patients in perfusion bioreactors to assess iTME responses to immunotherapy. Explants were exposed to antibodies blocking the immune checkpoints CD47, PD-1 or or their combination, and were analyzed by highly multiplexed microscopy (CODEX, co-detection by indexing) using an immune-focused 55-marker panel. Culture media were examined for changes of soluble factors including cytokines, chemokines and metabolites. CODEX enabled the spatially resolved identification and quantification of >850,000 single cells in explants, which were classified into 10 cell types by clustering. Explants from center and periphery differed significantly in their cell type composition, their levels of soluble factors, and their responses to immunotherapy. In a subset of explants, culture media displayed increased interferon-{gamma} levels, which correlated with shifts in immune cell composition within specific tissue compartments, including the enrichment of CD4+ and CD8+ T cells within an adaptive immune compartment. Furthermore, significant differences in the expression levels of functional molecules in innate and adaptive immune cell types were found between explants responding or not to immunotherapy. In non-responder explants, T cells showed higher expression of PD-1, LAG-3, TIM-3 and VISTA, whereas in responders, macrophages and microglia showed higher cathepsin D levels. Our study demonstrates that ex vivo immunotherapy of GBM explants enables an active antitumoral immune response within the tumor center in a subset of patients, and provides a framework for multidimensional personalized assessment of tumor response to immunotherapy.


title: Stabilization of E-cadherin adhesions by COX-2/GSK3β signaling is a targetable pathway in metastatic breast cancer

abstract: Metastatic progression and treatment-resistance of breast cancer has been associated with epithelial-mesenchymal-transition including downregulation of E-cadherin (CDH1) expression, which can be initiated by inflammatory mediators such as COX-2. Recently, E-cadherin-mediated, cluster-based metastasis and treatment resistance has become more appreciated, though the mechanisms that maintain E-cadherin expression in this context are unknown. Through studies of inflammatory breast cancer and an in vitro tumor cell emboli culture paradigm, we identified a role for COX-2, a target gene of C/EBP{delta}, or its metabolite PGE2 in promoting protein stability of E-cadherin, {beta}-catenin and p120 catenin through inhibition of GSK3{beta}, without affecting CDH1 mRNA. The COX-2 inhibitor celecoxib downregulated E-cadherin complex proteins and caused cell death. Co-expression of E-cadherin and COX-2 was seen in breast cancer patients with poor outcome and, along with inhibitory GSK3{beta} phosphorylation, in patient-derived xenografts of triple negative breast cancer. Celecoxib alone decreased E-cadherin protein expression within xenograft tumors, reduced circulating tumor cells and clusters, and in combination with paclitaxel attenuated or regressed lung metastases. This study uncovered a mechanism by which metastatic breast cancer cells can maintain E-cadherin-mediated cell-cell adhesions and cell survival, suggesting that patients with COX-2+/E-cadherin+ breast cancer may benefit from targeting of the PGE2 signaling pathway.


title: Heterosis and Combining Ability in Pumpkin Inbreds (Cucurbita moschata Duch. ex Poir.)

abstract: Twenty hybrids along with five parents evaluated in the study were mainly contemplated to find out the best cross combinations and the best general and specific combiners as well as to estimate the nature and magnitude of the gene action for different qualitative traits. Using Griffings and Haymans approach through a 5 x 5 full diallel cross fashion, an investigation on heterosis and combining ability in pumpkin was undertaken following RCBD design with three replications at the experimental field. Both positive and negative significant GCA and SCA variances were obtained from few parents and hybrids. Predominance of additive-additive gene action was noted for most of the characters except hollowness and dry matter content, where additive-dominance gene action was predominant; flesh thickness and brix (%), where dominance-dominance gene action were predominant. A single parent was not found as good combiner for more than two characters. The best specific combiners were IBD 40 X IBD 47 for beta carotene, total sugar and fruit yield; IBD23 X IBD40 for brix (%), hollowness and flesh thickness; IBD40 X IBD57 for fruit breadth; IBD47 X IBD50 for non reducing sugar; and IBD47 X IBD57 for reducing sugar. The Vr-Wr graphs exhibited complete, partial and over dominance effect of genes for different characters. Complete dominance was observed only for beta carotene whereas over dominance was noticed for hollowness and flesh thickness. Partial dominance was ensured for fruit breadth, dry matter, brix (%), reducing sugar, non-reducing sugar, total sugar and fruit yield. Significant heterosis of some crosses against mid parent and better parents were observed for some characters.


title: Genome-wide analysis of small RNA biogenesis proteins refine the evolution of Dicer-like and Argonaute gene families in flowering plants

abstract:
Several protein families play a role in the biogenesis and function of small RNAs (sRNAs) in plants. Among these are Dicer-like (DCL), RNA-directed RNA polymerase (RDR), and Argonaute (AGO) proteins, as well as accessory proteins such as double- stranded RNA-binding (DRB), SERRATE (SE), and SUPPRESSION OF SILENCING 3 (SGS3) that are co-factors of DCL and RDR proteins. Here, we present phylogenetic analyses of seven sRNA pathway protein families performed on 198 species in the Viridiplantae. We show that the eudicot DCL3 protein is orthologous to the monocot DCL5, indicating that the monocot DCL3 has diverged and specialized in heterochromatic siRNA production. We found 167 highly conserved amino acids conserved among monocot DCL3/DCL4/DCL5 sequences. Amino acids conserved between DCL3 and DCL5 were located in the Helicase and RNase domains while amino acids conserved between DCL4 and DCL5 were mostly in the dsRNA binding and PAZ domains. Absent in DCL4, we found protein regions common to DCL3/DCL5 near both extremities of the PAZ domain which can potentially increase the distance from the RNase III catalytic center and thus determine the product size. We identified multiple duplication events of AGOs that had been lost, retained or further duplicated in sub-groups, indicating that the evolution of AGOs is complex in monocots, suggesting that functional studies in model species might not be easily translated to other species. This work provides an evolutionarily coherent and standardized annotation for gene families involved in plant sRNA biogenesis.

