title: Detection of Multiple Types of Cancer Driver Mutations Us-ing Targeted RNA Sequencing in NSCLC

abstract:
Currently, DNA and RNA are used separately to capture different types of gene mutations. DNA is commonly used for the detection of SNVs, indels and CNVs; RNA is used for analysis of gene fusion and gene expression. To perform both DNA sequencing (DNA-seq) and RNA-seq, material is divided into two copies, and two different procedures are required for sequencing. Due to overconsumption of samples and experimental process complexity, it is necessary to create an experimental method capable of analyzing SNVs, indels, fusions and expression.

We developed an RNA-based hybridization capture panel targeting actionable driver oncogenes in solid tumors and corresponding sample preparation and bioinformatics workflows. Analytical validation with an RNA standard reference containing 16 known fusion mutations and 6 SNV mutations demonstrated a detection specificity of 100.0% [95% CI 88.7%~100.0%] for SNVs and 100.0% [95% CI 95.4%~100.0%] for fusions. The targeted RNA panel achieved a 0.73-2.63 copies/ng RNA lower limit of detection (LOD) for SNVs and 0.21-6.48 copies/ng RNA for fusions. Gene expression analysis revealed a correlation greater than 0.9 across all 15 cancer-related genes between the RNA-seq results and targeted RNA panel.

Among 1253 NSCLC FFPE tumor samples, multiple mutation types were called from DNA- and RNA-seq data and compared between the two assays. The DNA panel detected 103 fusions and 21 METex14 skipping events; 124 fusions and 26 METex14 skipping events were detected by the target RNA panel; 21 fusions and 4 METex14 skipping events were only detected by the target RNA panel. Among the 173 NSCLC samples negative for targetable mutations by DNA-seq, 15 (15/173, 8.67%) showed targetable gene fusions that may change clinical decisions with RNA-seq. In total, 226 tier I and tier II missense variants for NSCLC were analyzed at genomic (DNA-seq) and transcriptomic (RNA-seq) levels. The positive percent agreement (PPA) was 97.8%, and the positive predictive value (PPV) was 98.6%. Interestingly, variant allele frequencies were generally higher at the RNA level than at the DNA level, suggesting relatively dominant expression of mutant alleles. PPA was 97.6% and PPV 99.38% for EGFR 19del and 20ins variants.

We also explored the relationship of RNA expression with gene copy number and protein expression. The RPKM of EGFR transcripts assessed by the RNA panel showed a linear relationship with copy number quantified by the DNA panel, with an R of 0.8 in 1253 samples. In contrast, MET gene expression is regulated in a more complex manner. In IHC analysis, all 3+ samples exhibited higher RPKM levels; IHC level of 2+ and below showed lower RNA expression.

Parallel DNA- and RNA-seq and systematic analysis demonstrated the accuracy and robustness of the RNA sequencing panel in identifying multiple types of variants for cancer therapy.

Contact: zhaojia0327@126.com


title: Leaked genomic and mitochondrial DNA contribute to the host response to noroviruses in a STING-dependent manner

abstract:
The cGAS-STING pathway is central to the IFN response against DNA viruses. However, recent studies are increasingly demonstrating its role in the restriction of some RNA viruses. Here we show that the cGAS-STING pathway also contributes to the IFN response against noroviruses, positive-sense single-stranded RNA viruses that are now one of the most common causes of infectious gastroenteritis world-wide. We show a significant reduction in IFN-{beta} induction and a corresponding increase in viral replication in norovirus-infected cells following STING inhibition, knockdown or deletion. Upstream of STING, we show that cells lacking either cGAS or IFI16 also have severely impaired IFN responses. Further, we demonstrate that immunostimulatory host genome-derived DNA, and to a lesser extent mitochondrial DNA, accumulate in the cytosol of norovirus-infected cells. And lastly, overexpression of the viral NS4 protein was sufficient to drive the accumulation of cytosolic DNA. Together, our data elucidate a role for cGAS, IFI16 and STING in the restriction of noroviruses, and demonstrate for the first time the utility of host genomic DNA as a damage-associated molecular pattern in cells infected with an RNA virus.

