title: Infection by a eukaryotic gut parasite in wild Daphnia sp. associates with a distinct bacterial community

abstract: Host-associated bacterial communities can play an important role in host fitness and resistance to diseases. Yet, few studies have investigated tripartite interaction between a host, parasite and host-associated bacterial communities in natural settings. Here, we use 16S amplicon sequencing to compare gut- and body-bacterial communities of wild water fleas belonging to the Daphnia longispina complex, between uninfected hosts and those infected with the common and virulent eukaryotic gut parasite Caullerya mesnili (Family: Ichthyosporea). We report community-level changes in host-associated bacteria with the presence of the parasite infection; namely decreased alpha diversity and increased beta diversity at the site of infection, i.e. host gut (but not host body). We also report decreased abundance of bacterial taxa proposed elsewhere to be beneficial for the host, and an appearance of taxa specifically associated with infected hosts. Our study highlights the host-microbiota-infection link in a natural system and raises questions about the role of host-associated microbiota in natural disease epidemics as well as the functional roles of bacteria specifically associated with infected hosts.


title: ProbeTools: Designing hybridization probes for targeted genomic sequencing of diverse and hypervariable viral taxa

abstract:
BackgroundSequencing viruses in many specimens is hindered by excessive background material from hosts, microbiota, and environmental organisms. Consequently, enrichment of target genomic material is necessary for practical high-throughput viral genome sequencing. Hybridization probes are widely used for enrichment in many fields, but their application to viral sequencing faces a major obstacle: it is difficult to design panels of probe oligo sequences that broadly target many viral taxa due to their rapid evolution, extensive diversity, and genetic hypervariability. To address this challenge, we created ProbeTools, a package of bioinformatic tools for generating effective viral capture panels, and for assessing coverage of target sequences by probe panel designs in silico. In this study, we validated ProbeTools by designing a panel of 3,600 probes for subtyping the hypervariable haemagglutinin (HA) and neuraminidase (NA) genome segments of avian-origin influenza A viruses (AIVs). Using in silico assessment of AIV reference sequences and in vitro capture on egg-cultured viral isolates, we demonstrated effective performance by our custom AIV panel and ProbeTools suitability for challenging viral probe design applications.

ResultsBased on ProbeTools in silico analysis, our panel provided broadly inclusive coverage of 14,772 HA and 11,967 NA reference sequences. 90% of these HA and NA references sequences had 90.8% and 95.1% of their nucleotide positions covered in silico by the panel respectively. We also observed effective in vitro capture on a representative collection of 23 egg-cultured AIVs that included isolates from wild birds, poultry, and humans and representatives from all HA and NA subtypes. 42 of 46 HA and NA segments had over 98.3% of their nucleotide positions significantly enriched by our custom panel. These in vitro results were further used to validate ProbeTools in silico coverage assessment algorithm; 89.2% of in silico predictions were concordant with in vitro results.

ConclusionsProbeTools generated an effective panel for subtyping AIVs that can be deployed for genomic surveillance, outbreak prevention, and pandemic preparedness. Effective probe design against hypervariable AIV targets also validated ProbeTools design and coverage assessment algorithms, demonstrating their suitability for other challenging viral capture applications.

category: genomics


title: Distinct chromosomal niches in the genome of S. cerevisiae provide the background for genomic innovation and shape the fate of gene duplicates

abstract:
Gene duplication is a major source of genomic innovation in all eukaryotes, with large proportions of genes being the result of either small-scale (SSD) or genome-wide duplication (WGD) events. In the model unicellular eukaryote Saccharomyces cerevisiae, of which nearly one third of the genome corresponds to gene duplicates, the two modes of duplication have been shown to follow different evolutionary fates, with SSD genes being more prone to acquire novel functionalities (neofunctionalization) and WGD more likely to retain different parts of the original ancestral function (subfunctionalization). Having previously described aspects of functional compartmentalization for the genes of S. cerevisiae, in this work we set out to investigate the existence of positional preferences of gene duplicates. We found that SSD and WGD genes are organized in distinct gene clusters that are, furthermore, segregated, occupying different regions, with SSD being more peripheral and WGD more centrally positioned close to centromeric chromatin.

Duplicate gene clusters differ from the rest of the genome in terms of gene size and spacing, gene expression variability and regulatory complexity. What is more interesting, some of these properties are also shared by singleton genes residing in duplicate-rich regions in a position-dependent manner. Our analysis further reveals particular chromatin architectures in the promoters of duplicate genes, which are generally longer, with less pronounced nucleosome-free regions, strong structural constraints and a larger number of regulatory elements. Such structural features appear to be important for gene evolution as we find SSD gene-pair co-expression to be strongly associated with the similarity of nucleosome positioning patterns.

We propose that specific regions of the yeast genome provide a favourable environment for the generation and maintenance of small-scale gene duplicates. The existence of such genomic "niches" is supported by the enrichment of these regions in singleton genes bearing similarities with gene "relics", remnants of recent duplications that have reverted to single gene status. Our findings provide a valuable framework for the study of genomic innovation and suggest taking into account positional preferences in the study of gene emergence and fixation in experimentally and naturally evolving populations.


title: Three Mutations Convert the Selectivity of a Protein Sensor From Nicotinic Agonists to S-Methadone For Use in Cells, Organelles, and Biofluids

abstract: We report a reagentless, intensity-based S-methadone fluorescent sensor, iS-methadoneSnFR, consisting of a circularly permuted GFP inserted within the sequence of a mutated bacterial periplasmic binding protein (PBP). We evolved a previously reported nicotine-binding PBP to become a selective S-methadone-binding sensor, via three mutations in the PBPs second shell and hinge regions. iS-methadoneSnFR displays the necessary sensitivity, kinetics, and selectivity - notably enantioselectivity against R-methadone - for biological applications. Robust iS-methadoneSnFR responses in human sweat and saliva and mouse serum enable diagnostic uses. Expression and imaging in mammalian cells demonstrate that S-methadone enters at least two organelles and undergoes acid trapping in the Golgi apparatus, where opioid receptors can signal. This work shows a straightforward strategy in adapting existing PBPs to serve real-time applications ranging from subcellular to personal pharmacokinetics.

category: bioengineering


title: Structural dynamics of DNA strand break sensing by PARP-1 at a single-molecule level

abstract: Single stranded breaks (SSBs) are the most frequent DNA lesions threatening genomic integrity. A highly kinked DNA structure in complex with human PARP-1 led to the proposal that SSB sensing in Eukaryotes relies on dynamics of both the broken DNA double helix and PARP-1s multi-domain organization. Here, we directly probe this fundamental yet poorly understood process at the single-molecule level. Quantitative SM-FRET and structural ensemble calculations reveal how PARP-1 binding converts DNA SSBs from a largely unperturbed conformation into the highly kinked state. Binding of the second N-terminal zinc finger yields an intermediate DNA conformation that can be recognized by the first zinc finger. Thus, the data is inconsistent with a conformational selection model, instead an induced fit mechanism via a multi-domain assembly cascade drives SSBs sensing. Interestingly, a clinically used PARP-1 inhibitor niraparib shifts the equilibrium towards the unkinked state, whereas the inhibitor EB47 stabilizes the kinked state.

category: biophysics


title: PSMA-bearing extracellular vesicles secreted from prostate cancer convert the microenvironment to a tumor-supporting, pro-angiogenic state.

abstract: Extracellular vesicles (EV) are comprised of vesicles budding from cell membranes and smaller intracellular vesicles shed by cells. EV play a role in remodeling the tumor microenvironment (TME) and support tumor progression. Prostate-specific membrane antigen (PSMA) is a transmembrane glycoprotein with a carboxypeptidase function, frequently associated with poor clinical prognosis in prostate cancer (PCa). We previously identified an oncogenic PSMA signaling function in prostate cancer. Others demonstrated that EV isolated from the plasma of patients with high-grade PCa carry PSMA, but so far no pathophysiological effect has been associated with PSMA-bearing EV. Here we demonstrate that EV from PCa cells are able to transfer PSMA and its functionality to cells in the TME. The consequence of that EV-mediated PSMA transfer is an acute to long-term increased secretion of vascular endothelial growth factor-A (VEGF-A), angiogenin, pro-angiogenic and pro-lymphangiogenic mediators and increased 4E binding protein 1 (4EBP-1) phosphorylation in tumors. We compare EV from PCa cells with or without PSMA expression to address the role of PSMA-bearing EV in promoting pro-tumoral changes in the TME using classical molecular biology and novel molecular imaging approaches.

category: cancer biology


title: TGF-beta1 induced S100 family protein expression is associated with epithelial to mesenchymal transition states and poor survival in pancreatic cancer

abstract: Epithelial-mesenchymal transition (EMT) is a continuum that includes epithelial, partial EMT (P-EMT) and mesenchymal states, each of which are associated with cancer progression, invasive capabilities and ultimately metastasis. We have employed a lineage traced sporadic model of pancreatic cancer to generate a murine organoid biobank from primary and secondary tumors, including sublines that have undergone P-EMT and complete EMT (C-EMT). Using an unbiased proteomics approach, we found that the morphology of the organoids predicts the EMT state, with solid organoids associated with a P-EMT signature. We also observed that exogenous TGF{beta}1 induces a solid organoid morphology that is associated with changes in the S100 family, C-EMT and the formation of high-grade tumors. S100A4 may represent a useful biomarker to predict EMT state, disease progression and outcome for pancreatic cancer patients.


title: Development of a LAG-3 Immunohistochemistry Assay for Melanoma

abstract:
AimsA robust immunohistochemistry (IHC) assay was developed to detect lymphocyte-activation gene 3 (LAG-3) expression by immune cells (ICs) in tumor tissues. LAG-3 is an immuno-oncology target with demonstrable clinical benefit, and there is a need for a standardized, well-characterized assay to measure its expression. This study aims to describe LAG-3 scoring criteria and present the specificity, sensitivity, analytical precision, and reproducibility of this assay.

MethodsThe specificity of the assay was investigated by antigen competition and with LAG3 knockout cell lines. A melanin pigment removal procedure was implemented to prevent melanin interference in IHC interpretation. Formalin-fixed, paraffin-embedded (FFPE) human melanoma samples with a range of LAG-3 expression levels were used to assess the sensitivity and analytical precision of the assay with a [≥]1% cutoff to determine LAG-3-positivity. Interobserver and intraobserver reproducibility were evaluated with 60 samples in intralaboratory studies and 70 samples in interlaboratory studies.

