title: Circadian Clock Core Component Bmal1 Dictates Cell Cycle Rhythm of Proliferating Hepatocytes during Liver Regeneration

abstract:
Following partial hepatectomy (PH), the majority of remnant hepatocytes synchronously enter and rhythmically progress through the cell cycle for three major rounds to regain lost liver mass. Whether and how the circadian clock core component Bmal1 modulates this process remains elusive. We performed PH on Bmal1+/+ and hepatocyte-specific Bmal1 knockout (Bmal1hep-/-) mice and compared the initiation and progression of the hepatocyte cell cycle. After PH, Bmal1+/+ hepatocytes exhibited three major waves of nuclear DNA synthesis. In contrast, in Bmal1hep-/-hepatocytes, the first wave of nuclear DNA synthesis was delayed by 12 h, and the third such wave was lost. Following PH, Bmal1+/+ hepatocytes underwent three major waves of mitosis, whereas Bmal1hep-/-hepatocytes fully abolished mitotic oscillation. These Bmal1-dependent disruptions in the rhythmicity of hepatocyte cell cycle after PH were accompanied by suppressed expression peaks of a group of cell cycle components and regulators, and dysregulated activation patterns of mitogenic signaling molecules c-Met and EGFR. Moreover, Bmal1+/+ hepatocytes rhythmically accumulated fat as they expanded following PH, whereas this phenomenon was largely inhibited in Bmal1hep-/-hepatocytes. In addition, during late stages of liver regrowth, Bmal1 absence in hepatocytes caused the activation of redox sensor Nrf2, suggesting an oxidative stress state in regenerated liver tissue. Collectively, we demonstrated that during liver regeneration, Bmal1 partially modulates the oscillation of S-phase progression, fully controls the rhythmicity of M-phase advancement, and largely governs fluctuations in fat metabolism in replicating hepatocytes, and eventually determines the redox state of regenerated livers.

New and NoteworthyWe demonstrated that Bmal1 centrally controls the synchronicity and rhythmicity of the cell cycle and lipid accumulation in replicating hepatocytes during liver regeneration. Bmal1 plays these roles, at least in part, by ensuring formation of the expression peaks of cell cycle components and regulators, as well as the timing and levels of activation of mitogenic signaling molecules.

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category: pathology


title: Soma and Neurite Density MRI (SANDI) of the in-vivo mouse brain

abstract: Diffusion MRI (dMRI) provides unique insights into the neural tissue milieu by probing interaction of diffusing molecules and tissue microstructure. Most dMRI techniques focus on white matter tissues (WM) due to the relatively simpler modelling of diffusion in the more organized tracts; however, interest is growing in gray matter characterisations. The Soma and Neurite Density MRI (SANDI) methodology harnesses a model incorporating water diffusion in spherical objects (assumed to be associated with cell bodies) and in impermeable "sticks" (representing neurites), which potentially enables the characterisation of cellular and neurite densities. Recognising the importance of rodents in animal models of development, aging, plasticity, and disease, we here sought to develop SANDI for preclinical imaging and provide a validation of the methodology by comparing its metrics with the Allen mouse brain atlas. SANDI was implemented on a 9.4T scanner equipped with a cryogenic coil, and experiments were carried out on N=6 mice. Pixelwise, ROI-based, and atlas comparisons were performed, and results were also compared to more standard Diffusion Kurtosis MRI (DKI) metrics. We further investigated effects of different pre-processing pipelines, specifically the comparison of magnitude and real-valued data, as well as different acceleration factors. Our findings reveal excellent reproducibility of the SANDI parameters, including the sphere and stick fraction as well as sphere size. More strikingly, we find a very good rank correlation between SANDI-driven soma fraction and Allen brain atlas contrast (which represents the cellular density in the mouse brain). Although some DKI parameters (FA, MD) correlated with some SANDI parameters in some ROIs, they did not correlate nearly as well as SANDI parameters with the Allen atlas, suggesting a much more specific nature of the SANDI parameters. We conclude that SANDI is a viable preclinical MRI technique that can greatly contribute to research on brain tissue microstructure.

category: neuroscience


title: The light chain of the L9 antibody is critical for binding circumsporozoite protein minor repeats and preventing malaria

abstract: L9 is a potent human monoclonal antibody (mAb) that preferentially binds two adjacent NVDP minor repeats and cross-reacts with NANP major repeats of the Plasmodium falciparum circumsporozoite protein (PfCSP) on malaria-infective sporozoites. Understanding this mAbs ontogeny and mechanisms of binding PfCSP to neutralize sporozoites will facilitate vaccine development. Here, we isolated mAbs clonally related to L9 and showed that this B-cell lineage has baseline NVDP affinity and evolves to acquire NANP reactivity. Pairing the L9 kappa light chain (L9{kappa}) with clonally-related heavy chains resulted in chimeric mAbs that cross-linked two NVDP, cross-reacted with NANP, and more potently neutralized sporozoites compared to their original light chain. Structural analyses revealed that chimeric mAbs bound the minor repeat motif in a type-1 {beta}-turn seen in other repeat-specific antibodies. These data highlight the importance of L9{kappa} in binding NVDP on PfCSP to neutralize SPZ and suggest that PfCSP-based immunogens might be improved by presenting [≥]2 NVDP.

category: immunology


title: Single-cell lineage trajectories and chromatin regulators that initialize antiviral CD8 T cell ontogeny

abstract: Individual naive CD8 T cells activated in lymphoid organs differentiate into functionally diverse and anatomically distributed T cell phylogenies in response to intracellular microbes. During infections that resolve rapidly, including live viral vaccines1, distinct effector (TEFF) and memory (TMEM) cell populations develop that ensure long term immunity2. During chronic infections, responding cells progressively become dysfunctional and "exhaust"3. A diverse taxonomy of TEFF, TMEM and exhausted (TEX) CD8 T cell populations is known, but the initial developmental basis of this phenotypic variation remains unclear4-10. Here, we defined single-cell trajectories and identified chromatin regulators that establish antiviral CD8 T cell heterogeneity using unsupervised analyses of single-cell RNA dynamics11-13 and an in vivo RNAi screen14. Activated naive cells differentiate linearly into uncommitted effector-memory progenitor (EMP) cells, which initially branch into an analogous manifold during either acute or chronic infection. Disparate RNA velocities in single EMP cells initiate divergence into stem, circulating, and tissue-resident memory lineages that generate diverse TMEM and TEX precursor states in specific developmental orders. Interleukin-2 receptor (IL-2R) signals are essential for formation and transcriptional heterogeneity of EMP cells, and promote trajectories toward TEFF rather than TEX states. Nucleosome remodelers Smarca4 and Chd7 differentially promote transcription that delineates divergent TMEM lineages before cooperatively driving terminal TEFF cell differentiation. Thus, the lineage architecture is established by specific chromatin regulators that stabilize diverging transcription in uncommitted progenitors.


title: Exosomal miR-106b-5p promotes Mtb survival via targeting CREB5 followed by SOAT1-CIDEC and CASP9-CASP3 pathway

abstract: Tuberculosis(TB) is one of the top ten fatal diseases, but the research on the mechanism of TB is still not perfect. Exosome, as an important intercellular signal transmission signal vehicle and the mechanism of exosomes in the interaction between macrophages and Mycobacterium tuberculosis(Mtb), is crucial for TB treatment. In the study, we found that exosomes, derived from Mtb-infected macrophage, exhibited differential enrichment in different organs in mice, causing inflammatory cell infiltration in lungs. Further experiments in vitro showed that exosomes resulted in increased lipid synthesis and inhibition of apoptosis in normal macrophages. In order to further explore its molecular mechanism, bioinformatics analysis showed that miR-106b-5p was up-regulated in exosomes. Subsequently, we verified miR-106b-5p was increased through a large number of blood samples from TB patients. In addition, we demonstrated that miR-106b-5p was upregulated in exosomes from Mtb-infected macrophages, which can be engulfed by uninfected macrophages and further result in miR-106b-5p increase. We next found that miR-106b-5p mediated the same effect as the exosomes derived from infected macrophage. Through further research, we indicated that miR-106b-5p promoted lipid droplet accumulation through regulation of Creb5-SOAT1-CIDEC and suppressed macrophage apoptosis via Creb5-CASP9-CASP3 pathway, which ultimately led to Mtb survival. These findings provide a certain theoretical basis and ideas for the diagnosis and treatment of TB as well as the selection of biomarkers.

category: microbiology


title: Evaluation of the speed, accuracy and precision of the QuickMIC rapid antibiotic susceptibility testing assay in a clinical setting

abstract: The rapidly changing landscape of antimicrobial resistance poses a challenge for empirical antibiotic therapy in severely ill patients and highlights the need for fast antibiotic susceptibility diagnostics to guide treatment. Traditional methods for antibiotic susceptibility testing (AST) of bacteria such as broth microdilution (BMD) or the disc diffusion method (DDM) are comparatively slow and show high variability. Rapid AST methods under development often trade speed for resolution, sometimes only measuring responses at a single antibiotic concentration. QuickMIC is a recently developed lab-on-a-chip system for rapid AST. Here we evaluate the performance of the QuickMIC method with regard to speed, precision and accuracy in comparison to traditional diagnostic methods. 151 blood cultures of clinical Gram-negative isolates with a high frequency of drug resistance were tested with the QuickMIC system and compared with BMD for 12 antibiotics. To investigate sample turnaround time and functionality in a clinical setting, another 41 clinical blood culture samples were acquired from the Uppsala University Hospital and analyzed on site in the clinical laboratory with the QuickMIC system, and compared with DDM for 8 antibiotics routinely used in the clinical laboratory. The overall essential agreement between MIC values obtained by QuickMIC and BMD was 83.2%, with an average time to result of 3 h 2 min (SD: 24.8 min) for the QuickMIC method. For the clinical dataset, the categorical agreement was 94.9%, and the total turnaround time as compared to routine diagnostics reduced by 40% (33 h vs. 55 h). Interexperiment variability was low (average SD: 44.6% from target MIC) compared to the acceptable standard of {+/-}1 log2 unit (i.e. -50% to +100% deviation from target MIC) in BMD. We conclude that the QuickMIC method can provide rapid and accurate AST, and may be especially valuable in settings with high resistance rates and for antibiotics where wildtype and resistant MIC distributions are close or overlapping.


title: Quantitative analysis of nuclear pore complex organization in Schizosaccharomyces pombe

abstract: The number, distribution and composition of nuclear pore complexes (NPCs) in the nuclear envelope (NE) varies between cell types and changes during cellular differentiation and in disease. To understand how NPC density and organization is controlled, we analyzed NPC number and distribution in the fission yeast Schizosaccharomyces pombe using structured illumination microscopy. The small size of yeast nuclei, genetic features of fungi and our robust image analysis pipeline allowed us to study NPCs in intact nuclei under multiple conditions. Our data revealed that NPC density is maintained across a wide range of nuclear sizes. Regions of reduced NPC density are observed over the nucleolus and surrounding the spindle pole body (SPB). Lem2-mediated tethering of the centromeres to the SPB is required to maintain NPC exclusion, which is important for timely mitotic progression. These findings provide a quantitative understanding of NPC number and distribution in S. pombe and show that interactions between the centromere and the NE influences local NPC distribution.

category: cell biology


title: Identification and characterization of distinct cell cycle stages in cardiomyocytes using the FUCCI transgenic system

abstract:
Understanding the regulatory mechanism by which cardiomyocyte proliferation transitions to endoreplication and cell cycle arrest during the neonatal period is crucial for identifying proproliferative factors and developing regenerative therapies.