One sentence summaryAnalysis of small RNA biogenesis proteins from over 198 plant species refines the evolution of Dicer-like, Argonaute and other gene families, with insights into their expansion in flowering plants.


title: BOS1 is a positive regulator of wounding induced cell death and plant susceptibility to Botrytis

abstract: Programmed cell death (PCD) is required for many aspects of plant biology, including stress responses, immunity, and plant development including root and flower development. Our understanding of PCD regulation is incomplete, especially concerning regulators involved in multiple divergent processes. The botrytis-suscetible1 (bos1) mutant is one of the genotypes most susceptible to Botrytis cinerea (Botrytis) and has revealed the role of BOS1 in cell death propagation during plant responses to wounding. The bos1-1 allele harbours a T-DNA located in the 5UTR upstream from the start codon that results in elevated BOS1 transcript levels. Here, we resequenced the bos1-1 genome and found a MAS promoter at the ends of the T-DNAs. Expression of the BOS1 gene under control of the MAS promoter conferred the characteristic bos1-1 Botrytis- sensitivity and wounding phenotypes in wildtype plants. We used Crispr-Cas9 to create new bos1 alleles that disrupt exons. These lines lacked the typical bos1-1 wounding and Botrytis phenotypes, but exhibited reduced fertility, as previously observed in other bos1 T-DNA alleles. With multiple overexpression lines of BOS1, we demonstrate that BOS1 is involved in regulation of cell death propagation in a dosage dependent manner. Our data support that bos1-1 is a gain-of-function mutant and that BOS1 acts as a positive regulator of wounding and Botrytis-induced PCD. Taken together these finding suggest that BOS1 function in both fertility and Botrytis response could be unified under cell death control.


title: Coupling a Live Cell Directed Evolution Assay with Coevolutionary Landscapes to Engineer an Improved Fluorescent Rhodopsin Chloride Sensor

abstract:
Our understanding of chloride in biology can be accelerated through the application of fluorescent protein-based sensors in living cells. Laboratory-guided evolution can be used to diversify and identify sensors with specific properties. Recently, we established that the fluorescent proton-pumping rhodopsin wtGR from Gloeobacter violaceus can be converted into a fluorescent sensor for chloride. To unlock this non-natural function, a single point mutation at the Schiff counterion position from D121V was introduced into wtGR fused with cyan fluorescent protein (CFP) resulting in GR1-CFP. Here, we have integrated coevolutionary analysis with directed evolution to understand how the rhodopsin sequence space can be explored and engineered to improve this starting point. We first show how evolutionary couplings are predictive of functional sites in the rhodopsin family and how a fitness metric based on sequence can be used to quantify known proton-pumping activities of GR-CFP variants. Then, we couple this ability to predict potential functional outcomes with a screening and selection assay in live Escherichia coli to reduce the mutational search space of five residues along the proton-pumping pathway in GR1-CFP. This iterative selection process results in GR2-CFP with four additional mutations, E132K, A84K, T125C, and V245I. Finally, bulk, and single fluorescence measurements in live E. coli reveal that GR2-CFP is a reversible, ratiometric fluorescent sensor for extracellular chloride with an improved dynamic range. We anticipate that our framework will be applicable to other systems, providing a more efficient methodology to engineer fluorescent-protein based sensors with desired properties.

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title: A horizon for haptic perception

abstract: The spatial limits of sensory acquisition (its sensory horizon) is a fundamental property of any sensorimotor system. In the present study, we sought to determine the sensory horizon for the human haptic modality. At first blush, it seems obvious that the haptic system is bounded by the space where the body can interact with the environment (e.g., the arm span). However, the human somatosensory system is exquisitely tuned to sensing with tools--blind-cane navigation being a classic example of this. The horizon of haptic perception therefore extends beyond body space, but to what extent is unknown. We first used neuromechanical modelling to determine the theoretical horizon, which we pinpointed as six meters. We then used a psychophysical localization paradigm to behaviorally confirm that humans can haptically localize objects using a six-meter rod. This finding underscores the incredibly flexibility of the brains sensorimotor representations, as they can be adapted to sense with an object many times longer than the users own body.


title: Brain-wide continuous functional ultrasound imaging for real-time monitoring of hemodynamics during ischemic stroke

abstract: Ischemic stroke occurs with no warning, and therefore, very little is known about hemodynamic perturbations in the brain immediately after stroke onset. Here, functional ultrasound imaging was used to monitor variations in relative cerebral blood volume (rCBV) compared to baseline. rCBV levels were analyzed brain-wide and continuously at high spatiotemporal resolution (100m, 2Hz) until 70mins after stroke onset in rats. We compared two stroke models, with either a permanent occlusion of the middle cerebral artery (MCAo) or a tandem occlusion of both the common carotid and middle cerebral arteries (CCAo+MCAo). We observed a typical hemodynamic pattern, including a quick drop of the rCBV after MCAo, followed by spontaneous reperfusion of several brain regions located in the vicinity of the ischemic core. The severity and location of the ischemia were highly variable between animals. Still, both parameters were, on average, in good agreement with the final ischemic lesion volume measured 24hrs after stroke onset for the MCAo but not the CCAo+MCAo model. For the latter, we observed that the infarct was extended to regions that were initially not ischemic and located rostrally of the ischemic core. These regions strongly colocalize with the origin of transient hemodynamic events associated with spreading depolarizations.


title: Tissue volume estimation and age prediction using rapid structural brain scans

abstract: The multicontrast EPImix sequence generates 6 contrasts, including a T1-weighted scan, in [~]1 minute. EPImix shows comparable diagnostic performance to conventional scans under qualitative clinical evaluation, and similarities in simple quantitative measures including contrast intensity. However, EPImix scans have not yet been compared to standard MRI scans using established quantitative measures. In this study, we compared conventional and EPImix-derived T1-weighted scans of 64 healthy participants using tissue volume estimates and predicted brain-age. All scans were pre-processed using the SPM12 DARTEL pipeline, generating measures of grey matter, white matter and cerebrospinal fluid volume. Brain-age was predicted using brainageR, a Gaussian process regression model previously trained on a large sample of standard T1-weighted scans. Estimates of both global and voxel-wise tissue volume showed significantly similar results between standard and EPImix-derived T1-weighted scans. Brain-age estimates from both sequences were significantly correlated, although EPImix T1-weighted scans showed a systematic offset in predictions of chronological age. Supplementary analyses suggest that this is likely caused by the reduced field of view of EPImix scans, and the use of a brain-age model trained using conventional T1-weighted scans. However, this systematic error can be corrected using additional regression of T1-predicted brain-age onto EPImix-predicted brain-age. Finally, retest EPImix scans acquired for 10 participants demonstrated high test-retest reliability in all evaluated quantitative measurements. Quantitative analysis of EPImix scans holds potential to reduce scanning time, increasing participant comfort and reducing cost, as well as to support automation of scanning, utilising active learning for faster and individually-tailored (neuro)imaging.