ResultsThe LAG-3 IHC method demonstrated performance suitable for analysis of LAG-3 IC expression in clinical melanoma samples. The pretreatment step effectively removed melanin pigment that could interfere with interpretation. LAG-3 antigen competition and analysis of LAG3 knockout cell lines indicated that the 17B4 antibody clone binds specifically to LAG-3. The intrarun repeatability, interday, interinstrument, interoperator, and interreagent lot reproducibility demonstrated a high scoring concordance (>95%). The interobserver and intraobserver reproducibility and overall interlaboratory and intralaboratory reproducibility also showed high scoring concordance (>90%).

ConclusionsWe have demonstrated that the assay reliably assesses LAG-3 expression in FFPE human melanoma samples by IHC.

Key messagesO_ST_ABSWhat is already known on this topicC_ST_ABSLymphocyte-activation gene 3 (LAG-3) is an immune checkpoint receptor expressed on immune cells that limits T-cell activity and is being actively explored as a target for immunotherapy.

What this study addsAn immunohistochemistry assay was developed to detect the LAG-3 protein in formalin-fixed paraffin-embedded human tumor tissue specimens. This study describes scoring criteria and shows the specificity, sensitivity, analytical precision, and reproducibility of this assay as an aid to determine LAG-3 expression in melanoma patients using a [≥]1% expression on immune cells threshold.

How this study might affect research, practice or policyThe study describes a key immuno-oncology checkpoint immunohistochemistry assay that is robust and suitable for clinical trials. The assay was used in RELATIVITY-047 (NCT03470922), a phase 2/3 clinical trial that compared combined nivolumab and relatlimab treatment with nivolumab monotherapy, to stratify patients based on the percentage of LAG-3-positive immune cells within the tumor region. This assay is also being used in several ongoing clinical trials evaluating clinical response to relatlimab.

category: pathology


title: Nicotinamide Riboside alleviates exercise intolerance in ANT1-deficient mice.

abstract: Mitochondrial disorders are often characterized by muscle weakness and fatigue. Null mutations in the heart-muscle adenine nucleotide translocator isoform 1 (ANT1) of both humans and mice cause cardiomyopathy and myopathy associated with exercise intolerance and muscle weakness. We have analyzed the exercise physiology of mice deficient in ANT1, demonstrating a peripheral limitation of skeletal muscle mitochondrial respiration. Upon exercise, lack of Nicotinamide adenine dinucleotide+ (NAD+) results in a substrate limitation and stalling of the TCA cycle and mitochondrial respiration. Treatment of ANT1-deficient mice with nicotinamide riboside increased NAD+ levels in skeletal muscle and improved the exercise capacity and mitochondrial respiration. Thus, increasing NAD+ levels with nicotinamide riboside can alleviate the exercise intolerance associated with ANT1-deficiency, indicating the therapeutic potential of NAD+-stimulating compounds in specific mitochondrial myopathies.

category: physiology


title: The GENOMES UNCOUPLED1 protein has an ancient, highly conserved role in chloroplast gene expression but not in retrograde signalling

abstract:
O_LIThe pentatricopeptide repeat protein GENOMES UNCOUPLED1 (GUN1) is required for chloroplast-to-nucleus signalling in response to plastid stress during chloroplast development in Arabidopsis thaliana but its exact molecular function remains unknown.
C_LIO_LIWe analysed GUN1 sequences in land plants and streptophyte algae. We tested functional conservation by complementation of the Arabidopsis gun1 mutant with GUN1 genes from the streptophyte alga Coleochate orbicularis or the liverwort Marchantia polymorpha. We also analysed the transcriptomes of M. polymorpha gun1 knock-out mutant lines during chloroplast development.
C_LIO_LIGUN1 evolved within the streptophyte algal ancestors of land plants and is highly conserved among land plants but missing from the Rafflesiaceae that lack chloroplast genomes. GUN1 genes from C. orbicularis and M. polymorpha restore chloroplast retrograde signalling and suppress the cold-sensitive phenotype of the Arabidopsis gun1 mutant. However, GUN1 is not required for chloroplast retrograde signalling in the liverwort M. polymorpha.
C_LIO_LIOur findings suggest that GUN1 is an ancient protein that evolved within the streptophyte algal ancestors of land plants before the first plants colonised land more than 470 million years ago. Its primary role is likely to be in chloroplast gene expression and its role in chloroplast retrograde signalling probably evolved more recently.
C_LI

category: plant biology


title: Protection of Hamsters Challenged with SARS-CoV-2 after Two Doses of MVC-COV1901 Vaccine Followed by a Single Intranasal Booster with Nanoemulsion Adjuvanted S-2P Vaccine

abstract: Intramuscular vaccines have greatly reduced hospitalization and death due to severe COVID-19. However, most countries are experiencing a resurgence of infection driven predominantly by the Delta and Omicron variants of SARS-CoV-2. In response, booster dosing of COVID-19 vaccines has been implemented in many countries to address waning immunity and reduced protection against the variants. However, intramuscular boosting fails to elicit mucosal immunity and therefore does not solve the problem of persistent viral carriage and transmission, even in patients protected from severe disease. In this study, two doses of stabilized prefusion SARS-CoV-2 spike (S-2P)-based intramuscular vaccine adjuvanted with Alum/CpG1018, MVC-COV1901, were used as a primary vaccination series, followed by an intranasal booster vaccination with nanoemulsion (NE01)-adjuvanted S-2P vaccine in a hamster model to demonstrate immunogenicity and protection from viral challenge. Here we report that this vaccination regimen resulted not only in the induction of robust immunity and protection against weight loss and lung pathology following challenge with SARS-CoV-2, but also led to increased viral clearance from both upper and lower respiratory tracts. Our findings showed that intramuscular MVC-COV1901 vaccine followed by a booster with intranasal NE01-adjuvanted vaccine promotes protective immunity against both viral infection and disease, suggesting that this immunization protocol may offer a solution in addressing a significant, unmet medical need for both the COVID-19 and future pandemics.

category: immunology


title: Multi-omic analysis along the gut-brain axis points to a functional architecture of autism

abstract: Autism is a highly heritable neurodevelopmental disorder characterized by heterogeneous cognitive, behavioral and communication impairments. Disruption of the gut-brain axis (GBA) has been implicated in autism, with dozens of cross-sectional microbiome and other omic studies revealing autism-specific profiles along the GBA albeit with little agreement in composition or magnitude. To explore the functional architecture of autism, we developed an age and sex-matched Bayesian differential ranking algorithm that identified autism-specific profiles across 10 cross-sectional microbiome datasets and 15 other omic datasets, including dietary patterns, metabolomics, cytokine profiles, and human brain expression profiles. The analysis uncovered a highly significant, functional architecture along the GBA that encapsulated the overall heterogeneity of autism phenotypes. This architecture was determined by autism-specific amino acid, carbohydrate and lipid metabolism profiles predominantly encoded by microbial species in the genera Prevotella, Enterococcus, Bifidobacterium, and Desulfovibrio, and was mirrored in brain-associated gene expression profiles and restrictive dietary patterns in individuals with autism. Pro-inflammatory cytokine profiling and virome association analysis further supported the existence of an autism-specific architecture associated with particular microbial genera. Re-analysis of a longitudinal intervention study in autism recapitulated the cross-sectional profiles, and showed a strong association between temporal changes in microbiome composition and autism symptoms. Further elucidation of the functional architecture of autism, including of the role the microbiome plays in it, will require deep, multi-omic longitudinal intervention studies on well-defined stratified cohorts to support causal and mechanistic inference.

category: microbiology


title: Proteomic analysis of human milk reveals nutritional and immune benefits in the colostrum from mothers with COVID-19

abstract: The range of benefits breastfeeding provides neonates and infants include nutrition, improved neonatal survival, and reduced morbidity from certain diseases. It also aids maternal health by speeding postpartum recovery. However, due to concern about the risk of SARS-CoV-2 transmission and the lack of evidence of breastmilks protective effects against the virus, whether mothers with COVID-19 should be encouraged to breastfeed is under debate. Here, we present the results of proteomic and glycoproteomic studies of breast milk (colostrum and mature milk) from mothers with confirmed COVID-19. All colostrum samples exhibited significantly upregulated immune-related proteins, especially whey proteins with antiviral properties against SARS-CoV-2, and increased glycosylation levels and heterogeneity at those proteins. Such adaptive differences in milk from COVID-19 mothers tend to fade in mature milk from the same mothers one month postpartum. These results suggest the immune benefits of colostrum from mothers with COVID-19 and provide molecular-level insights that aid breastmilk feeding decisions in cases of active infection.

category: molecular biology


title: Genomic basis and phenotypic manifestation of (non-)parallel serpentine adaptation in Arabidopsis arenosa

abstract: Parallel evolution is common in nature and provides one of the most compelling examples of rapid environmental adaptation. In contrast to the recent burst of studies addressing genomic basis of parallel evolution, integrative studies linking genomic and phenotypic parallelism are scarce. Edaphic islands of toxic serpentine soils provide ideal systems for studying rapid parallel adaptation in plants, imposing strong, spatially replicated selection on recently diverged populations. We leveraged threefold independent serpentine adaptation of Arabidopsis arenosa and combined reciprocal transplants, ion uptake phenotyping, and available genome-wide polymorphisms to test if parallelism is manifested to a similar extent at both genomic and phenotypic levels. We found pervasive phenotypic parallelism in functional traits yet with varying magnitude of fitness differences that was congruent with neutral genetic differentiation between populations. Limited costs of serpentine adaptation suggest absence of soil-driven trade-offs. On the other hand, the genomic parallelism at the gene level was significant, although relatively minor. Therefore, the similarly modified phenotypes e.g., of ion uptake arose possibly by selection on different loci in similar functional pathways. In summary, we bring evidence for the important role of genetic redundancy in rapid adaptation involving traits with polygenic architecture.