We used a transgenic mouse model based on the fluorescent ubiquitination-based cell cycle indicator (FUCCI) system to isolate and characterize cycling cardiomyocytes at different cell cycle stages at a single-cell resolution. Single-cell transcriptome analysis of cycling and noncycling cardiomyocytes was performed at postnatal days 0 (P0) and 7 (P7).

The FUCCI system proved to be efficient for the identification of cycling cardiomyocytes with the highest mitotic activity at birth, followed by a gradual decline in the number of cycling and mitotic cardiomyocytes during the neonatal period. Cardiomyocytes showed premature cell cycle exit at G1/S shortly after birth and delayed G1/S progression during endoreplication at P7. Single-cell RNA-seq confirmed previously described signaling pathways involved in cardiomyocyte proliferation (Erbb2 and Hippo/YAP), cardiomyocyte motility, and maturation-related transcriptional changes during postnatal development, including the metabolic switch from glycolysis to fatty acid oxidation in cardiomyocytes. Additionally, we generated transcriptional profiles specific to cell division and endoreplication in cardiomyocytes.

Deciphering transcriptional changes at different developmental stages and in a cell cycle-specific manner may facilitate the identification of genes important for adult cardiomyocyte proliferation and heart regeneration.

Main findingsFUCCI reliably identifies cycling cardiomyocytes at distinct cell cycle stages in neonatal, juvenile, and adult hearts.

Cell cycle activity decreases as the metabolic switch transitions from glycolysis to fatty acid metabolism in postnatal cardiomyocytes.

Cell cycle arrest at G1/S is linked to the DNA damage response in postnatal cardiomyocytes. Distinct gene expression patterns are linked to different cell cycle phases in dividing and endoreplicating postnatal cardiomyocytes.


title: Open reading frame correction using antisense oligonucleotides for the treatment of cystic fibrosis caused by CFTR-W1282X

abstract:
CFTR gene mutations that result in the introduction of premature termination codons (PTCs) are common in cystic fibrosis (CF). This mutation type causes a severe form of the disease, likely because of low CFTR mRNA expression as a result of nonsense mediated mRNA decay (NMD), as well as production of a non-functional, truncated CFTR protein. Current therapeutics for CF, which target residual protein function, are less effective in patients with these types of mutations, due in part to low CFTR protein levels. Splice-switching antisense oligonucleotides (ASOs) designed to induce skipping of exons in order to restore the mRNA open reading frame have shown therapeutic promise pre-clinically and clinically for a number of diseases. We hypothesized that ASO-mediated skipping of CFTR exon 23 would recover CFTR activity associated with terminating mutations in the exon, including CFTR p.W1282X, the 5th most common mutation in CF. Here, we show that CFTR lacking the amino acids encoding exon 23 is partially functional and responsive to corrector and modulator drugs currently in clinical use. ASO-induced exon 23 skipping rescued CFTR expression and chloride current in primary human bronchial epithelial cells isolated from homozygote CFTR-W1282X patients. These results support the use of ASOs in treating CF patients with CFTR class I mutations in exon 23 that result in unstable CFTR mRNA and truncations of the CFTR protein.

Significance StatementFrameshift and nonsense mutations pose a major problem for disease therapeutic development. Eliminating these mutations from the mRNA by inducing exon skipping is a relatively unexplored treatment approach, though it has shown promise for some diseases. Here, we show that eliminating a common stop mutation associated with cystic fibrosis by inducing skipping of the exon it is located in, results in a restoration of the open reading frame and recovers CFTR protein function in a manner expected to be therapeutic in CF patients who dont currently have effective treatment options. These results are an important advancement for the cystic fibrosis community but also have implications for other diseases where terminating mutations are responsible for dysfunction.


title: Nuclear SUN1 stabilizes endothelial cell junctions to regulate blood vessel formation

abstract:
Endothelial cells line all blood vessels and coordinate blood vessel formation and the blood-tissue barrier via endothelial cell-cell junctions. The nucleus also regulates endothelial cell behaviors, but the mechanisms are poorly understood. Here we show that nuclear-localized SUN1, a LINC complex component that connects the nucleus to the cytoskeleton, regulates endothelial cell-cell junction communication and blood vessel formation. Loss of murine endothelial Sun1 impaired blood vessel formation and destabilized junctions. At the cellular level, SUN1 stabilized endothelial cell-cell junctions and promoted barrier function. Abnormal SUN1-depleted junctions resembled those seen with loss of microtubules, and they were accompanied by impaired microtubule dynamics and actomyosin hypercontractility. Angiogenic sprouts formed but retracted in SUN1-depleted endothelial cells, and vessels of zebrafish lacking SUN1 had abnormal extension and were defective in forming connections. Thus, endothelial SUN1 regulates peripheral cell-cell junctions from the nucleus, likely via microtubule-based interactions, and this long-range regulation is important for blood vessel formation and barrier function.

SUMMARYThe nuclear membrane protein SUN1 promotes blood vessel formation and barrier function by stabilizing endothelial cell-cell junctions. Communication between SUN1 and endothelial cell junctions relies upon microtubules, revealing long-range cellular communication that is important for vascular development and function.


title: Protein prediction for trait mapping in diverse populations

abstract:
Genetically regulated gene expression has helped elucidate the biological mechanisms underlying complex traits. Improved high-throughput technology allows similar interrogation of the genetically regulated proteome for understanding complex trait mechanisms. Here, we used the Trans-omics for Precision Medicine (TOPMed) Multi-omics pilot study, which comprises data from Multi-Ethnic Study of Atherosclerosis (MESA), to optimize genetic predictors of the plasma proteome for genetically regulated proteome-wide association studies (PWAS) in diverse populations. We built predictive models for protein abundances using data collected in TOPMed MESA, for which we have measured 1,305 proteins by a SOMAscan assay. We compared predictive models built via elastic net regression to models integrating posterior inclusion probabilities estimated by fine-mapping SNPs prior to elastic net. In order to investigate the transferability of predictive models across ancestries, we built protein prediction models in all four of the TOPMed MESA populations, African American (n=183), Chinese (n=71), European (n=416), and Hispanic/Latino (n=301), as well as in all populations combined. As expected, fine-mapping produced more significant protein prediction models, especially in African ancestries populations, potentially increasing opportunity for discovery. When we tested our TOPMed MESA models in the independent European INTERVAL study, fine-mapping improved cross-ancestries prediction for some proteins. Using GWAS summary statistics from the Population Architecture using Genomics and Epidemiology (PAGE) study, which comprises ~50,000 Hispanic/Latinos, African Americans, Asians, Native Hawaiians, and Native Americans, we applied S-PrediXcan to perform PWAS for 28 complex traits. The most protein-trait associations were discovered, colocalized, and replicated in large independent GWAS using proteome prediction model training populations with similar ancestries to PAGE. At current training population sample sizes, performance between baseline and fine-mapped protein prediction models in PWAS was similar, highlighting the utility of elastic net. Our predictive models in diverse populations are publicly available for use in proteome mapping methods at https://doi.org/10.5281/zenodo.4837328.

Author summaryGene regulation is a critical mechanism underlying complex traits. Transcriptome-wide association studies (TWAS) have helped elucidate potential mechanisms because each association connects a gene rather than a variant to the complex trait. Like genome-wide association studies (GWAS), most TWAS are still conducted exclusively in populations of European ancestry, which misses the opportunity to test the full spectrum of human genetic variation for associations with complex traits. Here, move beyond the transcriptome and because protein measurement assays are growing to allow interrogation of the proteome, we use data from TOPMed MESA to develop genetic predictors of protein abundance in diverse ancestry populations. We compare model-building strategies with the goal of providing the best resource for protein association discovery with available data. We demonstrate how these prediction models can be used to perform proteome-wide association studies (PWAS) in diverse populations. We show the most protein-trait associations were discovered, colocalized, and replicated in independent cohorts using proteome prediction model training populations with similar ancestries to individuals in the GWAS. We shared our protein prediction models and performance statistics publicly to facilitate future proteome mapping studies in diverse populations.

category: genomics


title: Medial temporal and prefrontal cortices encode structural task representations at different levels of abstraction

abstract: Memory generalisations may be underpinned by either encoding- or retrieval-based generalisation mechanisms and different training schedules may bias some learners to favour one of these mechanisms over the other. We used a transitive inference task to investigate whether generalisation is influenced by progressive vs randomly interleaved training, and overnight consolidation. On consecutive days, participants learnt pairwise discriminations from two transitive hierarchies before being tested during fMRI. Inference performance was consistently better following progressive training, and for pairs further apart in the transitive hierarchy. BOLD pattern similarity correlated with hierarchical distances in the left hippocampus (HIP) and medial prefrontal cortex (MPFC) following both training schedules. These results are consistent with the use of structural representations that directly encode hierarchical relationships between task features. However, such effects were only observed in the MPFC for recently learnt relationships and were strongest in participants who were least proficient at the inference task. We conclude that humans preferentially employ encoding-based mechanisms to store map-like relational codes that can be used for memory generalisation. These codes are expressed in the HIP and MPFC following both progressive and interleaved training but are not sufficient for accurate inference.