title: Mosquito odorant receptor sensitive to natural spatial repellents and inhibitory compounds

abstract: Previously, we have identified an odorant receptor (OR) from the southern house mosquito Culex quinquefasciatus, CquiOR32, which responded to both odorants (agonists) and inhibitory compounds (antagonists). CquiOR32/CquiOrco-expressing oocytes responded to methyl salicylate and other odorants with inward (regular) currents but gave currents in the reverse direction when challenged with eucalyptol and other inhibitors. To determine whether hitherto unknown ORs show this intrareceptor inhibition, we have now examined two other receptors in the same cluster, CquiOR27 and CquiOR28. We cloned and tested four variants of CquiOR28, but none of the 250 compounds in our panel of odorants, including an Orco ligand candidate (OCL12), elicited inward or outward currents. By contrast, CquiOR27/CquiOrco-expressing oocytes gave robust, dose-dependent inward currents when challenged with {gamma}-octalactone and other odorants. On the other hand, octylamine and other phenolic compounds elicited dose-dependent currents in the reverse direction. When stimulatory and inhibitory compounds were presented in binary mixtures, {gamma}-octalactone-elicited inward currents were attenuated in a dose-dependent manner according to the concentration of octylamine. As part of our chemical ecology approach, we tested the repellency activity of the best ligands in the surface landing and feeding assay and a newly reported hand-in cage assay. Protection elicited by {gamma}-octalactone did not differ significantly from that of DEET at the same dose. In the hand-in cage assay, a cream formulation of {gamma}-octalactone showed 97.0 {+/-} 1.3% protection, with 47.6 {+/-} 8.3% and 1.4 {+/-} 0.7% landings per trial in the hands covered with a control and {gamma}-octalactone cream, respectively (N=8, p=0.0078, Wilcoxon matched-pairs signed-rank test).


title: Task-related changes in aperiodic activity are related to visual working memory capacity independent of event-related potentials and alpha oscillations

abstract: Individual differences in visual working memory capacity are related to slow-wave event-related potentials (ERPs) and suppression of alpha-band oscillatory power measured during the delay period of memory tasks using electroencephalography (EEG). However, recent evidence suggests that changes in the non-oscillatory, aperiodic features of the EEG signal may also contribute to working memory performance. We assessed several features of task-related changes in aperiodic activity including the spatial distribution, the effect of memory load, and the relationship between memory capacity, slow-wave ERPs, and alpha suppression. Eighty-four healthy individuals performed a continuous recall working memory task consisting of 2, 4 or 6 coloured squares while EEG was recorded. Aperiodic activity during a baseline and delay period was quantified by fitting a model to the background of the EEG power spectra using the FOOOF toolbox, which returned parameters describing the slope (exponent) and broadband offset of the spectra. The aperiodic exponent decreased (i.e., slope flattened) over lateral parieto-occipital electrodes but increased (i.e., slope steepened) over fronto-central electrodes during the delay period, whereas the offset decreased over parieto-occipital electrodes. These task-related changes in aperiodic activity were not altered by memory load. Larger increases in the aperiodic exponent were associated with higher working memory capacity measured from both the EEG task and a separate battery of complex span tasks, and this relationship was independent of slow-wave ERPs and alpha suppression. Our findings suggest that task-related changes in aperiodic activity during working memory are region specific and reflect an independent neural mechanism important for general working memory ability.


title: Inhibition of the IL-17A axis Protects against Immune-related Adverse Events while Supporting Checkpoint Inhibitor Anti-tumor Efficacy

abstract: Checkpoint inhibitor (ICI) immunotherapy leverages the bodys own immune system to attack cancer cells but leads to unwanted autoimmune side effects in up to 60% of patients. Such immune related adverse events (IrAE) may lead to treatment interruption, permanent organ dysfunction, hospitalization and premature death. Thyroiditis is one of the most common IrAE, but the cause of thyroid IrAE remains unknown. Here we present a novel mouse model in which checkpoint inhibitor therapy leads to multi-organ autoimmune infiltrates and show that activation and infiltration of Type 3 immune cells including IL17A+ ROR{gamma}t+ CD4+ (T helper 17 or Th17) and gamma delta 17 ({gamma}{delta}T17) T cells promote thyroid IrAE development. IL-17A+ T cells were similarly found in thyroid specimens from cancer patients treated with ICI who developed thyroid IrAE. Furthermore, antibody-based inhibition of IL-17A, a clinically available therapy, significantly reduced thyroid IrAE development in ICI-treated mice. Finally, combination of IL-17A neutralization with ICI treatment in multiple tumor models did not reduce ICI anti-tumor efficacy. These studies suggest that targeting Th17 and {gamma}{delta}17 function may reduce IrAE without impairing ICI anti-tumor efficacy and may be a generalizable strategy to address IL17-mediated IrAE.


title: Therapeutic Interruption of T Cell Development Generates High-Affinity T Cells That Escape Exhaustion and Improve Cancer Immunotherapy

abstract: Availability of effective anti-tumor T cells is limited by cancer immunoediting, which depletes neoantigens, and central tolerance, which eliminates developing T cells with high-affinity T cell receptors (TCRs) against tumor self-antigens. Remaining tumor-reactive T cells are often exhausted after immune checkpoint blockade (ICB). Whether endogenous T cells with high- affinity TCRs against tumor self-antigens can be generated to circumvent exhaustion and reject neoantigen-poor tumors is unclear. We show that transiently interrupting central tolerance through RANKL blockade unleashes T cells possessing TCRs with self-reactive features that enable ICB to reject poorly immunogenic tumors. Upon recognition of tumor self-antigens, these T cells exhibit enhanced TCR signaling, enrichment in NFAT/AP-1 genes, and lymph node priming. Consequently, memory-precursor T cells against tumor self-antigens are generated, avoid exhaustion, and become effector-memory cells with transcriptional features associated with clinical ICB response. Thus, interrupting central tolerance provides T cells with tumor- directed autoreactivity that avoid exhaustion and improve immunotherapy.