category: evolutionary biology


title: Follow-up investigation and detailed mutational characterization of the SARS-CoV-2 Omicron variant lineages (BA.1, BA.2, BA.3 and BA.1.1)

abstract: Aided by extensive protein mutations, the SARS-CoV-2 Omicron (B.1.1.529) variant overtook the previously dominant Delta variant and rapidly spread around the world. It was shown to exhibit significant resistance to current vaccines and evasion from neutralizing antibodies. It is therefore critical to investigate the Omicron mutations trajectories. In this study, a literature search of published articles and SARS-CoV-2 databases was conducted, We explored the full list of mutations in Omicron BA.1, BA.1.1, BA.2, and BA.3 lineages. We described in detail the prevalence and occurrence of the mutations across variants, and how Omicron differs from them. We used GISAID as our primary data source, which provides open-access to genomics data of the SARS-CoV-2 virus, in addition to epidemiological and geographical data. We examined how these mutations interact with each other, their co-occurrence and clustering. Our study offers for the first time a comprehensive description of all mutations with a focus on non-spike mutations and demonstrated that mutations in regions other than the Spike (S) genes are worth investigating further. Our research established that the Omicron variant has retained some mutations reported in other SARS-CoV-2 variants, yet many of its mutations are extremely rare in other variants and unique to Omicron. Some of these mutations have been linked to the transmissibility and immune escape of the virus, and indicate a significant shift in SARS-CoV-2 evolution. The most likely theories for the evolution of the Omicron variant were also discussed.

category: genetics


title: Genomic and phenotypic comparisons reveal distinct variants of Wolbachia strain wAlbB

abstract:
The intracellular bacterium Wolbachia inhibits virus replication and is being harnessed around the world to fight mosquito-borne diseases through releases of mosquitoes carrying the symbiont. Wolbachia strains vary in their ability to invade mosquito populations and suppress viruses in part due to differences in their density within the insect and associated fitness costs. Using whole-genome sequencing, we demonstrate the existence of two variants in wAlbB, a Wolbachia strain being released in natural populations of Aedes aegypti mosquitoes. The two variants display striking differences in genome architecture and gene content. Differences in the presence/absence of 49 genes between variants include genes located in prophage regions and others potentially involved in controlling the symbionts density. Importantly, we show that these genetic differences correlate with variation in wAlbB density and its tolerance to heat stress, suggesting that different wAlbB variants may be better suited for field deployment depending on local environmental conditions. Finally, we found that the wAlbB genome remained stable following its introduction in a Malaysian mosquito population. Our results highlight the need for further genomic and phenotypic characterization of Wolbachia strains in order to inform ongoing Wolbachia-based programmes and improve the selection of optimal strains in future field interventions.

ImportanceDengue is a viral disease transmitted by Aedes mosquitoes that threatens around half of the world population. Recent advances in dengue control involve the introduction of Wolbachia bacterial symbionts with antiviral properties into mosquito populations which can lead to dramatic decreases in the incidence of the disease. In light of these promising results, there is a crucial need to better understand the factors affecting the success of such strategies, in particular the choice of Wolbachia strain for field releases and the potential for evolutionary changes. Here we characterized two variants of a Wolbachia strain used for dengue control that differ at the genomic level and in their ability to replicate within the mosquito. We also found no evidence for the evolution of the symbiont within the two years following its deployment in Malaysia. Our results have implications for current and future Wolbachia-based health interventions.


title: Structural basis for the simultaneous recognition of NEMO and acceptor ubiquitin by the HOIP NZF1 domain

abstract: Ubiquitination of NEMO by the linear ubiquitin chain assembly complex (LUBAC) is essential for activating the canonical NF-{kappa}B signaling pathway. While the NZF1 domain of the HOIP subunit of LUBAC recognizes the NEMO substrate, it is unclear how it cooperates with the catalytic domains in the ubiquitination process. Here, we report a crystal structure of NEMO in complex with HOIP NZF1 and linear diubiquitin chains, in which the two proteins bind to distinct sites on NEMO. Moreover, the NZF1 domain simultaneously interacts with NEMO and Ile44 surface of a proximal ubiquitin from a linear diubiquitin chain, where the C-term tail of the ubiquitin is in the proximity of the NEMO ubiquitination site (Lys285). We further propose a model for the linear ubiquitination of NEMO by HOIP. In the model, NZF1 binds the monoubiquitinated NEMO and recruits the catalytic domains to the ubiquitination site, thereby ensuring site-specific ubiquitination of NEMO.

category: biochemistry


title: Molecular modeling predicts novel antibody escape mutations in respiratory syncytial virus fusion glycoprotein

abstract:
Monoclonal antibodies are increasingly used for the prevention and/or treatment of viral infections. One caveat of their use is the ability of viruses to evolve resistance to antibody binding and neutralization. Computational strategies to predict which mutations will result in antibody resistance would be invaluable because current methods for identifying potential escape mutations are labor intensive and system-biased. Respiratory syncytial virus is an important pathogen for which monoclonal antibodies against the fusion (F) protein are used to prevent severe disease in high-risk infants. In this study, we used an approach that combines molecular dynamics simulations with FoldX to estimate changes in free energy in F protein folding and binding to the motavizumab antibody upon each possible amino acid change. We systematically selected 8 predicted escape mutations and tested them in an infectious clone. Consistent with our F protein stability predictions, replication-effective viruses were observed for each selected mutation. Six of the eight variants showed increased resistance to neutralization by motavizumab. Flow cytometry was used to validate the estimated (model-predicted) effects on antibody binding to F. Using surface plasmon resonance, we determined that changes in the on-rate of motavizumab binding were responsible for the reduced affinity for two novel escape mutations. Our study empirically validates the accuracy of our molecular modeling approach and emphasizes the role of biophysical protein modeling in predicting viral resistance to antibody-based therapeutics that can be used to monitor the emergence of resistant viruses and to design improved therapeutic antibodies.

ImportanceRespiratory syncytial virus (RSV) causes severe disease in young infants, particularly those with heart or lung diseases or born prematurely. As no vaccine is currently available, monoclonal antibodies are used to prevent severe RSV disease in high-risk infants. While it is known that RSV evolves to avoid recognition by antibodies, screening tools that can predict which changes to the virus will lead to antibody resistance are greatly needed.


title: Hypoconnectivity between anterior insula and amygdala in neonates with familial history of autism

abstract:
BackgroundAltered resting state functional connectivity (FC) involving the anterior insula (aINS), a key node in the salience network, has been reported consistently in autism.

MethodHere we examined, for the first time, FC between the aINS and the whole brain in a sample of full-term, postmenstrual age (PMA) matched neonates (mean 44.0 weeks, SD=1.5) who due to family history have high likelihood (HL) for developing autism (n=12) and in controls (n=41) without family history of autism (low likelihood, LL). Behaviors associated with autism were evaluated between 12 and 18 months (M=17.3 months, SD=2.5) in a subsample (25/53) of participants using the First Year Inventory (FYI).

ResultsCompared to LL controls, HL neonates showed hypoconnectivity between left aINS and left amygdala. Lower connectivity between the two nodes was associated with higher FYI risk scores in the social domain (r(25) = -.561, p=.003) and this association remained robust when maternal mental health factors were considered. Considering that a subsample of LL participants (n=14/41) underwent brain imaging during the fetal period at PMA 31 and 34 weeks, in an exploratory analysis, we evaluated prospectively development of the LaINS-Lamy connectivity and found that the two areas strongly coactivate throughout the third trimester of pregnancy.

ConclusionsThe study identifies left lateralized anterior insula - amygdala connectivity as a potential target of further investigation into neural circuitry that enhances likelihood of future onset of social behaviors associated with autism during neonatal and potentially prenatal periods.

category: neuroscience


title: Tango for Two: Modeling Synchronization and Social Interaction in Turn-Taking

abstract: Individuals can synchronize their body movements with regular auditory signals such as a metronome with a wide range of success. We investigate whether synchronizing with a regular auditory metronome is affected by each persons spontaneous production rate (SPR) and by hearing a partners synchronization in a social context. Musically trained and untrained participants synchronized their tapping with an auditory cue presented at different rates (their own SPR or their partners SPR) and in a Solo or Joint (turn-taking) condition. Linear and nonlinear oscillator models were fit to participants mean asynchronies (signed timing differences between the cued onsets and taps). Participants synchrony improved in Joint turn-taking trials when the auditory metronome was cued at the participants own SPR than at their partners SPR; in contrast, synchronization did not differ across rates in the Solo condition. Asynchronies in the Joint task became larger as the difference between partners spontaneous rates increased; the increased asynchronies were driven by the faster partner who did not slow down to match the rate of their slower partner. Nonlinear delay-coupled models (with time delay, coupling strength, and intrinsic frequency) outperformed linear models (intrinsic frequency only) in accounting for tappers synchronization adjustments. The nonlinear models coupling value was increased for musically trained participants, relative to untrained participants. Overall, these findings suggest that both intrinsic differences in partners spontaneous rates and the social turn-taking context contribute to the range of synchrony in the general population. Delay-coupled models are capable of capturing the wide range of individual differences in auditory-motor synchronization.


title: Sound-evoked responses of distinct neuron classes from the tail of the striatum

abstract: Given its inputs from auditory structures and neuromodulatory systems, the posterior tail of the striatum is ideally positioned to influence behavioral responses to acoustic stimuli according to context and previous rewards. Results from previous studies indicate that neurons in this striatal region display selective responses to sounds. However, it is not clear whether different striatal cell classes code for distinct features of sounds, nor how different striatal output pathways may use acoustic information to guide behavior. Here we compared the sound-evoked responses of posterior striatal neurons that form the striatal direct pathway (and express the dopamine receptor D1) to the responses of neighboring neurons in naive mice. We achieved this via optogenetic photo-identification of D1-expressing neurons during extracellular electrophysiological recordings in awake head-fixed mice of both sexes. We found that the frequency tuning of sound-responsive direct-pathway striatal neurons is comparable to that of their sound-responsive neighbors. Moreover, we found that both populations encode amplitude modulated sounds in a similar fashion. These results suggest that different classes of neurons in the posterior striatum of naive animals have similar access to acoustic features conveyed by the auditory system even outside the context of an auditory task.


title: Constructive Connectomics: how neuronal axons get from here to there using gene-expression maps derived from their family trees

abstract:
During brain development, billions of axons must navigate over multiple spatial scales to reach specific neuronal targets, and so build the processing circuits that generate the intelligent behavior of animals. However, the limited information capacity of the zygotic genome puts a strong constraint on how, and which, axonal routes can be encoded. We propose and validate a mechanism of development that can provide an efficient encoding of this global wiring task. The key principle, confirmed through simulation, is that basic constraints on mitoses of neural stem cells--that mitotic daughters have similar gene expression to their parent and do not stray far from one another--induce a global hierarchical map of nested regions, each marked by the expression profile of its common progenitor population. Thus, a traversal of the lineal hierarchy generates a systematic sequence of expression profiles that traces a staged route, which growth cones can follow to their remote targets. We have analyzed gene expression data of developing and adult mouse brains published by the Allen Institute for Brain Science, and found them consistent with our simulations: gene expression indeed partitions the brain into a global spatial hierarchy of nested contiguous regions that is stable at least from embryonic day 11.5 to postnatal day 56. We use this experimental data to demonstrate that our axonal guidance algorithm is able to robustly extend arbors over long distances to specific targets, and that these connections result in a qualitatively plausible connectome. We conclude that, paradoxically, cell division may be the key to uniting the neurons of the brain.