title: Hippocampal and medial prefrontal cortices encode structural task representations following progressive and interleaved training schedules


title: Analysis of combinatorial chemokine receptor expression dynamics using multi-receptor reporter mice

abstract: Inflammatory chemokines and their receptors are central to the development of inflammatory/immune pathologies. The apparent complexity of this system, coupled with lack of appropriate in vivo models, has limited our understanding of how chemokines orchestrate inflammatory responses and has hampered attempts at targeting this system in inflammatory disease. Novel approaches are therefore needed to provide crucial biological, and therapeutic, insights into the chemokine-chemokine receptor family. Here, we report the generation of transgenic multi-chemokine receptor reporter mice in which spectrally-distinct fluorescent reporters mark expression of CCRs 1, 2, 3 and 5, key receptors for myeloid cell recruitment in inflammation. Analysis of these animals has allowed us to define, for the first time, individual and combinatorial receptor expression patterns on myeloid cells in resting and inflamed conditions. Our results demonstrate that chemokine receptor expression is highly specific, and more selective than previously anticipated.


title: Organization and dynamics of the cortical complexes controlling insulin secretion in β-cells.

abstract: Insulin secretion in pancreatic {beta}-cells is regulated by cortical complexes that are enriched at the sites of adhesion to extracellular matrix facing the vasculature. Many components of these complexes, including Bassoon, RIM, ELKS and liprins, are shared with neuronal synapses. Here, we show that insulin secretion sites also contain non-neuronal proteins LL5{beta} and KANK1, which in migrating cells organize exocytotic machinery in the vicinity of integrin-based adhesions. Depletion of LL5{beta} or focal adhesion disassembly triggered by myosin II inhibition perturbed the clustering of secretory complexes and attenuated the first wave of insulin release. While previous analyses in vitro and in neurons suggested that secretory machinery might assemble through liquid-liquid phase separation, analysis of endogenously labeled ELKS in pancreatic islets indicated that its dynamics is inconsistent with such a scenario. Instead, fluorescence recovery after photobleaching and single molecule imaging showed that ELKS turnover is driven by binding and unbinding to low-mobility scaffolds. Both the scaffold movements and ELKS exchange were stimulated by glucose treatment. Our findings help to explain how integrin-based adhesions control spatial organization of glucose-stimulated insulin release.


title: Decellularized micro-scaffolds as stem cell carriers for regeneration of cartilage defects

abstract: Cartilage regeneration remains a great challenge in orthopedic treatment owing to their avascular in nature and lack of self-healing ability. Various clinical treatment options are widely used including osteochondral graft transplantation, micro-fracture, blood clot formation and tissue debridement to recover the damaged cartilage. In this circumstance, cartilage defect recovery via tissue engineered functional micro tissue delivery is becoming an emerging trend in musculoskeletal therapeutics. In this study, functional micro-scaffolds (MS) were generated from Capra ear cartilage, and were separated into size-wise groups. The scaffolds were decellularized via NaOH treatment. The cell adhesion study indicated that Capra adipose tissue derived mesenchymal stem cells (ADMSCs) adhesion is more in ~100 m MSs in comparison to 150-300 m MSs. It may be assumed that the cells are compatible to grow on fibrous surface area (100 m) in comparison to dense surface (150-300 m). Further, 100 m MSs were transformed into functional micro tissues (FMTs) in presence of high density ADMSCs in a hanging droplet culture system. The FMTs was transferred to F127 block polymer hydrogel for 3D culture. After 21 d cultures, the FMT clusters were evaluated for quantitative gene expression. To assess the in vivo cartilage defect regeneration potential, FMTs were delivered to rabbit auricular cartilage defect for 15, 30 and 60 d studies. The H&E-stained histological analysis showed that the cartilage defect is almost healed in 60 d study in comparison to 15 and 30 d study.

category: bioengineering


title: Size dependent regeneration capacity of functionalized Capra ear derived micro-tissue scaffolds for treatment of cartilage defects


title: ELICE16INDURES (R): a plant immune-priming activator targeting jasmonate metabolism via TIFY and RHOMBOID proteins in Hordeum vulgare

abstract:
Priming activity of plant-based allelochemicals is advanced research nowadays meaning a high potential in sustainable agriculture. The ELICE16INDURES(R) (RIMPH LTD, Hungary) plant conditioner of CO2 botanical extracts is rich in plant-active ingredients such as phenolic compounds, alkaloids, and flavonoids formulated in small multilamellar liposomes. This product was investigated in autumn barley (Hordeum vulgare). Field experiments of ELICE16INDURES showed augmented NDVI values interconnected with higher photosynthetic activity and yield increase. Background of the better vitality of plants was investigated by whole genomic gene expression profiling and showed an enhanced response to wounding, jasmonic acid, oxidative detoxification, and chloroplast activity. Among top50 differentially expressed genes the TIFY domain protein TIFY11B and RHOMBOID-like protein 2 related to JA signaling were up-regulated in field-collected samples. Phytotron experiments of barley were set up to validate and evaluate the transcriptomic effect of ELICE16INDURES. Well-studied priming active agents such as salicylic acid and beta-aminobutyric acid were compared with ELICE16INDURES and confirmed as priming inducer material with positive regulation of TIFY11B, TIFY3B, TIFY9, TIF10A, and RHOMBOID like protein 2 by using NGS GEx and RT-qPCR methods.

One-sentence summaryELICE16INDURES(R) is a plant conditioner agent with a high amount of allelochemicals encapsulated into small multilamellar liposomes and found as an immune priming activator tested in H. vulgare field and phytotron cultures.

category: plant biology


title: MRG proteins are shared by multiple protein complexes with distinct functions

abstract:
MRG15/MORF4L1 is a highly conserved protein in eukaryotes that contains a chromodomain recognizing H3K36me3 in chromatin. Intriguingly, it has been reported in the literature to interact with several different factors involved in chromatin modifications, gene regulation, alternative mRNA splicing and DNA repair by homologous recombination. In order to get a complete and reliable picture of associations in physiological conditions, we used genome editing and tandem affinity purification to analyze the stable native interactome of human MRG15, its paralog MRGX/MORF4L2 that lacks the chromodomain, and MRGBP (MRG-binding protein) in isogenic K562 cells. We found stable interchangeable association of MRG15 and MRGX with the NuA4/TIP60 histone acetyltransferase/chromatin remodeler, Sin3B histone deacetylase/demethylase, ASH1L histone methyltransferase and PALB2/BRCA2 DNA repair protein complexes. These associations were further confirmed and analyzed by CRISPR-tagging of endogenous proteins and comparison of expressed isoforms. Importantly, based on structural information, point mutations could be introduced that can specifically disrupt MRG15 association with some complexes but not others. Most interestingly, we also identified a new abundant native complex formed by MRG15/X-MRGBP-BRD8-EP400NL that is functionally similar to the yeast TINTIN (Trimer Independent of NuA4 for Transcription Interactions with Nucleosomes) complex. Our results show that EP400NL, being homologous to the N-terminal region of NuA4/TIP60 subunit EP400, creates TINTIN by competing for BRD8 association. Functional genomics indicate that human TINTIN plays a role in transcription of specific genes. This is most likely linked to the H4ac-binding bromodomain of BRD8 along the H3K36me3-binding chromodomain of MRG15 on the coding region of transcribed genes. Taken together, our data provide a complete detailed picture of human MRG proteins-associated protein complexes which is essential to understand and correlate their diverse biological functions in chromatin-based nuclear processes.

HighlightsO_LIMRG15 and MRGX are stably associated with several different protein complexes important for genome expression and stability.
C_LIO_LISeveral MRG-containing complexes are chromatin modifiers.
C_LIO_LISpecific point mutations in the MRG domain differentially affect associated complexes.
C_LIO_LIA major human complex homologous to the yeast TINTIN complex is identified.
C_LIO_LIThe protein EP400NL competes with EP400 to functionally separate TINTIN from the NuA4/TIP60 complex.
C_LIO_LITINTIN contains a bromodomain and a chromodomain to regulate transcription.
C_LI

category: molecular biology


title: Performance of Core Genome Multilocus Sequence Typing Compared to Capillary-Electrophoresis PCR Ribotyping and SNP Analysis of Clostridioides difficile

abstract: Clostridioides difficile is the most common cause of antibiotic-associated gastrointestinal infections. Capillary-electrophoresis (CE)-PCR ribotyping is currently the gold standard for C. difficile typing but lacks discriminatory power to study transmission and outbreaks in detail. New molecular methods have the capacity to differentiate better, but backward compatibility with CE-PCR ribotyping must be assessed. Using a well-characterized collection of diverse strains (N=630; 100 unique ribotypes [RTs]), we aimed to investigate PCR ribotyping prediction from core genome multilocus sequence typing (cgMLST). Additionally, we compared the discriminatory power of cgMLST (SeqSphere & EnteroBase) and whole genome MLST (wgMLST) (EnteroBase) with single nucleotide polymorphism (SNP) analysis). A unique cgMLST profile (>6 allele differences) was observed in 82/100 ribotypes, indicating sufficient backward compatibility. Intra-RT allele difference varied per ribotype and MLST clade. Application of cg/wgMLST and SNP analysis in two outbreak settings with ribotypes RT078 and RT181 (known with a low intra-ribotype allele difference) showed no distinction between outbreak- and non-outbreak strains, in contrast to wgMLST and SNP analysis. We conclude that cgMLST has the potential to be an alternative to CE-PCR ribotyping. The method is reproducible, easy to standardize and offers higher discrimination. However, in some ribotype complexes adjusted cut-off thresholds and epidemiological data are necessary to recognize outbreaks. We propose to decrease the current threshold of 6 to 3 alleles to better identify outbreaks.


title: cDNA-detector: Detection and removal of cDNA contamination in DNA sequencing libraries

abstract: Exogenous cDNA introduced into an experimental system, either intentionally or accidentally, can appear as added read coverage over that gene in next-generation sequencing libraries derived from this system. If not properly recognized and managed, this cross-contamination with exogenous signal can lead to incorrect interpretation of research results. Yet, this problem is not routinely addressed in current sequence processing pipelines. Here, we present cDNA-detector, a computational tool to identify and remove exogenous cDNA contamination in DNA sequencing experiments. We apply cDNA-detector to several highly-cited public databases (TCGA, ENCODE, NCBI SRA) and show that contaminant genes appear in sequencing experiments where they lead to incorrect coverage peak calls. Our findings highlight the importance of sensitive detection and removal of contaminant cDNA from NGS libraries before downstream analysis.