title: A GSVA based gene set synergizing with CD4+T cell bearing harmful factors yield risk signals in HBV related diseases via amalgamation of artificial intelligence

abstract: Genes encoding chemokines and extracellular matrix (ECM) play pivotal roles in chronic HBV infection (CHB), HBV related fibrosis (HBV-LF) and hepatocellular carcinoma (HBV-HCC). The landscape and potential of these genes in prognosis across diseases stages have not been fully and systemically understood. In this study, we defined an HBV-LF associated gene set comprised of chemokines and ECM related genes directly induced by initial HBV infection through GSVA algorithm that named as CLST (C stands for CXCL9, CXCL10, CCL19 and CCL20; L for LUM; S for SOX9 and SPP1; T for THBS1, THBS2) and evaluated its biomarker values in CHB and HBV-LF. Enrichment scores (ES) of CLST was subsequently observed synergized with activated CD4+T cells (aCD4) highly related to T helper cell 17 (TH17) associated genes and immune checkpoints and addressed as risk signals due to bearing harmful prognosis factors in tumor tissues of patients with HBV-HCC. Dual higher enrichment score (ES) of CLST and aCD4 in HBV-HCC patients exhibited worse overall survival (OS). Feature genes specific to these two gene sets showed promising clinical relevance in early-stage of HBV-HCC definition and OS prediction incorporating laboratory parameters via artificial intelligence (AI) systems. Finally, a novel mechanistic insight into the issue was proposed that PEG IFN- as an immunotherapy through modulating CLST signal in treatment responders and these immune signals down-regulation could be beneficial for HBV related diseases control and prevention. Together, our study provides GSVA and AI derived immunogenomic prognosis signatures and clinical utility of these signals will be benefit for HBV related diseases cure.


title: An early decline in ETPs reflects reduced pre-thymic progenitors and altered signals from the thymus microenvironment

abstract:
Age-related thymus involution results in decreased T-cell production, contributing to increased susceptibility to pathogens. Elucidating mechanisms underlying involution will inform strategies to restore thymopoiesis. The thymus is colonized by circulating bone marrow (BM)-derived thymus seeding progenitors (TSPs) that differentiate into early T-cell progenitors (ETPs). We find ETP cellularity declines as early as 3 months (3MO) in mice. This initial ETP reduction could reflect changes in pre-thymic progenitors and/or thymic stromal niches. We demonstrate that the number of functional TSP/ETP niches is not reduced with age. Instead, the number of pre-thymic BM and circulating lymphoid progenitors is substantially reduced by 3MO, although their intrinsic ability to seed and differentiate in the thymus is maintained. Additionally, Notch signaling in BM progenitors and ETPs declines by 3MO, indicating defective niche quality contributes to the reduction in ETPs. Together, these findings indicate that diminished BM lymphopoiesis and thymic stromal support contribute to the initial decline in ETPs, setting the stage for progressive thymus involution.

Summary statementThe number of early T-cell progenitors declines by 3 months of age in mice. This decline reflects a sharp drop in circulating thymus seeding progenitors, fewer bone marrow lymphoid progenitors, and reduced Notch signaling in both bone marrow and thymus.


title: Urban population structure and local movement ecology of a native Australian mosquito involved in disease transmission

abstract: Successful species management is reliant upon understanding key ecological features of the target species and the environment. Dispersal is a critical factor to assess when designing control measures as it determines the rate of migration out of or into controlled areas and influences the risk of human exposure to the species and its pathogens. This study uses spatial population genomics to investigate the movement ecology of Aedes notoscriptus, an important disease transmitting mosquito, at the Mornington Peninsula, Australia. We sampled Ae. notoscriptus eggs from the Mornington Peninsula at a single time point, and generated genomic data from 240 individuals from different locations. We also produced a draft genome assembly for this clade of Ae. notoscriptus. We used genomic data to detect close kin dyads, and the locations of dyads showed that specific acts of movement in the previous generation had occurred over distances >1 km. We also investigated isolation by distance patterns from the spatial autocorrelation of genetic distances. Significant genetic dissimilarity in Ae. notoscriptus began to be observed at >4 km separation, a fourfold higher distance than in a comparable sample of the dengue mosquito Ae. aegypti. These findings are evidence that the high mobility of Ae. notoscriptus influenced the success of a 2021 mosquito control trial at the Mornington Peninsula, because the dispersal ability of Ae. notoscriptus was likely to have exceeded the size of the intervention zones which were designed around ecological knowledge of Ae. aegypti. Further sampling within the same area was used to obtain counts of Ae. notoscriptus eggs at two timepoints 6 and 12 months after initial sampling. We found egg counts to be consistent across timepoints, and spatial variation in egg counts was found to covary with spatial variation in neighbourhood size (NS). As NS increases linearly with population density, this is evidence that egg counts may be useful for estimating relative density in Ae. notoscriptus. The overall results of this study draw attention to the importance of acquiring species-specific data when planning control measures, and contribute to the fundamental ecological understanding of this important vector species.


title: Spatially Aware Dimension Reduction for Spatial Transcriptomics

abstract: Spatial transcriptomics are a collection of genomic technologies that have enabled transcriptomic profiling on tissues with spatial localization information. Analyzing spatial transcriptomic data is computationally challenging, as the data collected from various spatial transcriptomic technologies are often noisy and display substantial spatial correlation across tissue locations. Here, we develop a spatially-aware dimension reduction method, SpatialPCA, that can extract a low dimensional representation of the spatial transcriptomics data with enriched biological signal and preserved spatial correlation structure, thus unlocking many existing computational tools previously developed in single-cell RNAseq studies for tailored and novel analysis of spatial transcriptomics. We illustrate the benefits of SpatialPCA for spatial domain detection and explores its utility for trajectory inference on the tissue and for high-resolution spatial map construction. In the real data applications, SpatialPCA identifies key molecular and immunological signatures in a newly detected tumor surrounding microenvironment, including a tertiary lymphoid structure that shapes the gradual transcriptomic transition during tumorigenesis and metastasis. In addition, SpatialPCA detects the past neuronal developmental history that underlies the current transcriptomic landscape across tissue locations in the cortex.

category: genetics


title: A natural fungal gene drive enacts killing through targeting DNA

abstract:
Fungal spore-killers are a class of selfish genetic elements that positively bias their own inheritance by killing non-inheriting gametes following meiosis. As killing takes place specifically within the developing fungal ascus, a tissue which is experimentally difficult to isolate, our understanding of the mechanisms underlying spore killers are limited. In particular, how these loci kill other spores within the fungal ascus is largely unknown. Here we overcome these experimental barriers by developing model systems in two evolutionary distant organisms, Escherichia coli (bacterium) and Saccharomyces cerevisiae (yeast). Using these systems, we show that the Podospora anserina spore killer protein Spok1 enacts killing through targeting DNA.