Author SummaryThe embryological development of each brain installs an essentially identical communication network between its cells that is roughly as complex as that between the billions of people living on Earth. Although vast scientific resources are currently applied to identifying the final pattern of connections, the connectome, there has until now been relatively little effort to answer the fundamental question of how this complex network across billions of neurons realized through the mitotic elaboration of the initial embryonic cell. The problem is sharpened by the constraints that construction of the network is limited by the information budget of the initial genome, and that it has no pre-existing address space for placing neurons and guiding axons. We explain how Biology can solve this problem by using the family tree of neurons to install a global space of molecular addresses, which axons can use to navigate from their source neuron to its relatives. We provide experimental evidence for this familial address space in gene expression patterns of the developing mouse brain, and demonstrate through simulation that the experimentally observed address space indeed supports global navigation to produce a qualitatively plausible default connectome.


title: Transient regulation of focal adhesion via Tensin3 is required for nascent oligodendrocyte differentiation

abstract: The differentiation of oligodendroglia from oligodendrocyte precursor cells (OPCs) to complex and extensive myelinating oligodendrocytes (OLs) is a multistep process that involves largescale morphological changes with significant strain on the cytoskeleton. While key chromatin and transcriptional regulators of differentiation have been identified, their target genes responsible for the morphological changes occurring during OL myelination are still largely unknown. Here, we show that the regulator of focal adhesion, Tensin3 (Tns3), is a direct target gene of Olig2, Chd7, and Chd8, transcriptional regulators of OL differentiation. Tns3 is transiently upregulated and localized to cell processes of immature OLs, together with integrin-{beta}1, a key mediator of survival at this transient stage. Constitutive Tns3 loss-of-function leads to reduced viability in mouse and humans, with surviving knockout mice still expressing Tns3 in oligodendroglia. Acute deletion of Tns3 in vivo, either in postnatal neural stem cells (NSCs) or in OPCs, leads to a two-fold reduction in OL numbers. We find that the transient upregulation of Tns3 is required to protect differentiating OPCs and immature OLs from cell death by preventing the upregulation of p53, a key regulator of apoptosis. Altogether, our findings reveal a specific time window during which transcriptional upregulation of Tns3 in immature OLs is required for OL differentiation likely by mediating integrin-{beta}1 survival signaling to the actin cytoskeleton as OL undergo the large morphological changes required for their terminal differentiation.


title: Transcription factor paralogs orchestrate alternative gene regulatory networks by context-dependent cooperation with multiple cofactors

abstract: In eukaryotes, members of large transcription factor families often exhibit similar DNA binding properties in vitro, yet initiate paralog-specific gene regulatory networks in vivo. The serially homologous first (T1) and third (T3) thoracic legs of Drosophila, which result from alternative gene regulatory networks specified by the Hox proteins Scr and Ubx, respectively, offer a unique opportunity to address this paradox in vivo. Genome-wide analyses using epitope-tagged alleles of both Hox loci in the T1 and T3 leg imaginal discs, which are the precursors to the adult appendages and ventral body regions, show that [~]8% of Hox binding is paralog-specific. Binding specificity is mediated by interactions with distinct cofactors in different domains: the known Hox cofactor Exd acts in the proximal domain and is necessary for Scr to bind many of its paralog-specific targets, while in the distal leg domain, we identified the homeodomain protein Distal-less (Dll) as a novel Hox cofactor that enhances Scr binding to a different subset of genomic loci. Reporter genes confirm the in vivo roles of Scr+Dll and suggest that [~]1/3 of paralog-specific Hox binding in enhancers is functional. Together, these findings provide a genome-wide view of how Hox paralogs, and perhaps paralogs of other transcription factor families, orchestrate alternative downstream gene networks and suggest the importance of multiple, context-specific cofactors.


title: A loss-of-function cysteine mutant in fibulin-3 (EFEMP1) forms aberrant extracellular disulfide-linked homodimers and alters extracellular matrix composition

abstract: Fibulin-3 (F3 or EFEMP1) is a disulfide-rich, secreted glycoprotein necessary for maintaining extracellular matrix (ECM) and connective tissue integrity. Two studies have identified distinct autosomal recessive F3 mutations in individuals with Marfan Syndrome-like phenotypes. Herein we characterized how one of these mutations, c.163T>C; p.Cys55Arg (C55R), disrupts F3 secretion, quaternary structure, and function by forming unique extracellular disulfide-linked homodimers. Dual cysteine mutants suggest that the C55R-induced disulfide species forms because of new availability of Cys70 on adjacent F3 monomers. Surprisingly, mutation of single cysteines located near C55 (i.e., C29, C42, C48, C61, C70, C159, and C171) also produced similar extracellular disulfide-linked dimers, suggesting that this is not a phenomenon isolated to the C55R mutant. To assess C55R functionality, F3 knockout (KO) retinal pigmented epithelial (RPE) were generated, followed by reintroduction of wild-type (WT) or C55R F3. F3 KO cells produced lower levels of the ECM remodeling enzyme, matrix metalloproteinase 2 and reduced formation of collagen VI ECM filaments, both of which were partially rescued by WT F3 overexpression. However, C55R F3 was unable to compensate for these same ECM-related defects. Our results highlight the unique behavior of particular cysteine mutations in F3 and uncover potential routes to restore C55R F3 loss-of-function.


title: Thermal regime during parental sexual maturation, but not during offspring rearing, modulates DNA methylation in brook charr (Salvelinus fontinalis)

abstract: Epigenetic inheritance can result in plastic responses to changing environments being faithfully transmitted to offspring. However, it remains unclear how epigenetic mechanisms such as DNA methylation can contribute to multigenerational acclimation and adaptation to environmental stressors. Brook charr (Salvelinus fontinalis), an economically important salmonid, is highly sensitive to thermal stress, and is of conservation concern in the context of climate change. We studied the effects of temperature during parental sexual maturation and offspring rearing on whole-genome DNA methylation in brook charr juveniles (fry). Parents were split between warm and cold temperatures during sexual maturation, mated in controlled breeding designs, then offspring from each family were split between warm (8{degrees}C) and cold (5{degrees}C) rearing environments. We found 188 differentially methylated regions (DMRs) due to parental maturation temperature after controlling for family structure. In contrast, offspring rearing temperature had a negligible effect on offspring methylation. Stable intergenerational inheritance of DNA methylation and minimal plasticity in progeny could result in transmission of acclimatory epigenetic states to offspring, priming them for a warming environment. Our findings have implications pertaining to the role of intergenerational epigenetic inheritance in response to ongoing climate change.


title: Vision-related convergent gene losses reveal SERPINE3's unknown role in the eye

abstract: Despite decades of research, knowledge about the genes that are important for development and function of the mammalian eye and are involved in human eye disorders remains incomplete. During mammalian evolution, mammals that naturally exhibit poor vision or regressive eye phenotypes have independently lost many eye-related genes. This provides an opportunity to predict novel eye-related genes based on specific evolutionary gene loss signatures. Building on these observations, we performed a genome-wide screen across 49 mammals for functionally uncharacterized genes that are preferentially lost in species exhibiting lower visual acuity values. The screen uncovered several genes, including SERPINE3, a putative serine proteinase inhibitor. A detailed investigation of 381 additional mammals revealed that SERPINE3 is independently lost in 18 lineages that typically do not primarily rely on vision, predicting a vision-related function for this gene. To test this, we show that SERPINE3 has the highest expression in eyes of zebrafish and mouse. In the zebrafish retina, serpine3 is expressed in Mueller glia cells, a cell type essential for survival and maintenance of the retina. A CRISPR-mediated knockout of serpine3 in zebrafish resulted in alterations in eye shape and defects in retinal layering. Furthermore, two human polymorphisms that are in linkage with SERPINE3 are associated with eye-related traits. Together, these results suggest that SERPINE3 has a role in vertebrate eyes. More generally, by integrating comparative genomics with experiments in model organisms, we show that screens for specific phenotype-associated gene signatures can predict functions of uncharacterized genes.


title: Human mesenchymal stem cells and derived extracellular vesicles reduce sensory neuron hyperexcitability and pain-related behaviors in a mouse model of osteoarthritis

abstract:
Osteoarthritis (OA) is a common degenerative joint disease characterized by joint pain and stiffness. In humans, mesenchymal stem cells (MSCs) and derived extracellular vesicles (MSC-EVs) have been reported to alleviate pain in knee OA. Here, we used the destabilization of the medial meniscus (DMM) mouse model of OA to investigate mechanisms by which MSCs and MSC-EVs influence pain-related behavior. We found that MSC and MSC-EV treated DMM mice displayed improved OA pain-related behavior (i.e. locomotion, digging and sleep) compared to untreated DMM mice. Improved behavior was not the result of reduced joint damage, but rather knee-innervating sensory neurons from MSC and MSC-EV treated mice did not display the hyperexcitability observed in untreated DMM mice. Furthermore, we found that MSC-EVs normalize sensory neuron hyperexcitability induced by nerve growth factor in vitro. Our study suggests that MSCs and MSC-EVs may reduce pain in OA by direct action on peripheral sensory neurons.