HighlightsO_LIcGAS, IFI16 and STING are required for a robust IFN response against noroviruses
C_LIO_LINuclear and mitochondrial DNA accumulate in the cytosols of infected cells
C_LIO_LIViral NS4 mediates accumulation of cytosolic DNA
C_LI

category: molecular biology


title: An Algorithm to Quantify Inducible Protein Condensates In Eukaryotic Cells

abstract:
Reorganization of cellular proteins into subcellular compartments, such as the rearrangement of RNA-binding proteins into cytoplasmic stress granules and P-bodies, is a well-recognized, widely studied physiological process currently under intense investigation. Using the assembly of a novel, inducible, nuclear granule formed from the yeast RNA-binding transcription termination factors Nab3 and Nrd1, we present a freely-accessible, high-throughput and unbiased algorithm written in MATLAB that detects and measures protein distribution, partitioning, and sequestration into subcellular compartments captured by fluorescence microscopy; an invaluable advancement to current image analysis methods which utilize experiment-specific custom scripts or subjective manual counting. Employing our algorithm, we quantified thousands of cells, ensuring rigorous examination of Nab3 granule formation across strains with reproducible statistical analyses. We document strain differences in Nab3 granule formation and an associated growth defect. Additionally, we applied our algorithm to immunofluorescent images of the inducible polymerization into filaments of an enzyme in human cells, demonstrating the algorithms versatility and adaptability.

SUMMARY STATEMENTWe describe a computational tool that enables the quantification of protein condensation during assembly of a subnuclear compartment. The algorithm scores assembly of fluorescently tagged proteins in yeast or human cells.


title: MicroRNA-27a-5p inhibits proliferation, migration and invasion and promotes apoptosis of Wilms tumor cell by targeting PBOV1

abstract: Wilms tumor is the most common type of renal tumor in children. MicroRNAs (miRNA) are small non-coding RNAs that play crucial regulatory roles in tumorigenesis. We aimed to study the expression profile and function of miR-27a-5p in Wilms tumor. MiR-27a-5p expression was downregulated in human Wilms tumor tissues. Functionally, overexpression of miR-27a-5p promoted cell apoptosis of Wilms tumor cells. Furthermore, upregulated miR-27a-5p delayed xenograft Wilms tumor tumorigenesis in vivo. Bioinformatics analysis predicted miR-27-5p directly targeted to the 3-untranslated region (UTR) of PBOV1 and luciferase reporter assay confirmed the interaction between miR-27a-5p and PBOV1. The function of PBOV1 in Wilms tumor was evaluated in vitro and knockdown of PBOV1 dampened cell migration. In addition, overexpression of PBOV1 antagonized the tumor-suppressive effect of miR-27a-5p in Wilms tumor cells. Collectively, our findings reveal the regulatory axis of miR-27-5p/PBOV1 in Wilms tumor and miR-27a-5p might serve as a novel therapeutic target in Wilms tumor.

category: cell biology


title: Evolutionarily conserved genetic interactions between nphp-4 and bbs-5 mutations exacerbate ciliopathy phenotypes