Significance StatementNatural gene drives have shaped the genomes of many eukaryotes and recently have been considered for applications to control undesirable species. In fungi these loci are called spore-killers. Despite their importance in evolutionary processes and possible applications our understanding of how they enact killing is limited. We show that the spore killer protein Spok1, which has homologues throughout the fungal tree of life, acts via DNA disruption. Spok1 is only the second spore killer locus in which the cellular target of killing has been identified and is the first known to target DNA. We also show that the DNA disrupting activity of Spok1 is functional in both bacteria and yeast suggesting a highly conserved mode of action.


title: Experimental Design and Power Calculation in Omics Circadian Rhythmicity Detection

abstract: Circadian clocks are 24-hour endogenous oscillators in physiological and behavioral processes. Though recent transcriptomic studies have been successful in revealing the circadian rhythmicity in gene expression, the power calculation and study design for omics circadian analysis have not been explored. In this paper, we develop a statistical package, namely CircaPower, to perform power calculation for circadian pattern detection. Our theoretical framework is determined by three key factors in circadian gene detection: sample size, intrinsic effect size and sampling design. Via simulations, we systematically investigate the impact of these key factors on circadian power calculation. We demonstrate that CircaPower not only has fast and accurate computing but also is robust against variety of violations of model assumptions. In real applications, we demonstrate the performance of CircaPower using mouse pan-tissue data and human postmortem brain data, and illustrate how to perform circadian power calculation using mouse skeletal muscle microarray pilot data as a case study. Our method CircaPower has been implemented in an R package, which is made publicly available on GitHub (https://github.com/circaPower/circaPower).


title: Rice Pan-genome Array (RPGA): an efficient genotyping solution for pan-genome-based accelerated crop improvement in rice

abstract: The advent of the pan-genome era has unraveled previously unknown genetic variation existing within diverse crop plants including rice. This untapped genetic variation is believed to account for a major portion of phenotypic variation existing in crop plants and might be responsible for missing heritability. However, the use of conventional single reference-guided genotyping often fails to capture large portion of this genetic variation leading to a reference bias. This makes it difficult to identify and utilize novel population/cultivar-specific genes for crop improvement. To overcome this challenge, we developed a rice pan-genome genotyping array (RPGA) includes 80K genome-wide SNPs which provides simple, user-friendly and cost-effective solution for rapid pan-genome-based genotyping in rice. The GWAS conducted using RPGA-SNP genotyping data of a rice diversity panel detected total of 42 loci, including previously known as well as novel genomic loci regulating grain size/weight traits in rice. Eight of the identified trait-associated loci (dispensable loci) could not be detected with conventional single reference genome-based GWAS and found to be missing from the commonly used Nipponbare reference genome. WD repeat-containing PROTEIN 12 gene underlying one of such dispensable locus on chromosome 7 (qLWR7) along with few other non-dispensable loci was subsequently detected using high-resolution QTL mapping confirming authenticity of RPGA-led GWAS. This demonstrates the potential of RPGA-based genotyping to overcome reference bias. Besides GWAS, the application of RPGA-based genotyping for natural allelic diversity and population structure analysis, seed purity and hybridity testing, ultra-high-density genetic map construction and chromosome level genome assembly, and marker-assisted foreground/background selection was successfully demonstrated. Based on these salient outcomes, a web application (http://www.rpgaweb.com) was also developed to provide easy to use platform for imputation of RPGA-based genotyping data using 3K Rice Reference Panel and subsequent GWAS in order to drive genetic improvement of rice.


title: Improving the Identification Coverage of Protein Interactome by Enhancing the Click Chemistry-based Cross-linking Enrichment Efficiency

abstract:
Chemical cross-linking coupled with mass spectrometry has emerged as a powerful strategy which enables global profiling of protein interactome with direct interaction interfaces in complex biological systems. The alkyne-tagged enrichable cross-linkers are preferred to improve the coverage of low-abundance cross-linked peptides, combined with click chemistry for biotin conjugation to allow the cross-linked peptides enrichment. However, a systematic evaluation on the efficiency of click approaches (protein-based or peptide-based) and diverse cleavable click chemistry ligands (acid, reduction, photo) for cross-linked peptides enrichment and release is lacking. Herein, together with in vivo chemical cross-linking by alkyne-tagged cross-linker, we explored the click chemistry-based enrichment approaches on protein and peptide level with three cleavable click chemistry ligands, respectively. By comparison, the approach of protein-based click chemistry conjugation with acid-cleavable tag was demonstrated to permit the most cross-linked peptides identification. The advancement of this strategy enhanced the proteome-wide cross-linking analysis, permitting a detection of 5,017 protein-protein interactions among 1,909 proteins across all subcellular compartments with wide abundance distribution in cell. Therefore, all these results demonstrated a guideline value of our work for efficient cross-linked peptides enrichment, thus facilitated the in-depth profiling of protein interactome for functional analysis.

GRAPHICAL ABSTRACT

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title: Longitudinal Morphological and Functional Characterization of Human Heart Organoids Using Optical Coherence Tomography

abstract: Organoids play an increasingly important role as in vitro models for studying organ development, disease mechanisms, and drug discovery. Organoids are self-organizing, organ-like three-dimensional (3D) cell cultures developing organ-specific cell types and functions. Recently, three groups independently developed self-assembling human heart organoids (hHOs) from human pluripotent stem cells (hPSCs). In this study, we utilized a customized spectral-domain optical coherence tomography (SD-OCT) system to characterize the growth of hHOs. Development of chamber structures and beating patterns of the hHOs were observed via OCT and calcium imaging. We demonstrated the capability of OCT to produce 3D images in a fast, label-free, and non-destructive manner. The hHOs formed cavities of various sizes, and complex interconnections were observed as early as on day 4 of differentiation. The hHOs models and the OCT imaging system showed promising insights as an in vitro platform for investigating heart development and disease mechanisms.


title: Predicting Mechanical Ventilation Effects on Six Human Tissue Transcriptomes

abstract:
BackgroundMechanical ventilation (MV) is a lifesaving therapy used for patients with respiratory failure. Nevertheless, MV is associated with numerous complications and increased mortality. The aim of this study is to define the effects of MV on gene expression of direct and peripheral human tissues.