category: bioinformatics


title: A gap-junction mutation in the mouse cochlea reveals cochlear amplification is driven by outer hair cell extracellular receptor potentials

abstract:
Cochlear amplification, whereby cochlear responses to low-to-moderate sound levels are amplified and compressed to loud sounds, is attributed to outer hair cell (OHC) electromotility driven by voltage changes across the OHC basolateral membranes due to sound-induced receptor-current modulation. Cochlear operation at high acoustic frequencies is enigmatic because the OHC intracellular receptor potential (RP) is severely attenuated at these frequencies. Clues to understanding the voltage control of OHC electromotility at different frequencies was provided by measurements from CD-1 mice with an A88V mutation of the gap-junction (GJ) protein connexin 30 (Cx30), which with Cx26, form heterogeneous GJs between supporting cells in the organ of Corti (OoC) and stria vascularis. The A88V mutation results in a smaller GJ conductance which may explain why the resistance across the OoC in CD-1Cx30A88V/A88V mutants is higher compared with wild-type mice. The endocochlear potential, which drives the OHC receptor current and, consequently, the OHC RPs, is smaller in CD-1Cx30A88V/A88V mutants. Even so, their high-frequency hearing sensitivity equals that of wild-type mice. Preservation of high-frequency hearing correlates with similar amplitude of extracellular receptor potentials (ERPs), measured immediately adjacent to the OHCs. ERPs are generated through OHC receptor current flow across the OoC electrical resistance, which is larger in CD-1Cx30A88V/A88V than in wild-type mice. Thus, smaller OHC receptor currents flowing across a larger OoC resistance in CD-1Cx30A88V/A88V mice may explain why their ERP magnitudes are similar to wild-type mice. It is proposed that the ERPs, which are not subject to low-pass electrical filtering, drive high-frequency cochlear amplification.

Significance StatementCochlear amplification, whereby responses to low-to-moderate sound levels are amplified and those to loud sounds are compressed, is attributed to outer hair cell (OHC) electromotility. Electromotility is driven by voltage changes across the OHC basolateral membranes due to modulation of receptor current flow during sound-induced sensory hair bundle displacement. Mechanisms of high-frequency cochlear amplification remain to be elucidated. A mutation of the gap-junction protein connexin 30 decreases OHC intracellular receptor potentials in CD-1 mice. Instead of decreasing auditory sensitivity, the mutation rescues high-frequency hearing by causing OHC extracellular receptor potentials to be similar in amplitude to those of sensitive wild-type mice. It is proposed extracellular, not intracellular, potentials drive high-frequency OHC motility and cochlear amplification at high acoustic frequencies.


title: Identification of Subpallial Neuronal Populations Across Zebrafish Larval Stages that Express Molecular Markers for the Striatum

abstract: Striatal neurons play a central role in vertebrate action selection; however, their location in larval zebrafish is not well defined. We assayed for conserved striatal markers in the zebrafish subpallium using fluorescent in situ hybridization (FISH) and immunohistochemistry. Whole mount FISH revealed an inhibitory neuronal cluster rostral to the anterior commissure that expresses tac1, a gene encoding substance P. This molecular profile is shared by mammalian striatal direct pathway neurons. A second partially overlapping population of inhibitory neurons was identified that expresses penka, a gene encoding enkephalin. This molecular profile is shared by striatal indirect pathway neurons. Immunostaining for substance P and enkephalin confirmed the presence of these peptides in the subpallium. The tac1 and penka populations were both found to increase linearly across larval stages. Together, these findings support the existence of a striatal homologue in larval zebrafish that grows to match the development and increasing behavioural complexity of the organism.


title: The spatio-temporal program of liver zonal regeneration

abstract: The liver carries a remarkable ability to regenerate rapidly after acute zonal damage. Single-cell approaches are necessary to study this process, given the spatial heterogeneity of multiple liver cell types. Here, we use spatially-resolved single cell RNA sequencing (scRNAseq) to study the dynamics of mouse liver regeneration after acute acetaminophen (APAP) intoxication. We find that hepatocytes proliferate throughout the liver lobule, creating the mitotic pressure required to repopulate the necrotic pericentral zone rapidly. A subset of hepatocytes located at the regenerating front transiently up-regulate fetal-specific genes, including Afp and Cdh17, as they reprogram to a pericentral state. Zonated endothelial, hepatic-stellate cell (HSC) and macrophage populations are differentially involved in immune recruitment, proliferation and matrix remodeling. We observe massive transient infiltration of myeloid cells, yet stability of lymphoid cell abundance, in accordance with global decline in antigen presentation. Our study provides a resource for understanding the coordinated programs of zonal liver regeneration.


title: Sticky Pi, an AI-powered smart insect trap for community chronoecology

abstract: In the face of severe environmental crises that threaten insect biodiversity, new technologies are imperative to monitor both the identity and ecology of insect species. Traditionally, insect surveys rely on manual collection of traps, which provide abundance data but mask the large intra- and inter-day variations in insect activity, an important facet of their ecology. Although laboratory studies have shown that circadian processes are central to insects biological functions, from feeding to reproduction, we lack the high-frequency monitoring tools to study insect circadian biology in the field. To address these issues, we developed the Sticky Pi, a novel, autonomous, open-source, insect trap that acquires images of sticky cards every twenty minutes. Using custom deep-learning algorithms, we automatically and accurately scored where, when and which insects were captured. First, we validated our device in controlled laboratory conditions with a classic chronobiological model organism, Drosophila melanogaster. Then, we deployed an array of Sticky Pis to the field to characterise the daily activity of an agricultural pest, Drosophila suzukii, and its parasitoid wasps. Finally, we demonstrate the wide scope of our smart trap by describing the sympatric arrangement of insect temporal niches in a community, without targeting particular taxa a priori. Together, the automatic identification and high sampling rate of our tool provide biologists with unique data that impacts research far beyond chronobiology; with applications to biodiversity monitoring and pest control as well as fundamental implications for phenology, behavioural ecology, and ecophysiology. We released the Sticky Pi project as an open community resource on https://doc.sticky-pi.com.

category: ecology


title: The rate of ecological acclimation is the dominant uncertainty in long-term projections of a key ecosystem service

abstract: Rapid climate change may exceed ecosystems capacity to respond through processes including phenotypic plasticity, compositional turnover and evolutionary adaption. However, research predicting impacts of climate change on ecosystem services rarely consider this rate of "ecosystem acclimation." Combining statistical models fit to historical climate data and remotely-sensed estimates of herbaceous productivity with an ensemble of climate models, we demonstrate that assumptions concerning acclimation rates are a dominant source of uncertainty: models assuming minimal acclimation project widespread decreases in forage production in the western US by 2100, while models assuming that acclimation keeps pace with climate change project widespread forage increases. Uncertainty related to ecosystem acclimation is larger than uncertainties from variation among climate models or emissions pathways. A better understanding of ecosystem acclimation is essential to improve long-term forecasts of ecosystem services, and shows that management to facilitate ecosystem acclimation may be necessary to maintain ecosystem services at historical baselines.


title: The transmission of African Animal Trypanosomiasis (nagana) in two districts in the forest zone of Ghana

abstract: The African Animal Trypanosomiasis (nagana) is caused by several species of trypanosome species which results in significant clinical diseases. This consequently leads to huge economic losses. We carried out a cross-sectional survey to investigate the composition of vectors, parasite diversity and intensities in two districts in the Eastern region of Ghana. Cytochrome c oxidase subunit 1 (COI) and internal transcribed spacer (ITS1) dependent PCR assays identified tsetse fly species and trypanosome parasites circulating in the area. A total of 229 tsetse flies, 65 pigs and 20 were sampled. Female and male flies 155 (51.8%), 74 (32.3%) respectively were collected with biconical traps. The overall vector density of 4.3 flies/trap/day was observed. Typanosome prevalence of 58.9% (95% CI: 52.5-65.1), 46.2% (95% CI: 34.6-58.1) and 0.0% (95% CI: 0.0-16.1) in tsetse flies, pigs and cattle respectively were detected. Trypanosoma congolense was the predominant species with a prevalence of 80.7% (95% CI: 73.3-86.5) in flies and 60.0% (95% CI: 42.3-75.4). There was evidence of multiple trypanosome infection with T. congolense/T. simiae occurring highest with prevalence of 38.0% (95% CI: 30.7-46.9). The parasite prevalence in pigs across the communities was high with significant differences associated between locations (Chi2 = 28.06, 95% CI: 0.05-0.81, P=0.0009). Blood meal with flies revealed feeding on both domestic Sus scrofa domesticus (pig) and Phacochoerus africanus (warthog). Tsetse flies in this area remain highly infected with trypanosomes and continue to transmit the parasites to livestock and human populations in the communities.

category: evolutionary biology


title: The transmission of African Animal Trypanosomiasis (Nagana) in two districts in the forest zone of Ghana


title: Site Pattern Probabilities Under the Multispecies Coalescent and a Relaxed Molecular Clock: Theory and Applications

abstract: 1The first step in statistical inference of the evolutionary histories of species is developing a probability model that describes the mutation process as accurately and realistically as possible. A major complication of this inference is that different loci on the genome can have histories that diverge from the common species history and each other. The multispecies coalescent process is commonly used to model one source of this divergence, incomplete lineage sorting, or ILS. Chifman and Kubatko (2015) computed the site pattern probabilities for four taxa under a full probability model based on the Jukes-Cantor substitution model when the molecular clock holds. This paper generalizes that work to a relaxed clock model, allowing for mutation rates to differ among species. This will enable better phylogentic inference in cases where the molecular clock does not hold.