abstract: Primary cilia are sensory and signaling hubs with a protein composition that is distinct from the rest of the cell due to the barrier function of the transition zone (TZ) at the base of the cilium. Protein transport across the TZ is mediated in part by the BBSome, and mutations disrupting TZ and BBSome proteins cause human ciliopathy syndromes. Ciliopathies have phenotypic variability even among patients with identical genetic variants, suggesting a role for modifier loci. To identify potential ciliopathy modifiers, we performed a mutagenesis screen on nphp-4 mutant C. elegans and uncovered a novel allele of bbs-5. Nphp-4;bbs-5 double mutant worms have phenotypes not observed in either individual mutant strain. To test whether this genetic interaction is conserved, we also analyzed zebrafish and mice mutants. While Nphp4 mutant zebrafish appeared overtly normal, Bbs5 mutants exhibited scoliosis. When combined, Nphp4;Bbs5 double mutant zebrafish did not exhibit synergistic effects, but the lack of a phenotype in Nphp4 mutants makes interpreting these data difficult. In contrast, viable Nphp4;Bbs5 double mutant mice were not obtained and there were fewer mice than expected carrying three mutant alleles. Additionally, postnatal loss of Bbs5 in mice using a conditional allele compromised survival when combined with a Nphp4 allele. As cilia are formed in the double mutant mice, the exacerbated phenotype is likely a consequence of disrupted ciliary signaling. Collectively, these data support an evolutionarily conserved genetic interaction between Bbs5 and Nphp4 alleles that may contribute to the variability in ciliopathy phenotypes.

category: genetics


title: The Specious Art of Single-Cell Genomics

abstract: Dimensionality reduction is standard practice for filtering noise and identifying relevant dimensions in large-scale data analyses. In biology, single-cell expression studies almost always begin with reduction to two or three dimensions to produce  all-in-one visuals of the data that are amenable to the human eye, and these are subsequently used for qualitative and quantitative analysis of cell relationships. However, there is little theoretical support for this practice. We examine the theoretical and practical implications of low-dimensional embedding of single-cell data, and find extensive distortions incurred on the global and local properties of biological patterns relative to the highdimensional, ambient space. In lieu of this, we propose semi-supervised dimensionality reduction to higher dimension, and show that such targeted reduction guided by the metadata associated with single-cell experiments provides useful latent space representations for hypothesis-driven biological discovery.

category: genomics


title: High temporal resolution RNA-seq time course data reveals mammalian lncRNA activationmirrors neighbouring protein-coding genes

abstract:
BackgroundThe advent of next-generation sequencing revealed extensive transcription beyond protein-coding genes, identifying tens of thousands of long non-coding RNAs (lncRNAs). Selected functional examples raised the possibility that lncRNAs, as a class, may maintain broad regulatory roles. Compellingly, lncRNA expression is strongly linked with adjacent protein-coding gene expression, suggesting a potential cis-regulatory function. Evidence for these regulatory roles may be obtained through careful examination of the precise timing of lncRNA expression relative to adjacent protein-coding genes.

ResultsWhere causal cis-regulatory relationships exist, lncRNA activation is expected to precede changes in adjacent target gene expression. Using an RNA-seq time course of uniquely high temporal resolution, we profiled the expression dynamics of several thousand lncRNAs and protein-coding genes in synchronized, transitioning human cells. Our findings reveal lncRNAs are expressed synchronously with adjacent protein-coding genes. Analysis of lipopolysaccharide-activated mouse dendritic cells revealed the same temporal relationship observed in transitioning human cells.

ConclusionOur findings suggest broad-scale cis-regulatory roles for lncRNAs are not common. The strong association between lncRNAs and adjacent genes may instead indicate an origin as transcriptional by-products from active protein-coding gene promoters and enhancers.

category: bioinformatics


title: Cell competition between wild-type and JAK2V617F mutant cells prevents disease relapse after stem cell transplantation in a murine model of myeloproliferative neoplasm

abstract: Disease relapse after allogeneic stem cell transplantation is a major cause of treatment-related morbidity and mortality in patients with myeloproliferative neoplasms (MPNs). The cellular and molecular mechanisms for MPN relapse are not well understood. Here, we established a murine model of MPN relapse, in which [~]60% of the MPN recipient mice develop disease relapse after receiving stem cell transplantation with wild-type marrow donor. Using this model, we find that impaired wild-type cell function is associated with MPN disease relapse. We also show that competition between wild-type and JAK2V617F mutant cells can modulate the immune cell composition and PD-L1 expression induced by the JAK2V617F oncogene. These results suggest that cell competition between wild-type donor cells and JAK2V617F mutant recipient cells can prevent MPN disease relapse after stem cell transplantation.