TeaserMesenchymal stem cells and secreted extracellular vesicles normalize sensory neuron excitability to reduce pain.


title: Using predictive validity to compare associations between brain damage and behavior

abstract: Lesion-behavior mapping (LBM) provides a statistical map of the association between voxel-wise brain damage and individual differences in behavior. To understand whether two behaviors are mediated by damage to distinct brain regions, researchers often compare LBM beta weight outputs by either the overlap method (emphasizing differences) or by the correlation method (emphasizing similarity). However, both methods lack a principled way to determine LBM distinctness and are disconnected from a major goal of LBMs, that of predicting behavior from brain damage. Without such criteria, researchers may draw conclusions from numeric differences between LBMs that are irrelevant to predicting behavior from brain damage. Here, we developed and validated a Predictive Validity Comparison method (PVC) that establishes such a criterion by directly comparing two LBMs based on the difference in predictive accuracy: two LBMs are significantly different if they provide unique predictive power for the two behaviors assessed. Applying the PVC to two lesion-behavior stroke data sets (Ding et al., 2020; Pustina et al., 2018) demonstrated the utility of a statistical criterion for determining when individual differences across behaviors result from the same vs. different lesion patterns. Using region-of-interest based simulations derived from proportion damage in Brodmann regions from a large group of participants (n=131), PVC detected when two behaviors were mediated by different regions (high sensitivity) and determined when two behaviors were mediated by the same region (high specificity). By objectively determining whether two behavioral deficits can be explained by a single vs. distinct patterns of brain damage, the PVC provides a critical advance in establishing the brain bases of behavior. We have released a GUI-driven, open-access web app of this method to encourage widespread adoption.


title: Activation of the actin/MRTF-A/SRF signalling pathway in pre-malignant mammary epithelial cells by P-cadherin is essential for transformation

abstract: Alterations in the expression or function of cell adhesion molecules have been implicated in all steps of tumour progression. Among those, P-cadherin expression is highly enriched in basal-like breast cancer, a molecular subset of triple-negative breast carcinomas, playing a central role in inducing cancer cell self-renewal, as well as collective cell migration and invasion capacity. To decipher the P-cadherin-dependent signalling network, we generated a humanised P-cadherin fly model, establishing a clinically relevant platform for functional exploration of P-cadherin effectors in vivo. We report that actin nucleators, MRTF and SRF are main effectors of P-cadherin functional effects. In addition, we validated these findings in a human mammary epithelial cell line with conditional activation of the Src oncogene, which recapitulates molecular events taking place during cellular transformation. We show that prior to triggering the gain of malignant phenotypes, Src induces a transient increase in P-cadherin expression levels, which correlates with MRTF-A accumulation, its nuclear translocation and the upregulation of SRF target genes. Moreover, knocking down P-cadherin, or preventing Factin polymerization with Latrunculin A, impairs SRF transcriptional activity. Furthermore, blocking MRTF-A nuclear translocation with CCG-203971 hampers proliferation, selfrenewal and invasion. Thus, in addition to sustaining malignant phenotypes, P-cadherin can also play a major role in the very early stages of breast carcinogenesis by promoting a transient boost of MRTF-A/SRF signalling through actin regulation.


title: Identification and characterisation of Klebsiella pneumoniae and Pseudomonas aeruginosa clinical isolates with atypical β-lactam susceptibility profiles using Orbitrap liquid chromatography-tandem mass spectrometry

abstract: There is significant interest in the possibility of predicting antibacterial drug susceptibility directly though the analysis of bacterial DNA or protein. We report the use of Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa and Acinetobacter baumannii transformants to define baseline predictive rules for the {beta}- lactam susceptibility profiles of {beta}-lactamase positive clinical isolates. We then deployed a robust and reproducible shotgun proteomics methodology to identify {beta}-lactamase positivity and predict {beta}-lactam susceptibility by reference to our baseline predictive rules both in cultured bacteria and in extracts of culture-positive blood. Proteomics and whole genome sequencing then allowed us to characterise K. pneumoniae and P. aeruginosa isolates that differed from the expected {beta}-lactam susceptibility profile, iteratively expanding our predictive rules. Proteomics added considerable value over and above the information generated by whole genome sequencing, allowing for gene expression, not just gene presence to be considered. Specifically, in K. pneumoniae, we identified key differences between acrR and ramR regulatory mutations and compared the effects of OmpK36 Aspartate-Threonine or Glycine-Aspartate dipeptide porin insertions on susceptibility to cefepime and carbapenems. In P. aeruginosa, we identified differences in the gene expression effects of mexR versus nalC mutations and related these to differences in {beta}-lactam MICs against isolates hyper-producing AmpC {beta}-lactamase and or producing a metallo-{beta}-lactamase.


title: What are the reference strains of Acinetobacter baumannii referring to?

abstract: We assembled the whole genome sequence (WGS) of a collection of 43 non-redundant modern clinical isolates and four broadly used reference strains of Acinetobacter baumannii. Comparison of these isolates and their WGS confirmed the high heterogeneity in capsule loci, sequence types, the presence of virulence and antibiotic resistance genes. However, a significant portion of clinical isolates strongly differ when compared to several reference strains in the light of colony morphology, cellular density, capsule production, natural transformability and in vivo virulence. These genetic and phenotypic differences between current circulating strains of A. baumannii and established reference strains could hamper the study of A. baumannii as an entity. The broadly used reference strains led to the current state of the art of the A. baumannii field, however, we propose that established reference strains in the A. baumannii field should be carefully used, because of the high genetic and phenotypic heterogeneities. In this study, we generated a collection of high-quality nucleotide sequences of 43 modern clinical isolates with the corresponding multi-level phenotypic characterizations. Beside the contribution of novel fundamental observations generated in this study, the phenotypic and genetic data, along with the bacterial strains themselves, will be further accessible using the first open access online platform called "Acinetobase". Therefore, a rational choice of modern strains will be possible to select the ones that suit the needs of specific biological questions.


title: Oropouche orthobunyavirus infection is mediated by the cellular host factor Lrp1

abstract: Oropouche orthobunyavirus (OROV; Peribunyaviridae) is a mosquito-transmitted virus that causes widespread cases of human febrile illness in South America, with occasional progression to neurologic effects. Host entry factors mediating cellular entry of OROV are undefined. Here, we show that OROV requires the host protein low density lipoprotein-related protein 1 (Lrp1) for cellular infection. Cells from evolutionarily distinct species lacking Lrp1 were less permissive to OROV infection than cells with Lrp1. Treatment of cells with either the high-affinity Lrp1 ligand receptor-associated protein (RAP) or recombinant ectodomain truncations of Lrp1 significantly reduced OROV infection. Thus, Lrp1 is an important host factor required for infection of cells by OROV. Taken together with our results showing that Lrp1 is a newly identified receptor for Rift Valley fever virus (RVFV), Lrp1 may play a broader role in cellular infection by bunyaviruses than previously appreciated.


title: Epigenetic signals that direct cell type specific interferon beta response in mouse cells

abstract: The antiviral response induced by type I interferon (IFN) via the JAK-STAT signaling cascade activates hundreds of IFN-stimulated genes (ISGs). While this response occurs essentially in all human and mouse tissues it varies between different cell types. However, the linkage between the underlying epigenetic features and the ISG pattern of a given cell is not well understood. We mapped ISGs, binding sites of the STAT1 and STAT2 transcription factors and chromatin features in three different mouse cell types (embryonic stem cells, neural progenitor cells and embryonic fibroblasts) before and after treatment with IFN{beta}. The analysis included gene expression, chromatin accessibility and histone H3 lysine modification by acetylation (ac) and mono-/tri-methylation (me1, me3). A large fraction of ISGs and STAT binding sites were cell type specific with promoter binding of a STAT1-STAT2 complex (STAT1/2) being a key driver of ISG induction. Furthermore, STAT1/2 binding to putative enhancers at intergenic and intronic sites induced ISG expression as inferred from a chromatin co-accessibility analysis. STAT1/2 binding was dependent on the chromatin context and positively correlated with pre-existing H3K4me1 and H3K27ac marks in an open chromatin state while the presence of H3K27me3 had an inhibitory effect. Thus, chromatin features present before stimulation represent an additional regulatory layer for the cell type specific antiviral response.


title: Obesity disrupts innate-adaptive immune network patterning in adipose tissue

abstract: Obesity drives significant changes in adipose tissue that precede development of tissue and systemic insulin resistance. Immune cell infiltration and inflammation are known contributors to these changes but there is limited understanding of their spatial context tissue-wide. We sought to identify spatial patterning in epididymal adipose tissue immune cells in a time course of diet-induced obesity in mice. Using spatial transcriptomics and single-cell RNA-sequencing, we identified dominant cell type signatures preserved in their anatomical context, quantified gene expression patterns at spots throughout adipose tissue, performed cell type network analysis, and investigated ligand-receptor colocalization. Our data support increased innate immune cells, including macrophages, monocytes, and innate lymphoid cells, with tissue-wide interspersion and dampened adaptive immune cell signatures with obesity. Network analysis identified increased heterogeneity in all major immune cell types, consistent with increased subtypes. To capture tissue dynamics at obesity onset, we draw on mathematical principles from linear algebra and spectral graph theory. We provide a framework for better understanding cell cooperation toward emergence of multicellular tissue function. Further, we adapt Turings mathematical theory on morphogenesis to show lapse of emergence in adipose tissue.