title: Decoding gene regulation in the fly brain

abstract: The Drosophila brain is a work horse in neuroscience. Single-cell transcriptome analysis 1-5, 3D morphological classification 6, and detailed EM mapping of the connectome 7-10 have revealed an immense diversity of neuronal and glial cell types that underlie the wide array of functional and behavioral traits in the fruit fly. The identities of these cell types are controlled by - still unknown - gene regulatory networks (GRNs), involving combinations of transcription factors that bind to genomic enhancers to regulate their target genes. To characterize the GRN for each cell type in the Drosophila brain, we profiled chromatin accessibility of 240,919 single cells spanning nine developmental timepoints, and integrated this data with single-cell transcriptomes. We identify more than 95,000 regulatory regions that are used in different neuronal cell types, of which around 70,000 are linked to specific developmental trajectories, involving neurogenesis, reprogramming and maturation. For 40 cell types, their uniquely accessible regions could be associated with their expressed transcription factors and downstream target genes, through a combination of motif discovery, network inference techniques, and deep learning. We illustrate how these "enhancer-GRNs" can be used to reveal enhancer architectures leading to a better understanding of neuronal regulatory diversity. Finally, our atlas of regulatory elements can be used to design genetic driver lines for specific cell types at specific timepoints, facilitating the characterization of brain cell types and the manipulation of brain function.


title: Predicting epitopes Based on TCR sequence using an embedding deep neural network artificial intelligence approach

abstract: T cells receptors are fundamental in recognizing antigens and mediating an appropriate specific immune response against them. Today TCR sequencing has contributed to forming a large repertoire for different immune-associated pathologies. However, predicting epitopes based on TCR sequences has not been satisfactory achieved. We formed a deep neural network using a combination of an embedding autoencoder and selu and relu layers to predict epitopes based on TCR TCR {beta}-chain CDR3. We trained our model using the VDJ database (VDJdb) and validated it using the manually curated catalog of pathology-associated T cell receptor sequences (McPAS-TCR). We used various metrics to measure the accuracy of our tool. We found that our tool can achieve an accuracy of 98 %. Overall our approach presents a step toward identifying microbes crosslinking epitopes that could be playing an important role in various immune diseases.


title: MODEL BASED OPTIMIZATION OF THE TERPENOIDS BIOSYNTHESIS IN E. coli

abstract:
Terpenoids are a family of compounds with high industrial interest and the development of biotechnological production methods is essential to achieve more sustainable alternatives to traditional extraction and synthesis methods. The modification and engineering of the catalytic activity (kcat) have been shown to be a feasible strategy in the biotechnological realm. Accordingly, we introduce a novel optimization strategy based in the modification of the kcat of the enzymes and applied it to the maximization of the terpenoids synthesis in E. coli. This approach is fairly general and can be applied alone or in conjunction with classic optimization strategies such as the modification of enzymatic specific activities.

For this purpose we first build up a reliable dynamic mathematical model of the alternative mevalonate pathway synthesis leading terpenoids biosynthesis in E. coli through the methyl-D-erythritol 4-phosphate (MEP) pathway. This model includes the 2-C-methyl-D-erythritol 2, 4-diphosphate (MEC) pumps that mediate MEC extracellular extrusion. Although the physiological significance of the MEC extrusion is still discussed and their biological function is not clear, we find that this process is a must to guarantee bacteria homeostasis and cell viability.

We have identified the enzyme IspA as a bottleneck of the terpenoids biosynthesis, which is dual substrate for the two bisubstrate final reactions. Here are presented different ways to overcome this enzymatic flux restriction by modification of the IspA kcat or by introduction of new enzymes with parallel function.

Our results show that the MEP pathway optimized solutions for kcat can yield a maximum of 17.68 fold increment in the terpenoids biosynthetic flux when the kcat are modified. This maximal solution involves the modification of 8 kcat and the corresponding Kms and Kds. Remarkably, this increase doesnt imply a change in the total enzyme concentration of the cell. This is favorable output since enzyme overproduction can compromise cell functionality.

We also apply the overexpression of the enzyme activity approach and then we have compared and combined both strategies including scenarios as the deletion or import of the genes expressing enzymes not naturally present in E. coli.

A combined strategy of enzymes concentrations and kcat modifications with changes up ti six enzyme levels allows a flux increment of 21.22 fold the basal value. It involves the incorporation of two IspA with similar GPP and IPP bisubstrate functions than the native one but low affinity for the DMAPP as substrate.

category: systems biology


title: Deep learning based behavioral profiling of rodent stroke recovery

abstract: Stroke research heavily relies on rodent behavior when assessing underlying disease mechanisms and treatment efficacy. Although functional motor recovery is considered the primary targeted outcome, tests in rodents are still poorly reproducible, and often unsuitable for unraveling the complex behavior after injury. Here, we provide a comprehensive 3D gait analysis of mice after focal cerebral ischemia based on the new deep learning-based software (DeepLabCut, DLC) that only requires basic behavioral equipment. We demonstrate a high precision 3D tracking of 10 body parts (including all relevant joints and reference landmarks) in several mouse strains with an accuracy of 99.4%. Building on this rigor motion tracking, a comprehensive post-analysis (with >100 parameters) unveils biologically relevant differences in locomotor profiles after a stroke over a time course of three weeks. We further refine the widely used ladder rung test using deep learning and compare its performance to human annotators. The generated DLC-assisted tests were then benchmarked to five widely used conventional behavioral set-ups (neurological scoring, rotarod, ladder rung walk, cylinder test, and single-pellet grasping) regarding sensitivity, accuracy, time use and costs. We conclude that deep learning-based motion tracking with comprehensive post-analysis provides accurate and sensitive data to describe the complex recovery of rodents following a stroke. The experimental set-up and analysis can also benefit a range of other neurological injuries that affect locomotion.


title: RNA sequencing on muscle biopsy from a 5-week bedrest study reveals the effect of exercise and potential interactions with dorsal root ganglion neurons

abstract: Lack of physical activity is a predictor of poor health outcomes that can be prevented or reversed by exercise. Sedentary lifestyle, chronic disease or microgravity can cause muscle deconditioning that then has an impact on other physiological systems. An example is the nervous system, which is adversely affected by decreased physical activity resulting in increased incidence of neurological problems such as chronic pain. We sought to better understand how this might occur by conducting RNA sequencing experiments on muscle biopsies from human volunteers in a 5-week bed-rest study with an exercise intervention arm. We also used a computational method for examining ligand-receptor interactions between muscle and human dorsal root ganglion (DRG) neurons, the latter of which play a key role in nociception and are generators of signals responsible for chronic pain. We identified 1352 differentially expressed genes (DEGs) in bed rest subjects without an exercise intervention but only 132 DEGs in subjects with the intervention. Thirty-six genes were shared between the exercise and no intervention groups. Among 591 upregulated muscle genes in the no intervention arm, 26 of these were ligands that have receptors that are expressed by human DRG neurons. We detected a specific splice variant of one of these ligands, placental growth factor (PGF), in deconditioned muscle that binds to neuropilin 1, a receptor that is highly expressed in DRG neurons and known to promote neuropathic pain. We conclude that exercise intervention protects muscle from deconditioning transcriptomic changes, and prevents changes in expression of ligands that might sensitize DRG neurons that promote pain. Our work creates a set of actionable hypotheses to better understand how deconditioned muscle may influence the function of sensory neurons that innervate the entire body.


title: Screening and identification of muscle-specific candidate genes via mouse microarray data analysis

abstract: Muscle tissue is involved with every stage of life activities and has roles in biological processes. For example, the blood circulation system needs the heart muscle to transport blood to all parts, and the movement cannot be separated from the participation of skeletal muscle. However, the process of muscle development and the regulatory mechanisms of muscle development are not clear at present. In this study, we used bioinformatics techniques to identify differentially expressed genes specifically expressed in multiple muscle tissues of mice as potential candidate genes for studying the regulatory mechanisms of muscle development. Mouse tissue microarray data from 17 tissue samples was selected from the GEO database for analysis. Muscle tissue as the treatment group, and the other 16 tissues as the control group. Genes expressed in the muscle tissue were different to those in the other 16 tissues and identified 272 differential genes with highly specific expression in muscle tissue, including 260 up-regulated genes and 12 down regulated genes. is the genes were associated with the myofibril, contractile fibers, and sarcomere, cytoskeletal protein binding, and actin binding. KEGG pathway analysis showed that the differentially expressed genes in muscle tissue were mainly concentrated in pathways for AMPK signaling, cGMP PKG signaling calcium signaling, glycolysis, and, arginine and proline metabolism. A PPI protein interaction network was constructed for the selected differential genes, and the MCODE module used for modular analysis. Five modules with Score > 3.0 are selected. Then the Cytoscape software was used to analyze the tissue specificity of differential genes, and the genes with high degree scores collected, and some common genes selected for quantitative PCR verification. The conclusion is that we have screened the differentially expressed gene set specific to mouse muscle to provide potential candidate genes for the study of the important mechanisms of muscle development.


title: Genome scale CRISPR Cas9a knockout screen reveals genes that controls glioblastoma susceptibility to the alkylating agent temozolomide

abstract:
Glioblastoma is the most fatal of all primary human brain tumors with 14 months survival, at best. The mainstay therapy for this tumor involves temozolomide, surgery, radiotherapy and tumor treating electric field. Cancer resistance to commonly available chemotherapeutics remains a major challenge in glioblastoma patients receiving treatment and unfavorably impact their overall survival and outcome. However, the lack of progress in this area could be attributed to lack of tools to probe unbiasedly at the genome wide level the coding and non-coding elements contribution on a large scale for factors that control resistance to chemotherapeutics. Understanding the mechanisms of resistance to chemotherapeutics will enable precision medicine in the treatment of cancer patients.