category: cancer biology


title: Utilizing Pine Needles to Temporally and Spatially Profile Per- and Polyfluoroalkyl Substances

abstract: As concerns continue to mount over exposure to per- and polyfluoroalkyl substances (PFAS), novel methods of profiling their presence and modifications are greatly needed as some have known toxic and bioaccumulative characteristics while others have unknown effects. This task however is not simple as over 5000 PFAS of interest have been named by the Environmental Protection Agency and this list continues to grow daily. In this work, we utilized widely available archived and field-sampled pine needles and a novel non-targeted analytical method to evaluate the temporal and spatial presence of numerous PFAS. Over 70 PFAS were detected in the pine needles from this study, providing information from the last six decades related to PFAS exposure, contamination, and reduction.

category: pharmacology and toxicology


title: L-citrulline ameliorates pathophysiology in a rat model of superimposed preeclampsia

abstract: Preeclampsia, characterized by hypertension, proteinuria, and fetal growth restriction, is one of the leading causes of maternal and perinatal mortality. By far, there is no effective pharmacological therapy for preeclampsia. The present study was conducted to investigate the effects of L-citrulline supplementation in Dahl salt-sensitive rat, a model of superimposed preeclampsia. Parental DSSR were treated with L-citrulline (2.5 g/L in drinking water) from the day of mating to the end of lactation period. Blood pressure of the rats was monitored throughout pregnancy and markers of preeclampsia were assessed. Endothelial function of the pregnant DSSR was assessed by wire myograph. L-citrulline supplementation significantly reduced gestational hypertension, proteinuria, and levels of circulating soluble fms-like tyrosine kinase 1 in DSSR. L-citrulline improved maternal endothelial function by augmenting the production of nitric oxide in the aorta and improving endothelium-derived hyperpolarizing factor-mediated vasorelaxation in resistance arteries. L-citrulline supplementation improved placental insufficiency and fetal growth, which were associated with an enhancement of angiogenesis and reduction of fibrosis and senescence in the placentas. In addition, L-citrulline downregulated genes involved in the toll-like receptor 4 and nuclear factor-{kappa}B signaling pathway. In conclusion, this study shows that L-citrulline supplementation reduces gestational hypertension, improves placentation and fetal growth in a rat model of superimposed preeclampsia. L-citrulline supplementation may represent an effective and safe therapeutic strategy for preeclampsia that benefit both the mother and the fetus.


title: Covariance-based information processing in reservoir computing systems

abstract: In biological neuronal networks, information representation and processing are achieved through plasticity learning rules that have been empirically characterized as sensitive to second and higher-order statistics in spike trains. However, most models in both computational neuroscience and machine learning aim to convert diverse statistical properties in inputs into first-order statistics in outputs, like in modern deep learning networks. In the context of classification, such schemes have merit for inputs like static images, but they are not well suited to capture the temporal structure in time series. In contrast, the recently developed covariance perceptron uses second-order statistics by mapping input covariances to output covariances in a consistent fashion. Here, we explore the applicability of covariance-based perceptron readouts in reservoir computing networks to classify synthetic multivariate time series structured at different statistical orders (first and second). We show that the second-order framework outperforms or matches the classical mean paradigm in terms of accuracy. Our results highlight a nontrivial relationship between input and reservoir properties in generating the output reservoir activity, which suggests an important role for recurrent connectivity in transforming information representations in biologically inspired architectures. Finally, we solve a speech recognition task for the classification of spoken digits to further demonstrate the potential of covariance-based decoding for real data.