category: cell biology


title: Rapid estimation of cryptic adult abundance and breeding dynamics in a critically endangered elasmobranch from close-kin mark recapture

abstract: AbstractElasmobranchs are one of the most highly-threatened vertebrate taxa. Estimating abundance of spawning adults is often extremely challenging, yet crucial for prioritization of conservation measures. Emblematic of these challenges, the speartooth shark (Glyphis glyphis, Muller and Henle, 1839) was initially known only from rare specimens collected in a few tropical regions river systems of Australia and Papua New Guinea. Listed as critically endangered in Australia (where only six adults have ever been recorded) and endangered by the IUCN, such scarcity and limited distribution prevent direct assessments of abundance or reproductive biology. We used close-kin mark-recapture to estimate the abundance of mature G. glyphis in a genetically isolated population in the Wenlock River, Queensland, Australia. From 224 juvenile/sub-adults and 2 adult samples taken over three years (2013-2016), 46 half-sibling and 33 full-sibling pairs were found. The adult population was estimated at 897 (80% Credible interval 531 - 1684) with a sex ratio (based on mitochondrial DNA) highly skewed to males (females - 0.09: males - 0.91). Based on juveniles sampled in different years that shared a mother, 71% of females were estimated to skip-spawn. In an average year we estimate 44 breeding females occupy the system. Importantly, these methods constitute a viable and relatively rapid approach to obtain robust estimates of absolute abundance and other key population parameters for similar rare species.

category: ecology


title: Trophic strategies explain the ocean niches of small eukaryotic phytoplankton

abstract: A large fraction of marine primary production is performed by diverse small protists, and many of these phytoplankton are mixotrophs that also consume bacterial prey. However, the mechanisms structuring this diversity and its biogeochemical consequences remain poorly understood. Here we use isolates from seven major taxa to demonstrate tradeoffs between phototrophic and phagotrophic abilities. We then show that trophic strategy along the autotrophy-mixotrophy spectrum correlates strongly with global niche differences, across depths and across gradients of stratification and chlorophyll a. A model of competition shows that community shifts can be explained by greater fitness of faster-grazing mixotrophs when nutrients are scarce and light is plentiful. Our results illustrate how basic physiological constraints and principles of resource competition can organize complexity in the surface ocean ecosystem.


title: Dynamic Interrogation of Stochastic Transcriptome Trajectories Using Disease Associated Genes Reveals Distinct Origins of Neurological and Neuropsychiatric Disorders

abstract:
1The advent of open access to genomic data offers new opportunities to revisit old clinical debates while approaching them from a different angle. We examine anew the question of whether psychiatric and neurological disorders are different from each other by assessing the pool of genes associated with disorders that are understood as psychiatric or as neurological. We do so in the context of transcriptome data tracked as human embryonic stem cells differentiate and become neurons. Building upon probabilistic layers of increasing complexity, we describe the dynamics and stochastic trajectories of the full transcriptome and the embedded genes associated with psychiatric and/or neurological disorders. From marginal distributions of a genes expression across hundreds of cells, to joint interactions taken globally to determine degree of pairwise dependency, to networks derived from probabilistic graphs along maximal spanning trees, we have discovered two fundamentally different classes of genes underlying these disorders and differentiating them. One class of genes boasts higher variability in expression and lower dependencies ("active genes"); the other has lower variability and higher dependencies ("lazy genes"). They give rise to different network architectures and different transitional states. Active genes have large hubs and a fragile topology, whereas lazy genes show more distributed code during the maturation toward neuronal state. Lazy genes boost differentiation between psychiatric and neurological disorders also at the level of tissue across the brain, spinal cord, and glands. These genes, with their low variability and asynchronous ON/OFF states that have been treated as gross data and excluded from traditional analyses, are helping us settle this old argument at more than one level of inquiry.

2 Manuscript Contribution to the FieldThere is an ongoing debate on whether psychiatric disorders are fundamentally different from neurological disorders. We examine this question anew in the context of transcriptome data tracked as human embryonic stem cells differentiate and become neurons. Building upon probabilistic layers of increasing complexity, we describe the dynamics and stochastic trajectories of the full transcriptome and the embedded genes associated with psychiatric and/or neurological disorders. Two fundamentally different types of genes emerge: "lazy genes" with low, odd, and asynchronous variability patterns in expression that would have been, under traditional approaches, considered superfluous gross data, and "active genes" likely included under traditional computational techniques. They give rise to different network architectures and different transitional dynamic states. Active genes have large hubs and a fragile topology, whereas lazy genes show more distributed code during the maturation toward neuronal state. Under these new wholistic approach, the methods reveal that the lazy genes play a fundamental role in differentiating psychiatric from neurological disorders across more than one level of analysis. Including these genes in future interrogation of transcriptome data may open new lines of inquiry across brain genomics in general.


title: The SPOC domain is a phosphoserine binding module that bridges transcription machinery with co- and post-transcriptional regulators

abstract: The heptarepeats of the C-terminal domain (CTD) of RNA polymerase II (Pol II) are extensively modified throughout the transcription cycle. The CTD coordinates RNA synthesis and processing by recruiting transcription regulation factors as well as RNA capping, splicing and 3end processing factors. The SPOC domain of PHF3 was recently identified as a new CTD reader domain specifically binding to phosphorylated serine-2 residues in adjacent CTD repeats. Here, we establish the SPOC domains of the human proteins DIDO, SHARP and RBM15 as phosphoserine binding modules that can act as CTD readers but also recognize other phosphorylated binding partners. We report the crystal structure of SHARP (SPEN) SPOC-CTD and identify the molecular determinants for its specific binding to phosphorylated serine-5. PHF3 and DIDO SPOC domains preferentially interact with the Pol II elongation complex, while RBM15 and SHARP SPOC domains engage with the m6A writer and reader proteins. Our findings establish the SPOC domain as a major interface between the transcription machinery and regulators of transcription and co-transcriptional processes.


title: Metagenomic binning with assembly graph embeddings

abstract:
Despite recent advancements in sequencing technologies and assembly methods, obtaining high-quality microbial genomes from metagenomic samples is still not a trivial task. Current metagenomic binners do not take full advantage of assembly graphs and are not optimized for long-read assemblies. Deep graph learning algorithms have been proposed in other fields to deal with complex graph data structures. The graph structure generated during the assembly process could be integrated with contig features to obtain better bins with deep learning.

We propose GraphMB, which uses graph neural networks to incorporate the assembly graph into the binning process. We test GraphMB on long-read datasets of different complexities, and compare the performance with other binners in terms of the number of High Quality (HQ) genome bins obtained. With our approach, we were able to obtain unique bins on all real datasets, and obtain more bins on most datasets. In particular, we obtained on average 17.5% more HQ bins when compared to state-of-the-art binners and 13.7% when aggregating the results of our binner with the others. These results indicate that a deep learning model can integrate contig-specific and graph-structure information to improve metagenomic binning. GraphMB is available from https://github.com/MicrobialDarkMatter/GraphMB.

category: bioinformatics


title: Transient DNA binding to gapped DNA substrates links DNA sequence to the single-molecule kinetics of protein-DNA interactions

abstract: Protein interactions with nucleic acids are central to all genetic processes and many biotechnological applications. While many sequence-dependent protein-DNA interactions have been studied in detail using single-molecule methods, there is no standard high-throughput way to link the complex single-molecule kinetics of protein-DNA interactions with the DNA sequence of a single molecule. Here we provide the missing link by introducing a single-molecule imaging method (Gap-Seq) that interrogates DNA sequences via transient binding of short fluorescent DNA to a single DNA molecule previously used to characterise a protein-DNA interaction. In Gap-Seq, we identify a base by the degree of binding of 6-9 nt-long DNAs to surface-immobilised DNA substrates featuring a short single-stranded gap. To facilitate detection, we also developed a fluorescence quenching strategy that allows single-molecule detection at up to 500 nM of unbound fluorescent DNA. We link single-base differences on single DNA molecules to the kinetics of protein-DNA interactions by studying the interaction of a transcription activator with its cognate site. Finally, we show that our assay can address mixed sequences by distinguishing between two different sequences immobilised on the same field of view, paving the way for interrogation of sequence libraries for both mechanistic work and biotechnological applications.


title: Cryo-EM structures of Doravirine and Rilpivirine with HIV-1 Reverse Transcriptase/DNA Aptamer: Nonnucleoside Inhibitor Resistance by E138K and M184I Mutations

abstract:
Structures trapping a verity of functional and conformational states of HIV-1 reverse transcriptase (RT) have been determined by X-ray crystallography. These structures have played important roles in understanding the mechanisms of catalysis, inhibition and drug resistance, and in driving drug design. However, structures of several desired complexes of RT could not be obtained even after many crystallization or crystal soaking experiments. The ternary complexes of doravirine and rilpivirine with RT/DNA are such examples.

Structural study of HIV-1 RT by single-particle cryo-EM has been challenging due to the enzymes relatively smaller size and higher flexibility. We optimized a protocol for rapid structure determination of RT complexes by cryo-EM and determined six structures of wild-type and E138K/M184I mutant RT/DNA in complexes with the nonnucleoside inhibitors rilpivirine, doravirine, and nevirapine. RT/DNA/rilpivirine and RT/DNA/doravirine complexes have structural differences between them and from the representative conformation of RT/DNA/nevirapine (or efavirenz); the primer grip in the RT/DNA/doravirine and the YMDD motif in the RT/DNA/rilpivirine complexes have large shifts. The DNA primer 3-end in the doravirine-bound structure is positioned at the active site, but the complex is in a non-productive state. In the mutant RT/DNA/rilpivirine structure, I184 is stacked with the DNA such that their relative positioning can influence rilpivirine in the pocket. Simultaneously, E138K mutation widens the NNRTI-binding pocket entrance, potentially contributing to a faster rate of rilpivirine dissociation by E138K/M184I mutant RT, as reported by earlier kinetic studies. These structural differences have implications for drug design and for understanding molecular mechanisms of drug resistance.


title: Transition between conformational states of the TREK-1 K2P channel promoted by interaction with PIP2

abstract: Members of the TREK family of two-pore domain (K2P) potassium channels are highly sensitive to regulation by membrane lipids, including phosphatidylinositol-4,5-bisphosphate (PIP2). This study used coarse-grained molecular dynamics (CG-MD) and atomistic MD simulations to model the PIP2 binding site on both the up and down state conformations of TREK-1. We also calculated the free energy of PIP2 binding relative to other anionic phospholipids in both conformational states using potential of mean force (PMF) and free energy perturbation (FEP) calculations. Our results identify state-dependent binding of PIP2 to sites involving the proximal C-terminus and we show that PIP2 promotes a conformational transition from a down state towards an intermediate that resembles the up state. These results are consistent with functional data for PIP2 regulation and together provide evidence for a structural mechanism of TREK-1 channel activation by phosphoinositides.