MethodsClassification models were applied to Genotype-Tissue Expression Project (GTEx) gene expression data of six representative tissues- liver, adipose, skin, nerve-tibial, muscle and lung, for performance comparison and feature analysis. We utilized 18 prediction models using the Random Forest (RF), XGBoost (eXtreme Gradient Boosting) decision tree and ANN (Artificial Neural Network) methods to classify ventilation and non-ventilation samples and to compare their prediction performance for the six tissues. In the model comparison, the AUC (area under receiver operating curve), accuracy, precision, recall, and F1 score were used to evaluate the predictive performance of each model. We then conducted feature analysis per each tissue to detect MV marker genes followed by pathway enrichment analysis for these genes.

ResultsXGBoost outperformed the other methods and predicted samples had undergone MV with an average accuracy for the six tissues of 0.951 and average AUC of 0.945. The feature analysis detected a combination of MV marker genes per each tested tissue, some common across several tissues. MV marker genes were mainly related to inflammation and fibrosis as well as cell development and movement regulation. The MV marker genes were significantly enriched in inflammatory and viral pathways.

ConclusionThe XGBoost method demonstrated clear enhanced performance and feature analysis compared to the other models. XGBoost was helpful in detecting the tissue-specific marker genes for identifying transcriptomic changes related to MV. Our results show that MV is associated with reduced development and movement in the tissues and higher inflammation and injury not only in direct tissues such as the lungs but also in peripheral tissues and thus should be carefully considered before being implemented.


title: Detecting Antibody Reactivities in Phage ImmunoPrecipitation Sequencing Data

abstract: Phage ImmunoPrecipitation Sequencing (PhIP-Seq) is a recently developed technology to assess antibody reactivity, quantifying antibody binding towards hundreds of thousands of candidate epitopes. The output from PhIP-Seq experiments are read count matrices, similar to RNA-Seq data; however some important differences do exist. In this manuscript we investigated whether the publicly available method edgeR1 for normalization and analysis of RNA-Seq data is also suitable for PhIP-Seq data. We find that edgeR is remarkably effective, but improvements can be made and introduce a Bayesian framework specifically tailored for data from PhIP-Seq experiments (Bayesian Enrichment Estimation in R, BEER).


title: Splicing factor BUD31 promotes ovarian cancer progression through sustaining the expression of anti-apoptotic BCL2L12

abstract: Dysregulated expression of splicing factors has important roles in cancer development and progression. However, it remains a challenge to identify the cancer-specific splicing variants. Here we demonstrated that spliceosome component BUD31 is increased in ovarian cancer, and its higher expression predicts worse prognosis. We characterized the BUD31 binding motif and found that BUD31 preferentially binds exon-intron regions near splicing sites by CLIP-seq. Further analysis revealed that BUD31 inhibition results in extensive exon skipping and decreased abundance of long CDS isoforms. In particular, we identified BCL2L12, an anti-apoptotic BCL2 family member, as a functional splicing target of BUD31. BUD31 stimulates the inclusion of exon 3 to generate full-length BCL2L12 and promotes ovarian cancer progression. Knockdown of BUD31 or splice-switching antisense oligonucleotide treatment promotes exon 3 skipping and results in a truncated isoform of BCL2L12 that undergoes nonsense-mediated mRNA decay, and the cells subsequently undergo apoptosis. Our findings reveal BUD31-regulated exon inclusion as a critical factor in ovarian cancer cell survival and progression.


title: mTOR is a Major Determinant of Chemosensitivity

abstract: Chemotherapy can eradicate a majority of cancer cells. However, a small population of tumor cells often survives drug treatments through genetic and/or non-genetic mechanisms, leading to tumor recurrence. Here we report a reversible chemoresistance phenotype regulated by the mTOR pathway. Through a genome-wide CRISPR knockout library screen in pancreatic cancer cells treated with chemotherapeutic agents, we have identified mTOR pathway as a prominent determinant of chemosensitivity. Pharmacological suppression of mTOR activity in cancer cells from diverse tissue origins leads to the persistence of a reversibly resistant population, which is otherwise eliminated by chemotherapeutic agents. Conversely, activation of mTOR pathway increases chemosensitivity in vitro and in vivo and predicts better survival among various human cancers. Persister cells display a senescence phenotype. Inhibition of mTOR does not induce cellular senescence per se, but rather promotes the survival of senescent cells through regulation of autophagy and G2/M cell cycle arrest, as revealed by a small molecule chemical library screen. Thus, mTOR plays a causal yet paradoxical role in regulating chemosensitivity; inhibition of the mTOR pathway, while suppressing tumor expansion, facilitates the development of a reversible drug-tolerant senescence state.


title: Adipose tissue dysfunction and visceral fat are associated to hepatic insulin resistance and severity of NASH even in lean individuals

abstract:
Background & AimsNonalcoholic fatty liver disease (NAFLD) is a heterogeneous disorder, but the factors that determine this heterogeneity remain poorly understood. Adipose tissue (AT) dysfunction is causally linked to NAFLD since it causes intrahepatic triglyceride (IHTG) accumulation through increased hepatic lipid flow, due to insulin resistance (IR) and pro-inflammatory adipokines release. While many studies in NAFLD have looked at total adiposity (that is mainly subcutaneous fat, SC-AT), it is still unclear the impact of visceral fat (VF). Thus, we investigated how VF vs. SC-AT were related to NAFLD in lean, overweight, and obese individuals compared to lean controls.

MethodsThirty-four non-diabetic NAFLD with liver biopsy and eight lean control individuals (CT) were enrolled in this study. We measured fat distribution (VF, SC-AT and IHTG) by magnetic resonance imaging (MRI), adiponectin concentration, free fatty acids (FFAs) and triglyceride (TAG) concentration and composition by mass spectrometry (MS), lipolysis and IR by tracer infusion.

ResultsIHTG was positively associated with lipolysis, adipose tissue IR, TG concentrations, and increased ratio of saturated/unsaturated fatty acids. VF was higher in NAFLD (including lean individuals) compared to controls, was increased with fibrosis stage and was associated with IR in liver, muscle and adipose tissue, increased lipolysis, and decreased adiponectin levels. Collectively, our results suggest that VF accumulation, given its location close to the liver, is one of the major risk factors for NAFLD.