category: neuroscience


title: Stop codon recoding is widespread in diverse phage lineages and has the potential to regulate translation of late stage and lytic genes

abstract: The genetic code is a highly conserved feature of life. However, some "alternative" genetic codes use reassigned stop codons to code for amino acids. Here, we survey stop codon recoding across bacteriophages (phages) in human and animal gut microbiomes. We find that stop codon recoding has evolved in diverse clades of phages predicted to infect hosts that use the standard code. We provide evidence for an evolutionary path towards recoding involving reduction in the frequency of TGA and TAG stop codons due to low GC content, followed by acquisition of suppressor tRNAs and the emergence of recoded stop codons in structural and lysis genes. In analyses of two distinct lineages of recoded virulent phages, we find that lysis-related genes are uniquely biased towards use of recoded stop codons. This convergence supports the inference that stop codon recoding is a strategy to regulate the expression of late stage genes and control lysis timing. Interestingly, we identified prophages with recoded stop codons integrated into genomes of bacteria that use standard code, and hypothesize that recoding may control the lytic-lysogenic switch. Alternative coding has evolved many times, often in closely related lineages, indicating that genetic code is plastic in bacteriophages and adaptive recoding can occur over very short evolutionary timescales.

category: microbiology


title: Frameshifting at collided ribosomes is modulated by elongation factor eEF3 and by Integrated Stress Response regulators Gcn1 and Gcn20

abstract: Ribosome stalls can result in ribosome collisions that elicit quality control responses, one function of which is to prevent frameshifting by the stalled ribosome, an activity that entails interaction of the conserved yeast protein Mbf1 with uS3 on colliding ribosomes. However, the full spectrum of factors that mediate frameshifting during ribosome collisions is unknown. To delineate such factors in the yeast Saccharomyces cerevisiae, we used genetic selections for mutants that either suppress or increase frameshifting from a known ribosome stall site, CGA codon repeats. We show that the general translation elongation factor eEF3 promotes frameshifting, while Integrated Stress Response (ISR) pathway components Gcn1 and Gcn20 suppress frameshifting. We found a mutant form of eEF3 that specifically suppressed frameshifting, but not translation inhibition by CGA codons. Thus, we infer that frameshifting at collided ribosomes requires eEF3, which facilitates tRNA-mRNA translocation and E-site tRNA release in yeast and other single cell organisms. By contrast, we found that removal of either Gcn1 or Gcn20, which bind collided ribosomes with Mbf1, increased frameshifting. Thus, we conclude that frameshifting is suppressed by Gcn1 and Gcn20, although these effects are not mediated through activation of the ISR. Furthermore, we examined the relationship of eEF3-mediated frameshifting to other quality control mechanisms, finding that the eEF3-mediated frameshifting competes with No-Go decay, Mbf1 and Gcn1/20. Thus, these results provide evidence of a direct link between translation elongation and frameshifting at collided ribosomes, as well as evidence that frameshifting competes with other quality control pathways that act on collided ribosomes.


title: Developmental Venous Anomalies are a Genetic Primer for Cerebral Cavernous Malformations

abstract: Cerebral cavernous malformations (CCM) are a neurovascular anomaly that may occur sporadically in otherwise healthy individuals, or be inherited by autosomal dominant mutations in the genes that encode the proteins of the CCM signaling complex (KRIT1, CCM2, or PDCD10)1-4. CCMs have long been known to follow a genetic two-hit model where lesion formation is initiated by somatic mutations resulting in biallelic loss of a CCM complex gene5-8. Recent studies have shown that somatic mutations in MAP3K3 and PIK3CA also contribute to CCM pathogenesis9-11; however, it remains unclear how these mutations contribute to sporadic versus familial cases. Here we show that somatic mutations in MAP3K3 are mutually exclusive with mutations in CCM complex genes and that mutations in MAP3K3 contribute to sporadic, but not familial CCM. Using single-nucleus DNA sequencing, we show that co-occurring MAP3K3 and PIK3CA mutations are present within the same clonal population of cells. Furthermore, we identify PIK3CA mutations in CCM-associated developmental venous anomalies (DVA). It has long been known that sporadic CCM often develop in the vicinity of a DVA. However, the underlying cause of this association is unknown12-14. In this first report of the molecular pathology of CCM-associated DVA, we find that the identical PIKC3A mutation is found in both the DVA and its associated CCM, but that an activating MAP3K3 mutation appears only in the CCM. These results support a mechanism where DVA develop as the result of a PIK3CA mutation, creating a region of the brain vasculature that functions as a genetic primer for CCM development following acquisition of an additional somatic mutation.