title: Eribulin normalizes pancreatic cancer-associated fibroblasts by simulating TGFβ inhibition

abstract: Less than 11% of pancreatic cancer patients survive 5-years post-diagnosis. The unique biology of pancreatic cancer includes a significant expansion of its desmoplastic tumor microenvironment, wherein cancer-associated fibroblasts (CAFs) and their self-produced extracellular matrix are key components. CAF functions are both tumor-supportive and tumor-suppressive, while normal fibroblastic cells are solely tumor-suppressive. Knowing that CAF-eliminating drugs are ineffective and can accelerate cancer progression, therapies that "normalize" CAF function are highly pursued. Eribulin is a well-tolerated anti-microtubule drug used to treat a plethora of neoplasias, including advanced/metastatic cancers. Importantly, eribulin can inhibit epithelial to mesenchymal transition via a mechanism akin to blocking pathways induced by transforming growth factor-beta (TGF{beta}). Notably, canonical TGF{beta} signaling also plays a pivotal role in CAF activation, which is necessary for the development and maintenance of desmoplasia. Hence, we hypothesized that eribulin could modulate, and perhaps "normalize" CAF function. To test this premise, we used a well-established in vivo-mimetic fibroblastic cell-derived extracellular matrix (CDM) system and gauged the effects of eribulin on human pancreatic CAFs and cancer cells. The pathophysiologic cell-culturing system was also used to query eribulin effects on CDM-regulated cancer cell survival and invasive spread. Results demonstrated that intact CDMs modestly restricted eribulin from obstructing cancer cell growth. Nonetheless, eribulin-treated CAFs generated CDMs that limited cancer cell survival under nutritional stress, similar to reported tumor-suppressive CDMs generated by TGF{beta}-deficient CAFs. Data from this study support the central proposed premise and suggest that eribulin could be used as a CAF-normalizing drug.


title: A metabolically engineered bacterium controls autoimmunity by remodeling the pro-inflammatory microenvironment

abstract: Immunotherapy has led to impressive advances in the treatment of autoimmune and pro-inflammatory disorders; yet, its clinical outcomes remain limited by a variety of factors including the pro-inflammatory microenvironment (IME). Discovering effective immunomodulatory agents, and the mechanisms by which they control disease, will lead to innovative strategies for enhancing the effectiveness of current immunotherapeutic approaches. We have metabolically engineered an attenuated bacterial strain (i.e., Brucella melitensis 16M {Delta}vjbR, Bm{Delta}vjbR) to produce indole, a tryptophan metabolite that controls the fate and function of regulatory T cells (Tregs). We demonstrated that treatment with this strain polarized M2 macrophages (M{varphi}) which produced anti-inflammatory cytokines (e.g., IL-10) and promoted Treg function; moreover, when combined with adoptive cell transfer (ACT) of Tregs, a single treatment with our engineered bacterial strain dramatically reduced the incidence and score of autoimmune arthritis and decreased joint damage. These findings show how a metabolically engineered bacterium can constitute a powerful vehicle for improving the efficacy of immunotherapy, defeating autoimmunity and reducing inflammation by remodeling the IME and augmenting Treg function.


title: Human HLFneg placental erythro-myeloid progenitors give rise to HLA Class IIneg Hofbauer cells

abstract: The earliest macrophages are generated during embryonic development from erythro-myeloid progenitors (EMPs) via primitive haematopoiesis. This process is still poorly understood in humans but is generally thought to be spatially restricted to the yolk sac. Human fetal placental macrophages, Hofbauer cells (HBC), arise during the primitive haematopoietic wave, yet are unlikely to be yolk sac derived as they appear prior to placental vascularisation. Here we identify a population of placental erythro-myeloid progenitors (PEMPs) in the early human placenta that give rise to HBC. PEMP are fetal CD34+CD43+ progenitors found exclusively at early gestational timepoints. Transcriptomic analyses reveal that PEMP have a unique transcriptome with some conserved features of primitive yolk sac EMPs, including the lack of HLF expression. Using in vitro single-cell culture experiments we show that PEMP generate HBC-like cells which lack HLA-DR expression, a conserved feature of all fetal primitive macrophages in humans. These findings indicate that HBC are derived locally from PEMP and demonstrate that human primitive haematopoiesis is not restricted to the yolk sac, occurring also in the placenta.


title: NEDD8 activity is important for direct antigen MHC class I antigen presentation

abstract: Successful direct MHC class I antigen presentation is dependent on the protein degradation machinery of the cell to generate antigenic peptides which can be loaded onto MHC class I molecules for surveillance by CD8+ T cells of the immune system. Most often this process involves the ubiquitin-proteasome system, however other ubiquitin-like (UBL) proteins have also been implicated in protein degradation and direct antigen presentation. Here, we examine the role of neuronal precursor cell-expressed developmentally down-regulated protein 8 (NEDD8) in direct antigen presentation. NEDD8 is the UBL with highest similarity to ubiquitin and fusion of NEDD8 to the amino-terminus of a target protein can lead to the target proteins degradation. We find that appending NEDD8 to the N-terminus of the model antigen ovalbumin resulted in degradation by both the proteasome and autophagy protein degradation pathways, but only proteasomal degradation, involving the proteasomal subunit NEDD8 ultimate buster 1 (NUB1), resulted in peptide presentation. When directly compare to ubiquitin, NEDD8-fusion was less efficient at generating peptides. However, inactivation of the NEDD8-conugation machinery by treating cells with MLN4924, inhibited the presentation of peptides from Defective Ribosomal Products (DRiPs) derived from a model antigen. These results demonstrate that NEDD8 activity in the cell is important for direct antigen presentation, but not by directly targeting proteins for degradation.


title: Multiomic profiling defines cell fate plasticity of in vitro-derived islets

abstract: Insulin-secreting {beta}-cells within the pancreatic islets of Langerhans are lost in patients with type 1 diabetes1. Islets can be generated in vitro by differentiation of human pluripotent stem cells2-5, but understanding of the molecular events governing this process is limited. Here, we use single-cell multiomics to measure chromatin accessibility and transcriptional profiles of 120,064 cells to characterize and delineate maturation of islets from in vitro differentiation. We find distinguishing chromatin accessibility and gene expression signatures as well as dynamic profiles for each major islet cell type produced from in vitro differentiation. Furthermore, based on chromatin accessibility, {beta}, , and {delta} cells from in vitro differentiation are more ambiguous in their cellular identity than from isolated primary islets. However, extended time in vitro or with transplantation into mice drives more distinct states of chromatin accessibility for each cell type. Modulation of CTCF expression redirects cell fate from pancreatic islet endocrine to an intestinal enteroendocrine-like cell type. Additionally, knockdown of ARID1B enhances {beta}-cell maturation transcriptionally and by chromatin accessibility. These results provide a comprehensive atlas and insights into cell fate identity and maturation of stem cell-derived islets, which will inform on their utility for therapy and disease modeling6-8.


title: Hypoxia promotes osteogenesis via regulation of the mito-nuclear communication

abstract: Bone marrow mesenchymal stem cells (MSCs) reside in a hypoxic niche that maintains their differentiation potential. Several studies have highlighted the critical role of hypoxia (low oxygen concentration) in the regulation of stem cell function, reporting differentiation defects following a switch to normoxia (high oxygen concentration). However, the molecular events triggering changes in stem cell fate decisions in response to high oxygen remain elusive. Here, we study the impact of normoxia in the mito-nuclear communication, with regards to stem cell differentiation. We show that normoxia-cultured MSCs undergo profound transcriptional alterations which cause irreversible osteogenesis defects. Mechanistically, high oxygen promotes chromatin compaction and histone hypo-acetylation, particularly on promoters and enhancers of osteogenic genes. Although normoxia induces rewiring of metabolism, resulting in high acetyl-CoA levels, histone hypo-acetylation occurs due to trapping of acetyl-CoA inside mitochondria, likely due to lower CiC activity. Strikingly, restoring the cytosolic acetyl-CoA pool via acetate supplementation remodels the chromatin landscape and rescues the osteogenic defects. Collectively, our results demonstrate that the metabolism-chromatin-osteogenesis axis is heavily perturbed in response to high oxygen and identify CiC as a novel, oxygen-sensitive regulator of MSC function.


title: Maternal thyroid hormone increases neural cell diversity during zebrafish spinal cord neurodevelopment.

abstract: Maternally derived thyroid hormone (MT3) is a fundamental factor for vertebrate neurodevelopment. In humans, mutations on the T3 exclusive transporter monocarboxylic acid transporter 8 (MCT8) lead to the Allan-Herndon-Dudley syndrome (AHDS). Patients with AHDS present severe underdevelopment of the central nervous system with cognitive and locomotory consequences. Functional impairment of the zebrafish T3 exclusive membrane transporter MCT8 has been shown to phenocopy the symptoms observed in human patients with AHDS, thus providing an outstanding animal model to study this human condition. In this zebrafish model, MT3 acts as an integrator of different key developmental pathways during zebrafish neurodevelopment. Here we expand this knowledge by determining the developmental time of action of MT3 that occurs in very defined temporal intervals during zebrafish neurodevelopment. We have determined that MT3 is not involved in neural specification but is fundamental for developing particular neural progenitors and the consequent neural lineages that originate from them. Our data provide evidence that MT3 achieves this likely by modulation NOTCH signalling in a cell non-autonomous way. The findings show that MT3 expands the cell diversity output of neural progenitor cells, establishing a cellular background behind human AHDS and inherited limited CNS development.

category: developmental biology


title: Siderophore production and utilization by microbes in the North Pacific Ocean

abstract: Siderophores are strong iron-binding molecules produced and utilized by microbes to acquire the limiting nutrient iron (Fe) from their surroundings. Despite their importance as a component of the iron-binding ligand pool in seawater, data on the distribution of siderophores and the microbes that use them are limited. Here we measured the concentrations and types of dissolved siderophores during two cruises in April 2016 and June 2017 that transited from the iron-replete, low-macronutrient North Pacific Subtropical Gyre (NPSG) through the North Pacific Transition Zone (NPTZ) to the iron-deplete, high-macronutrient North Pacific Subarctic Frontal Zone (SAFZ). Surface siderophore concentrations in 2017 were higher in the NPTZ (4.0 - 13.9 pM) than the SAFZ (1.2 - 5.1 pM), which may be partly attributed to stimulated siderophore production by environmental factors such as dust-derived iron concentrations (up to 0.51 nM). Multiple types of siderophores were identified on both cruises, including ferrioxamines, amphibactins and iron-free forms of photoreactive siderophores, which suggest active production and use of diverse siderophores across latitude and depth. Additionally, the widespread genetic potential for siderophore biosynthesis and uptake across latitude and the variability in the taxonomic composition of bacterial communities that transcribe putative ferrioxamine, amphibactin and salmochelin transporter genes at different latitudes suggest that particular microbes actively produce and use siderophores, altering siderophore distributions and the bioavailability of iron across the North Pacific.