CRISPR Cas9a has emerged as a functional genomics tool to study at genome level the factors that control cancer resistance to drugs. Recently, we used genome wide CRISPR-Cas9a screen to identify genes responsible for glioblastoma susceptibility to etoposide. We extended our inquiry to understand genes that control glioblastoma response to temozolomide by using genome scale CRISPR. This study shows that the unbiased genome-wide loss of function approach can be applied to discover genes that influence tumor resistance to chemotherapeutics and contribute to chemoresistance in glioblastoma.

category: cancer biology


title: Kindlin-2 preserves integrity of the articular cartilage to protect against osteoarthritis

abstract: Osteoarthritis (OA) is an aging-related degenerative joint disease, which has no cure partly due to limited understanding of its pathological mechanism(s). Here we report that the focal adhesion protein Kindlin-2, but not Kindlin-1 or -3, is highly expressed in articular chondrocytes of the hyaline cartilage, which is dramatically decreased in the degenerated articular cartilage of aged mice and patients with OA. Inducible deletion of Kindlin-2 in chondrocytes at adult stage leads to spontaneous OA and much severe OA lesions in the mice receiving the surgery of destabilization of the medial meniscus. Mechanistically, Kindlin-2 deficiency promotes mitochondrial oxidative stress and activates Stat3 in articular chondrocytes, leading to Runx2-mediated chondrocyte hypertrophic differentiation and catabolism. In vivo, systemic pharmacological blockade of Stat3 activation or genetic ablation of Stat3 in chondrocytes reverses aberrant accumulation of Runx2 and ECM-degrading enzymes and limits OA deteriorations caused by Kindlin-2 deficiency. Furthermore, genetic inactivation of Runx2 in chondrocytes reverses structural changes and OA lesions caused by Kindlin-2 deletion without down-regulating p-Stat3 in articular chondrocytes. Of translational significance, intraarticular injection of Kindlin-2-expressing adeno-associated virus decelerates progression of aging- and instability-induced knee joint OA in mice. Collectively, we identify a novel pathway comprising of Kindlin-2, Stat3 and Runx2 in articular chondrocytes responsible for maintaining integrity of the articular cartilage and define a potential therapeutic target for OA.


title: A NOVEL PREGNANT RAT MODEL FOR LABOR INDUCTION AND AUGMENTATION WITH OXYTOCIN

abstract:
BackgroundDespite the widespread use of oxytocin for induction of labor, mechanistic insights into maternal and neonatal wellbeing are lacking because of the absence of an animal model that recapitulates modern obstetric practice.

ObjectiveThe objectives of this research were to create and validate a hi-fidelity animal model that mirrors labor induction with oxytocin in parturients and to assess its translational utility.

Study DesignThe study was performed in timed-pregnant Sprague Dawley dams. The model consisted of a subcutaneously implanted microprocessor-controlled infusion pump on gestational day 18 that was pre-programmed to deliver an escalating dose of intravenous oxytocin on gestational day 21 to induce birth. Once predictable delivery of healthy pups was achieved, we validated the model with molecular biological experiments on the uterine myometrium and telemetry-supported assessment of changes in intrauterine pressure. Finally, we applied this model to test the hypothesis that labor induction with oxytocin was associated with oxidative stress in the newborn brain with a comprehensive array of biomarker assays and oxidative stress gene expression studies.

ResultsDuring the iterative model development phase, we confirmed the optimal gestational age for pump implantation, the concentration of oxytocin, and the rate of oxytocin administration. Exposure to anesthesia and surgery during pump implantation was not associated with significant changes in the cortical transcriptome. Activation of pump with oxytocin on gestational day 21 resulted in predictable delivery of pups within 8-12 hours. Increased frequency of change of oxytocin infusion rate was associated with dystocic labor. Labor induction and augmentation with oxytocin was associated with increased expression of the oxytocin receptor gene in the uterine myometrium, decreased expression of the oxytocin receptor protein on the myometrial cell membrane, and cyclical increases in intrauterine pressure. Examination of the frontal cortex of vaginally delivered newborn pups born after oxytocin-induced labor did not reveal an increase in oxidative stress compared to saline-treated control pups. Specifically, there were no significant changes in oxidative stress biomarkers involving both the oxidative stress (reactive oxygen/nitrogen species, 4-hydroxynonenal, protein carbonyl) and the antioxidant response (total glutathione, total antioxidant capacity). In addition, there were no significant differences in the expression of 16 genes emblematic of the oxidative stress response pathway.

ConclusionsCollectively, we provide a viable and realistic animal model for labor induction and augmentation with oxytocin. We demonstrate its utility in addressing clinically relevant questions in obstetric practice that could not be mechanistically ascertained otherwise. Based on our findings, labor induction with oxytocin is not likely to cause oxidative stress in the fetal brain. Adoption of our model by other researchers would enable new lines of investigation related to the impact of perinatal oxytocin exposure on the mother-infant dyad.

category: pharmacology and toxicology


title: Physiologically based modeling of the effect of physiological and anthropometric variability on indocyanine green based liver function tests

abstract: Accurate evaluation of liver function is a central task in hepatology. Dynamic liver function tests (DLFT) based on the time-dependent elimination of a test substance provide an important tool for such a functional assessment. These tests are used in the diagnosis and monitoring of liver disease as well as in the planning of hepatobiliary surgery. A key challenge in the evaluation of liver function with DLFTs is the large inter-individual variability. Indocyanine green (ICG) is a widely applied test compound used for the evaluation of liver function. After an intravenous administration, pharmacokinetic (PK) parameters are calculated from the plasma disappearance curve of ICG which provide an estimate of liver function. The hepatic elimination of ICG is affected by physiological factors such as hepatic blood flow or binding of ICG to plasma proteins, anthropometric factors such as body weight, age, and sex, or the protein amount of the organic anion-transporting polypeptide 1B3 (OATP1B3) mediating the hepatic uptake of ICG. Being able to account for and better understand these various sources of inter-individual variability would allow to improve the power of ICG based DLFTs and move towards an individualized evaluation of liver function. Within this work we systematically analyzed the effect of various factors on ICG elimination by the means of computational modeling. For the analysis, a recently developed and validated physiologically based pharmacokinetics (PBPK) model of ICG distribution and hepatic elimination was utilized. Key results are (i) a systematic analysis of the variability in ICG elimination due to hepatic blood flow, cardiac output, OATP1B3 abundance, liver volume, body weight and plasma bilirubin level; (ii) the evaluation of the inter-individual variability in ICG elimination via a large in silico cohort of n=100000 subjects based on the NHANES cohort with special focus on stratification by age, sex, and body weight; (iii) the evaluation of the effect of various degrees of cirrhosis on variability in ICG elimination. The presented results are an important step towards individualizing liver function tests by elucidating the effects of confounding physiological and anthropometric parameters in the evaluation of liver function via ICG.


title: Androgen synthesis inhibition but not gonadectomy reduces persistence during strategy set-shifting and reversal learning in male rats

abstract: Androgens regulate behavioural flexibility, which is essential to adapt to a changing environment and depends on the medial prefrontal cortex (mPFC). Testosterone (T) administration decreases behavioural flexibility. It is well known that T is produced in the gonads, but T is also produced in the mesocorticolimbic system, which modulates behavioural flexibility. It is unclear how T produced in the brain versus the gonads influences behavioural flexibility. Here, we assess the effects of the androgen synthesis inhibitor abiraterone acetate (ABI) and long-term gonadectomy (GDX) on behavioural flexibility in two paradigms. In Experiment 1, ABI independent of GDX reduced the number of trials to criterion and perseverative errors in a strategy set-shifting task. Similarly, in Experiment 2, ABI but not GDX reduced perseverative errors in a reversal learning task. In subjects from Experiment 1, we also examined tyrosine hydroxylase immunoreactivity (TH-ir), and ABI but not GDX increased TH-ir in the mPFC. Our findings suggest that neurally-produced androgens modulate behavioural flexibility via modification of dopamine signalling in the mesocorticolimbic system. These results suggest novel roles for neurosteroids and possible side effects of ABI treatment for prostate cancer.


title: The Proteomic Architecture of Schizophrenia Cerebral Organoids Reveals Alterations in GWAS and Neuronal Development Factors

abstract: Schizophrenia (Scz) is a brain disorder that has a typical onset in early adulthood but otherwise maintains unknown disease origins. Unfortunately, little progress has been made in understanding the molecular mechanisms underlying neurodevelopment of Scz due to ethical and technical limitations in accessing developing human brain tissue. To overcome this challenge, we have previously utilized patient-derived Induced Pluripotent Stem Cells (iPSCs) to generate self-developing, self-maturating, and self-organizing 3D brain-like tissue known as cerebral organoids. As a continuation of this prior work [1], here we provide a molecular architectural map of the developing Scz organoid proteome. Utilizing iPSCs from n = 25 human donors (n = 8 healthy Ctrl donors, and n = 17 Scz patients), we generated 3D human cerebral organoids, employed 16-plex isobaric sample-barcoding chemistry, and simultaneously subjected samples to comprehensive high-throughput liquid-chromatography/mass-spectrometry (LC/MS) quantitative proteomics. Of 3,705 proteins identified by high-throughput proteomic profiling, we identified that just ~2.62% of the organoid global proteomic landscape was differentially regulated in Scz organoids. In sum, just 43 proteins were up-regulated and 54 were down-regulated in Scz patient-derived organoids. Notably, a range of neuronal factors were depleted in Scz organoids (e.g., MAP2, TUBB3, SV2A, GAP43, CRABP1, NCAM1 etc.). Based on global enrichment analysis, alterations in key pathways that regulate nervous system development (e.g., axonogenesis, axon development, axon guidance, morphogenesis pathways regulating neuronal differentiation, as well as substantia nigra development) were perturbed in Scz patient-derived organoids. We also identified prominent alterations in two novel GWAS factors, Pleiotrophin (PTN) and Podocalyxin (PODXL), in Scz organoids. In sum, this work serves as both a report and a resource whereby researchers can leverage human-derived neurodevelopmental data from Scz patients, which can be used to mine, compare, contrast, or orthogonally validate novel factors and pathways related to Scz risk identified in datasets from observational clinical studies and other model systems.


title: Orthogonal representation of task-related information in theta phase-based multiple place fields of single units in the subiculum

abstract: The subiculum is positioned at a critical juncture at the interface of the hippocampus with the rest of the brain. However, the exact roles of the subiculum in most hippocampal-dependent memory tasks remain largely unknown. One obstacle to make analytical comparisons of neural firing patterns between the subiculum and hippocampal CA1 is the broad firing fields of the subicular cells. Here, we used spiking phases in relation to theta rhythm to parse the broad firing field of a subicular neuron into multiple subfields to find the unique functional contribution of the subiculum while male rats performed a hippocampal-dependent visual scene memory task. Some of the broad firing fields of the subicular neurons were successfully divided into multiple subfields by using the theta-phase precession cycle. The resulting phase-based fields in the subiculum were more similar to those in CA1 in terms of the field size and phase-precession strength. The new method significantly improved the detection of task-relevant information in subicular cells without affecting the information content represented by CA1 cells. Notably, multiple fields of a single subicular neuron, unlike those in the CA1, could carry heterogeneous task-related information such as visual context and choice response. Our findings suggest that the subicular cells integrate multiple task-related factors by using theta rhythm to associate environmental context with action.