title: Pro70 Hydroxylation of Actin Impairs Cell Motility Through Binding with VHL and Blocking His73 Methylation

abstract: Protein hydroxylation is a post translational modification happens on various amino acids, which is catalyzed by the oxoglutarate and oxygen dependent dioxygenases. The best characterized hydroxylated protein is the hypoxia inducible factor (HIF), which is degraded by VHL/elongin C/elongin B/cullin 2/RBX1 (VCB/CR) E3 complex under normal oxygen conditions. Hypoxia or inhibitors (including FG4592 and MK8617) of PHDs stabilize HIF1a and regulate its downstream targets. Prolyl hydroxylase, including PHD2 and PHD3 has been reported in regulating actin polymerization and cell motility. Here, we found MK8617 regulated cell motility in Von Hippel Lindau (VHL) dependent manner. Through the protein hydroxylation proteome experiment upon MK8617 treatment, we identified Pro70 in actin could be hydroxylated and near to His73, which has been reported be methylated and stabilize actin polymerization. Using biochemical assay, we found that binding of VHL with hydroxylated actin (Pro70) decrease the His73 methylation by blocking the interaction of actin with SETD3, the His73 methyltransferase, and further regulated actin polymerization and cell motility. In summary, our study revealed that hypoxia and deficiencies in the VHL, in a HIF independent and prolyl hydroxylation dependent manner, regulate actin polymerization and cell motility through the PTM (Post Translational Modifications) crosstalk.

category: biochemistry


title: Genomic drivers of large B-cell lymphoma resistance to CD19 CAR-T therapy

abstract: Chimeric antigen receptor-reprogrammed autologous T cells directed to CD19 are breakthrough immunotherapies for heavily pretreated patients with aggressive B-cell lymphomas but still fail to cure most patients. Host inflammatory and tumor microenvironmental factors associate with CAR-19 resistance, but the tumor-intrinsic factors underlying these phenomena remain undefined. To characterize genomic drivers of resistance, we interrogated whole genome sequencing of 30 tumor samples from 28 uniformly CAR-19-treated large-cell lymphoma patients. We reveal that patterns of genomic complexity (i.e., chromothripsis and APOBEC mutational activity), and distinct genomic alterations (deletions of RB1 or RHOA) associate with more exhausted immune microenvironments and poor outcome after CAR-19 therapy. Strikingly, pretreatment reduced expression or sub-clonal mutation of CD19 did not affect responses, suggesting CAR-19 therapy successes are due not only to direct antigen-dependent cytotoxicity but require surmounting immune exhaustion in tumor microenvironments to permit broader host responses that eliminate tumors.


title: Diterpenoid Vinigrol activates ATF4/DDIT3-mediated PERK/eIF2 arm of unfolded protein response to drive breast cancer cell death

abstract: Vinigrol is a natural diterpenoid with unprecedented chemical structure, driving great efforts into its total synthesis and the chemical analogs in the past decades. Despite its pharmacological efficacies reported on anti-hypertension and anti-clot, comprehensive functional investigations on Vinigrol and the underlying molecular mechanisms are entirely missing. In this study, we carried out a complete functional prediction of Vinigrol using a transcriptome-based strategy, Connectivity Map, and identified "anti-cancer" as the most prominent biofunction ahead of anti-hypertension and anti-depression/psychosis. A broad cytotoxicity was subsequently confirmed on multiple cancer types. Further mechanistic investigation on MCF7 cells revealed that its anti-cancer effect is mainly through activating PERK/eIF2 arm of unfolded protein response (UPR) and subsequent upregulation of p53/p21 to halt the cell cycle. The other two branches of UPR, IRE1 and ATF6, are functionally irrelevant to Vinigrol-induced cell death. CRISPR/Cas9-based gene activation, repression, and knockout systems identified essential contribution of ATF4/DDIT3 not ATF6 to the death process. This study unraveled a broad anti-cancer function of Vinigrol and its underlying targets and regulatory mechanisms, and also paved the way for further inspection on the structure-efficacy relationship of the whole compound family, making them a novel cluster of chemical hits for cancer therapy.