title: Microbes are potential key players in the evolution of life histories and aging in Caenorhabditis elegans

abstract: Microbes can have profound effects on host fitness and health and the appearance of late-onset diseases. Host-microbe interactions thus represent a major environmental context for healthy aging of the host and might also mediate trade-offs between life-history traits in the evolution of host senescence. Here we have used the nematode Caenorhadbitis elegans to examine whether host-microbe interactions might modulate the evolution of host life-history and aging. We first characterized the effects of two non-pathogenic Escherichia coli strains, a pathogenic E. coli strain and a pathogenic Serratia marcescens strain on the reproductive schedule and survival of an outbred C. elegans population, to be used in an experimental evolution study. Secondly, to investigate the dependency of these effects on host genotype, we assayed population growth rates and survival of five representative C. elegans inbred strains in response to these microbes. Our results show that host-microbe interactions have a substantial, host-genotype-dependent impact on reproductive effort and survival of the nematode host. Although pathogenic bacteria reduced host survival, as expected, they did not necessarily decrease host fertility or population growth rate. Given such microbe-specific genotypic differences in host life history, we predict that the evolution of reproductive schedules and senescence in this system might be critically contingent upon host-microbe interactions, a hypothesis which we will be testing using experimental evolution in future work.


title: Regional centromeres protect organisms from lethal mutations

abstract: Shorter centromeres have fewer kinetochore microtubules (kMTs) bound as evident in Saccharomyces cerevisiae where each chromosome is associated with a single kMT. Cells can also be conditioned in recruiting excess kMTs highlighting the flexibility of the kinetochore as the kMT binding interface. Here, we addressed the evolutionary significance of the inherent flexibility in conditionally recruiting excess kMTs. We identified a conserved arginine residue at the C- terminus of an essential outer kinetochore protein Dad2 that remained positively selected for [~]700 million years in fungi. While the conserved R126 residue in Dad2 is essential for viability in S. cerevisiae harboring short point centromeres, the corresponding arginine residues are functionally important but no longer essential for viability in either Candida albicans or Cryptococcus neoformans, each possessing longer regional centromeres. We revealed a gradual decline in the requirement of the conserved arginine for chromosome biorientation and mitotic progression with increasing centromere length. Finally, our results suggested that the mutational tolerance in organisms with regional centromeres is imparted by higher kinetochore protein levels known to recruit multiple kMTs per chromosome.


title: Protein-protein interaction prediction for targeted protein degradation

abstract:
Protein-protein interactions (PPIs) play a fundamental role in various biological functions; thus, detecting PPI sites is essential for understanding diseases and developing new drugs. PPI prediction is of particular relevance for the development of drugs employing targeted protein degradation, as their efficacy relies on the formation of a stable ternary complex involving two proteins. However, experimental methods to detect PPI sites are both costly and time-intensive. In recent years, computer-aided approaches have been developed as screening tools, but these tools are primarily based on sequence information and are therefore limited in their ability to address spatial requirements and have thus far not been applied to targeted protein degradation.

Here, we present a new deep learning architecture based on the concept of graph representation learning that can predict interaction sites and interactions of proteins based on their surface representations. We demonstrate that our model reaches state-of-the-art performance using AUROC scores on the established MaSIF dataset. We furthermore introduce a new dataset with more diverse protein interactions and show that our model generalizes well to this new data. These generalization capabilities allow our model to predict the PPIs relevant for targeted protein degradation, which we show by demonstrating the high accuracy of our model for PPI prediction on the available ternary complex data. Our results suggest that PPI prediction models can be a valuable tool for screening protein pairs while developing new drugs for targeted protein degradation.


title: Meaningful Local Signaling in SA Node Center Identified by Random Matrix Theory

abstract: AO_SCPLOWBSTRACTC_SCPLOWThe sinoatrial node (SAN) is the pacemaker of the heart. Recently calcium signals, believed to be crucially important in rhythm generation, have been imaged in intact SAN and shown to be heterogeneous in various regions of the SAN. However, calcium imaging is noisy, and the calcium signal heterogeneity has not been mathematically analyzed to distinguish meaningful signals from randomness or to identify signalling regions in an objective way. In this work we apply methods of random matrix theory (RMT) developed for financial data and used for analysis of various biological data sets including {beta}-cell collectives and EEG data to analyse correlations in calcium signals in the SAN. We find eigenvalues of the correlation matrix that deviate from RMT predictions, and thus are not explained by randomness but carry additional meaning. We use localization properties the eigenvectors corresponding to high eigenvalues to locate particular signalling modules. We find that the top eigenvector captures the common response of the SAN to action potential. In some cases, the eigenvector corresponding to the second highest eigenvalue appears to yield a possible pacemaker region as its calcium signals predate the action potential. Next we study the relationship between covariance coefficients and distance and find that in many cases even inside the central zone, there are non-trivial long range correlations, indicating intercellular interactions in most cases. Lastly, we perform an analysis of nearest neighbor eigenvalue distances and find that it coincides with the universal Wigner surmise for all videos under all available experimental conditions, which means that it is the same as in energies of heavy atoms, buses in Cuernavaca, correlations of financial returns, and a multitude of other systems. On the other hand, the number variance, which captures eigenvalue correlations, is a parameter that is sensitive to experimental conditions. Thus RMT application to SAN allows to remove noise and the global effects of the action potential and thereby isolate the correlations in calcium signalling which are local. Thus indentified local signals in the SAN center can be meaningful and represent the emergent pacemaker or the signal integrator module of the SAN.


title: Polyelectrolyte brush bilayers at thermal equilibrium: Density functional theory and molecular dynamic simulations

abstract:
In this article, the equilibrium properties of Polyelectrolyte brush bilayers is studied. The density functional theory framework as well as the benchmark molecular dynamic simulations are employed to indicate the correct equation of state. Both techniques turn out that the equation of state neutral polymer brush bilayers scales with the wall distance as D-4. Additionally, it turns out that the electrostatic interactions does not change the universal power laws. The only difference is that the charged systems posses larger pressure due to stored electrostatic energy.

SIGNIFICANCEStudying Polyelectrolyte brush bilayers is important because it exists widely in nature, biology and industry. In biology, they could be found in mammalian synivial joints as well as in the cellular structures.


title: Activation of the neonatal immune system in healthy neonates within hours of birth.

abstract: It is now established that immune maturation occurs along a defined trajectory in the weeks and months after birth, but the immediate changes that occur within immune cells following birth is less clear. In this study, we monitored the immune profile of neonates via analysis of paired samples (n= 28) of cord blood and heel prick blood taken at varying times post term delivery by caesarean section. This paired approach accounted for the between-subject variability often observed over the first week of life. We identified rapid changes in immune cell populations within hours of birth. Specifically, we observed increased proliferation in effector T cells (but not regulatory T cells) that exhibited an increase in cytokine producing ability and also an increase in the percentage of CD3 T cells over this short time frame. This indicates that the mobilisation of the immune system is immediate post birth, presumably as a response to sudden exposure to the external environment, antigen or stress. Hence, immune development may start to occur more rapidly than previously proposed and as such, to study this trajectory, blood sampling should begin as soon after birth as possible.


title: Identification of novel leaf rust seedling resistance loci in Iranian bread wheat germplasm using genome-wide association mapping

abstract: Leaf or brown rust caused by Puccinia triticina Eriks. (Pt) is a major biotic constraint threatening bread wheat production worldwide. The continued evolution of new races of Pt necessitates a constant search for the identification of new resistance genes, or QTLs, to enhance the resistance durability of bread varieties. On a panel of 320 bread wheat accessions, we used a genome-wide association study (GWAS) technique to map loci associated with Pt resistance using single-nucleotide polymorphism markers (SNPs) generated by genotyping-by-sequencing (GBS). The panel was tested with five Pt races gathered from different regions of IRAN to identify loci associated with seedling resistance. After estimating genetic relatedness and population structure among accessions, GWAS discovered a total of 19 SNPs on chromosomes 1B, 2B, 3A, 3B, 4A, 5B, 5D, 6A, 6B, 6D, 7B, and 7D that were significantly associated with seedling stage resistance. The three SNP markers rs12954, rs34220, and rs42447 on chromosomes 5D, 6A, and 7D, respectively, associated with resistance to Pt race PKTTS expressing potential new loci for leaf rust resistance. Overall, this research gives an integrated perspective of leaf rust resistance resources in Iranian bread wheat and recognizes new resistance loci that will be valuable to expand the set of resistance genes available to control this serious disease.


title: Meta-Analysis of the Functional Neuroimaging Literature with Probabilistic Logic Programming

abstract: Inferring reliable brain-behavior associations requires synthesizing evidence from thousands of functional neuroimaging studies through meta-analysis. However, existing meta-analysis tools are limited to investigating simple neuroscience concepts and expressing a restricted range of questions. Here, we expand the scope of neuroimaging meta-analysis by designing NeuroLang: a domain-specific language to express and test hypotheses using probabilistic first-order logic programming. By leveraging formalisms found at the crossroads of artificial intelligence and knowledge representation, NeuroLang provides the expressivity to address a larger repertoire of hypotheses in a meta-analysis, while seamlessly modelling the uncertainty inherent to neuroimaging data. We demonstrate the languages capabilities in conducting comprehensive neuroimaging meta-analysis through use-case examples that address questions of structure-function associations. Specifically, we infer the specific functional roles of three canonical brain networks, support the role of the visual word-form area in visuospatial attention, and investigate the heterogeneous organization of the fronto-parietal control network.