title: Iba1+ cell density and morphology classification at postnatal day 10 in four hippocampal subregions of female and male C57BL/6J mice

abstract: Microglia maintain normal brain function and support the brains response to disease and injury. The hippocampus is an area of focus for microglial study due to its central role in numerous behavioral and cognitive functions. Interestingly, microglia and related cells in the hippocampus and throughout the brain are distinct in male vs. female rodents, even in early life. Indeed, postnatal day (P)-dependent sex differences in number, density, and morphology of microglia-like cells have been reported in certain hippocampal subregions. For example, P3 female mice have more phagocytic microglia in dentate gyrus (DG) molecular layer (Mol) and CA1-3 stratum oriens (SO) regions vs. male mice, while P8 -- but not P15 -- male rats have more volume immunoreactive for markers of microglia-like cells (Iba1 and CD68) in the CA1 stratum radiatum (SR) vs. female rats. In the mouse, P10 is roughly equivalent to human term gestation, making it a common timepoint to study for many translationally-relevant neurobiological processes. However, sex differences in hippocampal microglia have not been examined in the P10 mouse hippocampus. In addition, key subregions of the hippocampus -- CA3 SR, DG hilus -- have not yet been assessed for sex differences in microglia. To address these knowledge gaps, we quantified Iba1+ cell densities and classified Iba1+ cell morphology in P10 male and female C57BL/6J mice. Four subregions in the bilateral anterior hippocampus were analyzed in 40-micrometer coronal sections: DG Mol (Mol), DG Hilus, CA1 SR and stratum lacunosum moleculare (CA1), and CA3 SR and stratum lucidum (CA3). Light microscope images (40x) were analyzed offline for Iba1+ cell density and morphology by an observer blind to sex. The morphology of each Iba1+ cell was used to place cells into one of four previously-published categories: Round or ameboid (round-ish soma, no processes), Stout (round-ish soma, short process), Thick (irregular soma with few, thick processes), or Thin (irregular soma with multiple thin processes). Analysis of Iba1+ cell density shows no difference between male and female mice in Mol, Hilus, CA3, or CA1 (male n=6, female n=7). However, morphology classification shows a sex-dependent difference in the Mol and Hilus, with female mice having a greater percentage of Thick Iba1+ cells vs. male mice (Mol, Hilus), and a lower percentage of Thin Iba1+ cells vs. male mice (Mol). With our analysis, it is unclear whether this greater percentage of thick and lower percentage of thin Iba1+ cells in the female vs. male hippocampus means Iba1+ microglia in female mice are  younger or  more active than those in male mice. However, these data are important as they reveal sex differences in Iba1+ microglia in the P10 mouse hippocampus. We discuss these results in the context of the large literature on sex differences in rodent microglia in the early postnatal period.


title: Sulfonylureas target the neurovascular response to decrease Alzheimer's pathology

abstract:
Hyperexcitability is a defining feature of Alzheimers disease (AD), where aberrant neuronal activity is both a cause and consequence of AD. Therefore, identifying novel targets that modulate cellular excitability is an important strategy for treating AD. ATP-sensitive potassium (KATP) channels are metabolic sensors that modulate cellular excitability. Sulfonylureas are KATP channel antagonists traditionally used to combat hyperglycemia in diabetic patients by inhibiting pancreatic KATP channels, thereby stimulating insulin release. However, KATP channels are not limited to the pancreas and systemic modulation of KATP channels has pleotropic physiological effects, including profound effects on vascular function. Here, we demonstrate that human AD patients have higher cortical expression of vascular KATP channels, important modulators of vasoreactivity. We demonstrate that peripheral treatment with the sulfonylurea and KATP channel inhibitor, glyburide, reduced the aggregation and activity-dependent production of amyloid-beta (A{beta}), a hallmark of AD, in mice. Since glyburide does not readily cross the blood brain barrier, our data suggests that glyburide targets vascular KATP channel activity to reduce arterial stiffness, improve vasoreactivity, and normalize pericyte-endothelial cell morphology, offering a novel therapeutic target for AD.

Graphical abstractTargeting vascular KATP channel activity for the treatment of Alzheimers disease pathology.

O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY


title: The absence of the autoimmune regulator gene (AIRE) impairs the three-dimensional structure of medullary thymic epithelial cell spheroids

abstract: Besides controlling the expression of peripheral tissue antigens, the autoimmune regulator (AIRE) gene also regulates the expression of adhesion genes in medullary thymic epithelial cells (mTECs), an essential process for mTEC-thymocyte interaction for triggering the negative selection in the thymus. For these processes to occur, it is necessary that the medulla compartment forms an adequate three-dimensional (3D) architecture, preserving the thymic medulla. Previous studies have shown that AIRE knockout (KO) mice have a small and disorganized thymic medulla; however, whether Aire influences the mTEC-mTEC interaction in the maintenance of the 3D structure has been little explored. Considering that AIRE controls cell adhesion genes, we hypothesized that this gene affects 3D mTEC-mTEC interaction. To test this, we constructed an in vitro model system for mTEC spheroid formation, in which cells adhere to each other, establishing a 3D structure. The effect of Aire on mTEC-mTEC adhesion was evaluated by comparing AIRE wild type (AIREWT) versus Aire KO (AIRE-/-) mTECs. Considering the 3D spheroid model evaluated, we reported that the absence of AIRE disorganizes the 3D structure of mTEC spheroids, promotes a differential regulation of mTEC classical surface markers, and modulates genes encoding adhesion and other molecules.


title: Independently paced calcium oscillations in progenitor and differentiated cells in an ex vivo epithelial organ

abstract: Cytosolic calcium is a highly dynamic, tightly regulated, and broadly conserved cellular signal. Calcium dynamics have been studied widely in cellular monocultures, yet in vivo most organs comprise heterogeneous populations of stem and differentiated cells. We examined calcium dynamics in each cell type of the adult Drosophila intestine, a self-renewing epithelial organ where multipotent stem cells give rise to mature absorptive enterocytes and secretory enteroendocrine cells. Here we perform live imaging of whole organs ex vivo, and we employ orthogonal expression of red and green calcium sensors to determine whether calcium oscillations between different cell types are coupled. We show that stem cell daughters adopt strikingly distinct patterns of calcium oscillations when they acquire their terminal fates: Enteroendocrine cells exhibit single-cell calcium oscillations, while enterocytes exhibit rhythmic, long-range calcium waves. These multicellular waves do not propagate through progenitor cells (stem cells and enteroblasts), whose oscillation frequency is approximately half that of enteroendocrine cells. Organ-scale inhibition of gap junctions eliminates calcium oscillations in all three cell types, even, intriguingly, in progenitor and enteroendocrine cells that are surrounded only by enterocytes. Our findings establish that cells adopt fate-specific modes of calcium dynamics as they terminally differentiate and reveal that the oscillatory dynamics of different cell types in a single, coherent epithelium are paced independently.


title: Live imaging YAP signaling in vivo with fusion reporter mice

abstract: YAP protein is a critical regulator of mammalian embryonic development. By generating a near-infrared fusion YAP reporter mouse line, we have achieved high-resolution live imaging of YAP localization during mouse embryonic development. We have validated the reporter by demonstrating its predicted responses to blocking LATs kinase activity or blocking cell polarity. By time lapse imaging preimplantation embryos, we revealed a mitotic reset behavior of YAP nuclear localization. We also demonstrated deep tissue live imaging in post-implantation embryos and revealed an intriguing nuclear YAP pattern in migrating cells. The YAP fusion reporter mice and imaging methods will open new opportunities for understanding dynamic YAP signaling in vivo in many different situations.

category: developmental biology


title: Live imaging YAP signaling in mouse embryo development


title: Maternal and paternal sugar consumption interact to modify offspring life history and physiology

abstract: Intergenerational effects on offspring phenotypes occur in response to variation in both maternal and paternal nutrition. Because the combined maternal and paternal effects are rarely considered together however, their relative contributions, and the capacity for interactions between parental diets to shape offspring life history and physiology are not understood. To address this, we altered sucrose levels of adult fruit flies (Drosophila melanogaster) prior to mating, across two generations, producing parent-parent and parent-offspring combinations that were either matched or mismatched in dietary sucrose. We then measured lifespan, fecundity, body mass, and triglyceride levels in parents and offspring. We reveal complex non-additive interactions, that involve diets of each parent and offspring to shape offspring phenotypes, but the effects were generally not consistent with an adaptive response to parental diet. Notably, we find that interacting parental flies (sires and dams) lived longer when their sucrose treatments were matched, but they produced shorter-lived offspring.


title: A reference genome sequence resource for the sugar beet root rot pathogen Aphanomyces cochlioides

abstract: Aphanomyces cochlioides, the causal agent of damping-off and root rot of sugar beet (Beta vulgaris L.), is a soil-dwelling oomycete responsible for yield losses in all major sugar beet growing regions. Currently, genomic resources for A. cochlioides are limited. Here we report a de novo genome assembly using a combination of long-read MinION (Oxford Nanopore Technologies) and short-read Illumina sequence data for A. cochlioides isolate 103-1, from Breckenridge, MN. The assembled genome was 76.3 Mb, with a contig N50 of 2.6 Mb. The reference assembly was annotated and was composed of 32.1% repetitive elements and 20,274 gene models. This high-quality genome assembly of A. cochlioides will be a valuable resource for understanding genetic variation, virulence factors, and comparative genomics of this important sugar beet pathogen.


title: Genome-guided discovery of natural products through multiplexed low coverage whole-genome sequencing of soil Actinomycetes on Oxford Nanopore Flongle

abstract:
Genome-mining is an important tool for discovery of new natural products; however, the number of publicly available genomes for natural product-rich microbes such as Actinomycetes, relative to human pathogens with smaller genomes, is small. To obtain contiguous DNA assemblies and identify large (ca. 10 to greater than 100 Kb) biosynthetic gene clusters (BGCs) with high-GC (>70%) and -repeat content, it is necessary to use long-read sequencing methods when sequencing Actinomycete genomes. One of the hurdles to long-read sequencing is the higher cost.

In the current study, we assessed Flongle, a recently launched platform by Oxford Nanopore Technologies, as a low-cost DNA sequencing option to obtain contiguous DNA assemblies and analyze BGCs. To make the workflow more cost-effective, we multiplexed up to four samples in a single Flongle sequencing experiment while expecting low-sequencing coverage per sample. We hypothesized that contiguous DNA assemblies might enable analysis of BGCs even at low sequencing depth. To assess the value of these assemblies, we collected high-resolution mass-spectrometry data and conducted a multi-omics analysis to connect BGCs to secondary metabolites.