title: Using herbarium samples for NGS methods - a methodological comparison

abstract: Herbaria harbor a tremendous amount of plant specimens that are rarely used for plant systematic studies. The main reason is the difficulty to extract a decent quantity of good quality DNA from the preserved plant material. While the extraction of ancient DNA in animals is well established, studies including old plant material are still underrepresented. In our study we compared the standard Qiagen DNeasy Plant Mini Kit and a specific PTB-DTT protocol on two different plant genera (Xanthium L. and Salix L.). The included herbarium material covered about two centuries of plant collections. A selected subset of samples was used for a standard library preparation as well as a target enrichment approach. The results revealed that PTB-PTT resulted in higher quantity and quality regarding DNA yield. For relatively recent herbarium specimens, and despite the lower overall yield of DNA, the Qiagen Kit resulted in better sequencing results regarding the number of filtered and mapped reads. We were able to successfully sequence a sample from 1820 and conclude that it is possible to include old herbarium specimens in NGS approaches. This opens a treasure box for phylogenomic research.

category: plant biology


title: A quantitative mapping approach to identify direct interactions within complexomes

abstract:
Complementary methods are required to fully characterize all protein complexes, or the complexome, of a cell. Affinity purification coupled to mass-spectrometry (AP-MS) can identify the composition of complexes at proteome-scale. However, information on direct contacts between subunits is often lacking. In contrast, solving the 3D structure of protein complexes can provide this information, but structural biology techniques are not yet scalable for systematic, proteome-wide efforts. Here, we optimally combine two orthogonal high-throughput binary interaction assays, LuTHy and N2H, and demonstrate that their quantitative readouts can be used to differentiate direct interactions from indirect associations within multiprotein complexes. We also show that LuTHy allows accurate distance measurements between proteins in live cells and apply these findings to study the impact of the polyglutamine expansion mutation on the structurally unresolved N-terminal domain of Huntingtin. Thus, we present a new framework based on quantitative interaction assays to complement structural biology and AP-MS techniques, which should help to provide first-approximation contact maps of multiprotein complexes at proteome-scale.

Graphical Abstract

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category: systems biology


title: Health assessment of the pink land iguana, Conolophus marthae.

abstract: The pink land iguana, Conolophus marthae, is one of four species of iguanas (three terrestrial and one marine) in the Galapagos Islands, and the only one listed as critically endangered by the IUCN. The species can only be found on the north-west slopes of the highest volcano on the island of Isabela and was first described to science in 2009. As part of a population telemetry study, a health assessment was authorized by the Galapagos National Park. Wild adult iguanas were captured on Wolf Volcano in September 2019 and April 2021 to record morphological and physiological parameters including body temperature, heart rate, body measurements, intraocular pressures, tear formation, and infrared iris images. Blood samples were also collected and analyzed. An i-STAT portable blood analyzer was used to obtain values for base excess in the extracellular fluid compartment (BEECF), glucose, hematocrit, hemoglobin, ionized calcium (iCa), partial pressure of carbon dioxide (pCO2), partial pressure of oxygen (pO2), percent oxygen saturation (sO2%), pH, potassium (K), and sodium (Na). Standard laboratory hematology techniques were employed for packed-cell-volume (PCV) determination. When possible, data were compared to previously published and available data for the other Galapagos iguanas. The results reported here provide baseline values that may be useful in detecting changes in health status among pink land iguanas affected by climate change, invasive species, anthropogenic threats, or natural disturbances.