ConclusionsThese findings propose VF as an early indicator of NAFLD independently of BMI, which may allow for evidence-based prevention and intervention strategies.

category: physiology


title: Ultra-flexible and Stretchable Intrafascicular Peripheral Nerve Recording Device with Axon-dimension, Cuff-less Microneedle Electrode Array

abstract: Peripheral nerve mapping tools with higher spatial resolution are needed to advance systems neuroscience, and potentially provide a closed-loop biomarker in neuromodulation applications. Two critical challenges of microscale neural interfaces are (i) how to apply them to small peripheral nerves, and (ii) how to minimize chronic reactivity. We developed a flexible microneedle nerve array (MINA), which is the first high-density penetrating electrode array made with axon-sized silicon microneedles embedded in low-modulus thin silicone. We present the design, fabrication, acute recording, and chronic reactivity to an implanted MINA. Distinctive units were identified in the rat peroneal nerve. We also demonstrate a long-term, cuff-free, and suture-free fixation manner using rose bengal as a light-activated adhesive for two timepoints. The tissue response at 1-week included a sham (N=5) and MINA-implanted (N=5) group, and the response at 6-week also included a sham (N=3) and MINA-implanted (N=4) group. These conditions were quantified in the left vagus nerve of rats using histomorphometry. Micro-CT was added to visualize and quantify tissue encapsulation around the implant. MINA demonstrated a reduction in encapsulation thickness over previously quantified interfascicular methods. Future challenges include techniques for precise insertion of the microneedle electrodes and demonstrating long-term recording.


title: Implicit adaptation is modulated by the relevance of feedback

abstract: The motor system adapts to changes in the body and the environment by learning from sensory prediction errors, the difference between predicted and actual visual feedback. This process is highly automatic and implicit, with the response to the visual feedback occurring even when the resultant change from the sensory prediction error worsens task performance. Given that informative and relevant feedback in the real world is often intertwined with distracting and irrelevant feedback, we asked how the relevancy of visual feedback impacts implicit adaptation. To tackle this question, we presented multiple cursors as visual feedback in a center-out reaching task and varied the task relevance of these cursors. In other words, participants were instructed to hit a target with a specific task-relevant cursor, while ignoring the other cursors. In Experiment 1, we found that reach aftereffects were attenuated by the mere presence of distracting cursors, compared to reach aftereffects in response to a single task-relevant cursor. The degree of attenuation did not depend on the position of the distracting cursors. In Experiment 2, we examined the interaction between task relevance and attention. Participants were asked to adapt to a task-relevant cursor/target pair, while ignoring the task-irrelevant cursor/target pair. Critically, we jittered the location of the relevant and irrelevant target in an uncorrelated manner, allowing us to index attention via how well participants tracked the position of target. As expected, participants who were better at tracking the task-relevant target/cursor pair showed greater aftereffects, and interestingly, the same correlation applied to the task-irrelevant target/cursor pair. Together, these results highlight the novel role of task relevancy on modulating implicit adaptation, perhaps by giving greater attention to informative sources of feedback, increasing the saliency of the sensory prediction error.


title: Tracking the dynamics of perisaccadic visual signals with magnetoencephalography

abstract: Many brain functions are difficult to localize, as they involve distributed networks that reconfigure themselves on short timescales. One example is the integration of oculomotor and visual signals that occurs with each eye movement: The brain must combine motor signals about the eye displacement with retinal signals, to infer the structure of the surrounding environment. Our understanding of this process comes primarily from single-neuron recordings, which are limited in spatial extent, or fMRI measurements, which have poor temporal resolution. We have therefore studied visual processing during eye movements, using magnetoencephalography (MEG), which affords high spatiotemporal resolution. Human subjects performed a task in which they reported the orientation of a visual stimulus while executing a saccade. After removal of eye movement artifacts, time-frequency analysis revealed a signal that propagated in the beta-frequency band from parietal cortex to visual cortex. This signal had the characteristics of perisaccadic "remapping", a neural signature of the integration of oculomotor and visual signals. These results reveal a novel mechanism of visual perception and demonstrate that MEG can provide a useful window into distributed brain functions.


title: Behavioral and neurochemical effects of novel N-Benzyl-2-phenylethylamine derivatives in adult zebrafish

abstract: Serotonergic hallucinogenic drugs potently affect human brain and behavior, and have recently emerged as potentially promising agents in psychopharmacotherapy. Complementing rodent studies, zebrafish (Danio rerio) is a powerful animal model for screening neuroactive drugs, including serotonergic agents. Here, we test ten different N-Benzyl-2-phenylethylamine (NBPEA) derivatives with the 2,4- and 3,4-dimethoxy substitutions in the phenethylamine moiety and the - OCH3, -OCF3, -F, -Cl and -Br substitutions in the ortho position of phenyl ring of N-benzyl fragment, assessing their behavioral and neurochemical effects in adult zebrafish. Overall, substitutions in N-benzyl fragment primarily affected zebrafish locomotion, and in phenethylamine moiety - anxiety-like behavior, also modulating brain serotonin and/or dopamine turnover. We also identified several behavioral clusters, including anxiogenic/hypolocomotor (24H-NBF, 24H-NBOMe and 34H-NBF), behaviorally inert (34H-NBBr, 34H-NBCl and 34H- NBOMe), anxiogenic/hallucinogenic-like (24H-NBBr, 24H-NBCl and 24H-NBOMe(F)), and anxiolytic/hallucinogenic-like (34H-NBOMe(F)) agents. The 24H-NBOMe(F) and 34H-NBOMe(F) also reduced despair-like behavior in zebrafish. The artificial intelligence-driven phenotyping supports association of multiple compounds with NMDA antagonists and/or MDMA, supporting their potential hallucinogenic-like properties, as well as other valuable psychoactive effects. In silico functional molecular activity modelling also supports existing of similarities between studied NBPEAs drugs, MDMA, and ketamine. Functional analysis implicates potential involvement of serotonin release stimulating activity, calcium channel (voltage-sensitive) activity, some serotonin receptors activity and variety of psychiatric and neurologic disorders treatments activities. Overall, we report potent neuroactive properties of several novel synthetic N-benzylphenylethylamines in an in vivo vertebrate model system (zebrafish), raising the possibility of their potential use in clinical practice.