In total, we assembled genomes for 20 distinct strains across seven sequencing experiments. In each experiment, 50% of the bases were in reads longer than 10 Kb, which facilitated the assembly of reads into contigs with an average N50 value of 3.5 Mb. The programs antiSMASH and PRISM predicted 629 and 295 BGCs, respectively. We connected BGCs to metabolites for N,N-dimethyl cyclic-ditryptophan, a novel lassopeptide and three known Actinomycete-associated siderophores, namely mirubactin, heterobactin and salinichelin.

ImportanceShort-read sequencing of GC-rich genomes such as Actinomycetes results in a fragmented genome assembly and truncated biosynthetic gene clusters (often 10 to >100 Kb long), which hinders our ability to understand the biosynthetic potential of a given strain and predict the molecules that can be produced. The current study demonstrates that contiguous DNA assemblies, suitable for analysis of BGCs, can be obtained through low-coverage, multiplexed sequencing on Flongle, which provides a new low-cost workflow ($30-40 per strain) for sequencing Actinomycete strain libraries.


title: Towards the extended barcode concept: Generating DNA reference data through genome skimming of danish plants

abstract:
BackgroundRecently, there has been a push towards the extended barcode concept of utilising chloroplast genomes (cpGenome) and nuclear ribosomal DNA (nrDNA) sequences for molecular identification of plants instead of the standard barcode regions. These extended barcodes has a wide range of applications, including biodiversity monitoring and assessment, primer design, and evolutionary studies. However, these extended barcodes are not well represented in global reference databases. To fill this gap, we generated cpGenomes and nrDNA reference data from genome skims of 184 plant species collected in Denmark. We further explored the application of our generated reference data for molecular identifications of plants in an environmental DNA metagenomics study.

ResultsWe assembled partial cpGenomes for 82.1% of sequenced species and full or partial nrDNA sequences for 83.7% of species. We added all assemblies to GenBank, of which chloroplast reference data from 101 species and nuclear reference data from 6 species were not previously represented. On average, we recovered 45 genes per species. The rate of recovery of standard barcodes was higher for nuclear barcodes (>89%) than chloroplast barcodes (< 60%). Extracted DNA yield did not affect assembly outcome, whereas high GC content did so negatively. For the in silico simulation of metagenomic reads, taxonomic assignments using the reference data generated had better species resolution (94.9%) as compared to GenBank (18.1%) without any identification errors.

ConclusionsGenome skimming generates reference data of both standard barcodes and other loci, contributing to the global DNA reference database for plants.


title: Forgetting Enhances Episodic Control with Structured Memories

abstract: Forgetting is a normal process in healthy brains, and evidence suggests that the mammalian brain forgets more than is required based on limitations of mnemonic capacity. Episodic memories, in particular, are liable to be forgotten over time. Researchers have hypothesized that it may be beneficial for decision making to forget episodic memories over time. Reinforcement learning offers a normative framework in which to test such hypotheses. Here, we show that a reinforcement learning agent that uses an episodic memory cache to find rewards in maze environments can forget a large percentage of older memories without any performance impairments, if they utilize mnemonic representations that contain structural information about space. Moreover, we show that some forgetting can actually provide a benefit in performance compared to agents with unbounded memories. Our analyses of the agents show that forgetting reduces the influence of outdated information and states which are not frequently visited on the policies produced by the episodic control system. These results support the hypothesis that some degree of forgetting can be beneficial for decision making, which can help to explain why the brain forgets more than is required by capacity limitations.

category: animal behavior and cognition


title: Experimental evidence that chronic outgroup conflict reduces reproductive success in a cooperatively breeding fish

abstract: Conflicts with conspecific outsiders are common in group-living species, from ants to primates, and are argued to be an important selective force in social evolution. However, whilst an extensive empirical literature exists on the behaviour exhibited during and immediately after interactions with rivals, only very few observational studies have considered the cumulative fitness consequences of outgroup conflict. Using a cooperatively breeding fish, the daffodil cichlid (Neolamprologus pulcher), we conducted the first experimental test of the effects of chronic outgroup conflict on reproductive investment and output.  Intruded groups received long-term simulated territorial intrusions by neighbours that generated consistent group-defence behaviour; matched  Control groups (each the same size and with the same neighbours as an Intruded group) received no intrusions in the same period. Intruded groups had longer inter-clutch intervals and produced eggs with less protein than Control groups. Despite this lower egg investment, Intruded groups provided more parental care, achieving similar hatching success to Control groups. Ultimately, however, Intruded groups had fewer and smaller surviving offspring than Control groups at 1-month post-hatching. We therefore provide experimental evidence that outgroup conflict can decrease fitness via cumulative effects on reproductive success, confirming the selective potential of this empirically neglected aspect of sociality.


title: MOV10 facilitates messenger RNA decay in an N6-methyladenosine (m6A) dependent manner to maintain the mouse embryonic stem cells state

abstract: N6-methyladenosine (m6A) is the most predominant internal mRNA modification in eukaryotes, recognised by its reader proteins (so-called m6A-readers) for regulating subsequent mRNA fates -- splicing, export, localisation, decay, stability, and translation -- to control several biological processes. Although a few m6A-readers have been identified, yet the list is incomplete. Here, we identify a new m6A-reader protein, Moloney leukaemia virus 10 homologue (MOV10), in the m6A pathway. MOV10 recognises m6A-containing mRNAs with a conserved GGm6ACU motif. Mechanistic studies uncover that MOV10 facilitates mRNA decay of its bound m6A-containing mRNAs in an m6A-dependent manner within the cytoplasmic processing bodies (P-bodies). Furthermore, MOV10 decays the Gsk-3{beta} mRNA through m6A that stabilises the {beta}-CATENIN expression of a WNT/{beta}-CATENIN signalling pathway to regulate downstream NANOG expression for maintaining the mouse embryonic stem cells (mESCs) state. Thus, our findings reveal how a newly identified m6A-reader, MOV10 mediates mRNA decay via m6A that impact embryonic stem cell biology.

category: biochemistry


title: Data and Diversity Driven Development of a Shotgun Crystallisation Screen using the Protein Data Bank

abstract: Protein crystallisation has for decades been a critical and restrictive step in macro-molecular structure determination via X-ray diffraction. Crystallisation typically involves a multi-stage exploration of the available chemical space, beginning with an initial sampling (screening) followed by iterative refinement (optimisation). Effective screening is important for reducing the number of optimisation rounds required, reducing the cost and time required to determine a structure. Here, we propose an initial screen (Shotgun II) derived from analysis of the up-to-date Protein Data Bank (PDB) and compare it with the previously derived (2014) Shotgun I screen. In an update to that analysis, we clarify that the Shotgun approach entails finding the crystallisation conditions which cover the most diverse space of proteins by sequence found in the PDB - which can be mapped to the well known Maximum Coverage problem in computer science. With this realisation we are able to apply a more effective algorithm for selecting conditions, such that the Shotgun II screen outperforms the Shotgun I screen both in protein coverage and quantity of data input. Our data demonstrates that the Shotgun I screen, compared with alternatives, has been remarkably successful over the seven years it has been in use, indicating that Shotgun II is likely to be a highly effective screen.


title: Selecting Clustering Algorithms for IBD Mapping

abstract:
BackgroundGroups of distantly related individuals who share a short segment of their genome identical-by-descent (IBD) can provide insights about rare traits and diseases in massive biobanks via a process called IBD mapping. Clustering algorithms play an important role in finding these groups. We set out to analyze the fitness of commonly used, fast and scalable clustering algorithms for IBD mapping applications. We designed a realistic benchmark for local IBD graphs and utilized it to compare clustering algorithms in terms of statistical power. We also investigated the effectiveness of common clustering metrics as replacements for statistical power.

ResultsWe simulated 3.4 million clusters across 850 experiments with varying cluster counts, false-positive, and false-negative rates. Infomap and Markov Clustering (MCL) community detection methods have high statistical power in most of the graphs, compared to greedy methods such as Louvain and Leiden. We demonstrate that standard clustering metrics, such as modularity, cannot predict statistical power of algorithms in IBD mapping applications, though they can help with simulating realistic benchmarks. We extend our findings to real datasets by analyzing 3 populations in the Population Architecture using Genomics and Epidemiology (PAGE) Study with 51,000 members and 2 million shared segments on Chromosome 1, resulting in the extraction of 39 million local IBD clusters across three different populations in PAGE. We used cluster properties derived in PAGE to increase the accuracy of our simulations and comparison.

ConclusionsMarkov Clustering produces a 30% increase in statistical power compared to the current state-of-art approach, while reducing runtime by 3 orders of magnitude; making it computationally tractable in modern large-scale genetic datasets. We provide an efficient implementation to enable clustering at scale for IBD mapping and poplation-based linkage for various populations and scenarios.


title: Looking at the BiG picture: Incorporating bipartite graphs in drug response prediction

abstract:
MotivationThe increasing number of publicly available databases containing drugs chemical structures, their response in cell lines, and molecular profiles of the cell lines has garnered attention to the problem of drug response prediction. However, many existing methods do not fully leverage the information that is shared among cell lines and drugs with similar structure. As such, drug similarities in terms of cell line responses and chemical structures could prove to be useful in forming drug representations to improve drug response prediction accuracy.

ResultsWe present two deep learning approaches, BiG-DRP and BiG-DRP+, for drug response prediction. Our models take advantage of the drugs chemical structure and the underlying relationships of drugs and cell lines through a bipartite graph and a heterogenous graph convolutional network that incorporate sensitive and resistant cell line information in forming drug representations. Evaluation of our methods and other state-of-the-art models in different scenarios shows that incorporating this bipartite graph significantly improves the prediction performance. Additionally, genes that contribute significantly to the performance of our models also point to important biological processes and signaling pathways. Analysis of predicted drug response of patients tumors using our model revealed important associations between mutations and drug sensitivity, illustrating the utility of our model in pharmacogenomics studies.

Availability and ImplementationAn implementation of the algorithms in Python is provided in github.com/ddhostallero/BiG-DRP.

Contactamin.emad@mcgill.ca

Supplementary InformationOnline-only supplementary data is available at the journals website.