title: Reduced feedback barely slows down proprioceptive recalibration

abstract: Introducing altered visual feedback of the hand results in quick adaptation of reaching movements. And while this may be partly due to explicit strategies, our lab has shown that implicit changes like reach aftereffects and shift in estimates of the unseen hand, can also emerge and even saturate within a few training trials. The goal of the current study is to determine whether these rapid changes in unseen hand position that occur during classical visuomotor adaptation are diminished or slowed when feedback during training is reduced. We reduced feedback by either providing visual feedback only at the end of the reach (terminal feedback) or constraining hand movements to reduce efferent contribution (exposure). We measured changes as participants completed reaches with a 30{degrees} rotation, a -30{degrees} rotation and clamped visual feedback, with these two "impoverished" training conditions, along with classical visuomotor adaptation training, while continuously estimating their felt hand position. Classic continuous-cursor training produced exemplary learning curves and rapid and robust shifts in felt hand position. Training with terminal feedback slightly reduced the initial rate of change in overall adaptation and but not the magnitude of shifts in felt hand position. Finally using a robot to constrain and deviate hand movement direction, called exposure training, only delayed saturation of proprioceptive changes by a single trial and these changes were slightly smaller than those during classical training. Taken together, adaptation and shifts in felt hand position are a rapid and robust responses to sensory mismatches and are only slightly modulated when feedback is reduced. This means that, given a visuo-proprioceptive mismatch, the resulting shift in sense of limb position can contribute to movements from the start of adaptation.


title: Tree size drives diversity and community structure of microbial communities on the bark of beech (Fagus sylvatica)

abstract: Tree bark constitutes ideal habitat for microbial communities, because it is a stable substrate, rich in micro-niches. Bacteria, fungi, and terrestrial microalgae together form microbial communities, which in turn support more bark-associated organisms, such as mosses, lichens, and invertebrates, thus contributing to forest biodiversity. We have a limited understanding of the diversity and biotic interactions of the bark-associated microbiome, as investigations have mainly focussed on agriculturally relevant systems and on single taxonomic groups. Here we implemented a multi-kingdom metabarcoding approach to analyse diversity and community structure of the green algal, bacterial, and fungal components of the bark-associated microbial communities of beech, the most common broadleaved tree of Central European forests. We identified the most abundant taxa, hub taxa, and co-occurring taxa. We found that tree size (as a proxy for age) is an important driver of community assembly, suggesting that environmental filtering leads to less diverse fungal and algal communities over time. Conversely, forest management intensity had negligible effects on microbial communities on bark. Our study suggests the presence of undescribed, yet ecologically meaningful taxa, especially in the fungi, and highlights the importance of bark surfaces as a reservoir of microbial diversity. Our results constitute a first, essential step towards an integrated framework for understanding microbial community assembly processes on bark surfaces, an understudied habitat and neglected component of terrestrial biodiversity. Finally, we propose a cost-effective sampling strategy to study bark-associated microbial communities across large spatial or environmental scales.


title: Mapping cherry blossoms from geotagged street-level photos

abstract: Floral phenology is useful information as an indicator on climate change and ecosystem services, however its observation is not straightforward over space and time. Satellite remote sensing and official and volunteer-based in-situ observations have been conducting, but the long-term and accurate data collection is challenging due to the insufficient quality and quantity of observations and the lack of financial and human resources to sustain. Here, we demonstrate a flower detection model from street-level photos, which can be the core function of a semi-automatic observation system to tackle those issues above. We detected cherry blossoms by this model from geotagged images with the observation date, obtained from Mapillary, which is one of social sensing data sources, and mapped dates of flowering in a study site, Aizuwakamatsu, Japan in April 2018. This approach enables us to collect floral phenology information semi-automatically as a data-driven approach. It is expected to collect a large number of observations with a certain level of quality by avoiding human-induced biases for the observations.


title: Quantifying the effects of recent glacial history and future climate change on a unique population of mountain goats

abstract: Human disturbance and climate change can impact populations by disrupting movement corridors and reducing important habitat. Characterizing how animals respond to such environmental changes is valuable for conservation as many species, especially habitat specialists, can experience reduced genetic diversity when deleterious habitat change occurs, leading to an increased likelihood of extirpation. Mountain goats (Oreamnos americanus) exemplify this conservation challenge; their geographically isolated habitat can inhibit gene flow, making them susceptible to population declines in the face of anthropogenic-induced landscape change. To facilitate biologically informed population management of mountain goats in Glacier Bay National Park, Alaska, we characterized the fine-scale genetic population structure and examined how future climate change could impact the population density of these mountain goats. We used DNA samples to estimate diversity and depict the genealogical history. Climate response models allowed us to simulate changes to suitable habitat and predict how this might influence future population structure. Our results indicated that three genetically distinct subpopulations exist in Glacier Bay and that the population structure is reflective of the historic landscape patterns. Climate modeling predicted that demographic productivity was likely to be reduced for all subpopulations; additionally, we found that climate change likely degrades the suitability of movement corridors that facilitate gene flow between subpopulations, ultimately increasing the cost of travel. Understanding such fine-scale patterns are key to managing subpopulations, particularly with impending changes to the landscape.


title: Bergmann's rule in Microlophus lizards: testing for latitudinal and climatic gradients of body size.

abstract: Broadscale environmental gradients exert selection pressures that can result in macroevolutionary patterns of phenotypic diversity. Bergmanns rule, one of the most studied ecogeographical patterns, states that among related species or populations, larger body sizes should occur at higher latitudes or colder climates given the efficacy of larger sizes to conserve heat. However, there is still much debate regarding the mechanisms that underlie the rule and whether they should be different between endotherms and ectotherms. Here we test for Bergmanns rule in Microlophus lizards, a genus distributed in western South America and the Galapagos Islands, and test for associations between body size and climatic variables. Our results indicate that mainland Microlophus species follow Bergmanns rule, meaning that larger species are found towards the southernmost latitudes encompassed by the genus distribution. This relationship was similar for males and females, suggesting that patterns of sexual size dimorphism are not driven by differing latitudinal gradients of body size between the sexes. We identified negative associations between body size and temperature, precipitation, and isothermality, meaning that larger species are found in colder, drier, and more thermally variable environments. We discuss the possible mechanisms behind this pattern, the implications of insularity and ecology for the evolution of body size in this genus, and emphasize the need for additional natural history information to allow for the elucidation of the mechanisms behind the patterns presented in this work.