category: zoology


title: IDENTIFICATION OF A NOVEL BDNF-SPECIFIC CORTICOSTRIATAL CIRCUITRY

abstract:
BDNF is released from axon terminals originating in the cerebral cortex onto striatal neurons. Here, we characterized BDNF in the corticostriatal circuitry. First, we utilized Bdnf-Cre and Ribotag transgenic mouse lines to label BDNF-positive cells in the cortex, and detected BDNF expression in the motor cortex, medial prefrontal cortex (mPFC) and the orbitofrontal cortex (OFC). Next, we used a retrograde viral tracing strategy, in combination with Bdnf-Cre knockin mice, to map the cortical outputs of BDNF neurons in the dorsal striatum. We found that the BDNF-positive prefrontal regions differentially project to the dorsal striatum. Specifically, BDNF-expressing neurons located in the mPFC project to both dorsolateral striatum (DLS) and dorsomedial striatum (DMS), and those located in the motor cortex project to the DLS. Surprisingly however, the BDNF-expressing OFC neurons differentially target the dorsal striatum depending on their mediolateral location. Specifically, the DMS is mainly innervated by the medial part of the OFC (mOFC) whereas, the DLS receives projections specifically from the ventrolateral region of the OFC (vlOFC). Next, using an anterograde viral tracing strategy, we confirmed the presence of a BDNF-specific vlOFC-DLS circuit. Finally, we show that overexpression of BDNF in the vlOFC activates TrkB signaling specifically in the DLS but not in the DMS demonstrating the functionality of this circuit. Our study uncovers a previously unknown neural circuit composed of BDNF-positive vlOFC neurons projecting to the DLS. These findings could have important implications for the role of BDNF signaling in the OFC as well as in other corticostriatal circuitries.

Significance StatementBDNF is released in axons upon neuronal depolarization. Surprisingly, careful mapping of BDNF projecting neurons in CNS has not been conducted. Using anterograde and retrograde viral strategies in combination with transgenic reporter mice, we mapped out corticostriatal BDNF circuits. We found that mPFC BDNF neurons project to both the DMS and DLS, and the motor cortex to the DLS only. In contrast, BDNF neurons in the OFC are anatomically segregated. Whereas BDNF in mOFC neurons project to the DMS, BDNF in vlOFC project specifically to the DLS. Our findings could be important to the study of BDNF in corticostriatal circuitries.


title: The PH/MyTH4/FERMmolecule MAX-1 inhibits UNC-5 activity in regulation of VD growth cone protrusion in Caenorhabditis elegans

abstract: UNC-6/Netrin is a secreted conserved guidance cue that regulates dorsal-ventral axon guidance of C. elegans and in the vertebral spinal cord. In the polarity/protrusion model of VD growth cone guidance away from ventrally-expressed UNC-6 (repulsion), UNC-6 first polarizes the growth cone via the UNC-5 receptor such that filopodial protrusions are biased dorsally. UNC-6 then regulates a balance of protrusion in the growth cone based upon this polarity. UNC-5 inhibits protrusion ventrally, and the UNC-6 receptor UNC-40/DCC stimulates protrusion dorsally, resulting in net dorsal growth cone outgrowth. UNC-5 inhibits protrusion through the flavin monooxygenases FMO-1, 4, and 5 and possible actin destabilization, and inhibits pro-protrusive microtubule entry into the growth cone utilizing UNC-33/CRMP. The PH/MyTH4/FERM myosin-like protein was previously shown to act with UNC-5 in VD axon guidance utilizing axon guidance endpoint analysis. Here, we analyzed the effects of MAX-1 on VD growth cone morphology during outgrowth. We found that max-1 mutant growth cones were smaller and less protrusive than wild-type, the opposite of the unc-5 mutant phenotype. Furthermore, genetic interactions suggest that MAX-1 might normally inhibit UNC-5 activity, such that in a max-1 mutant growth cone, UNC-5 is overactive. Our results, combined with previous studies suggesting that MAX-1 might regulate UNC-5 levels in the cell or plasma membrane localization, suggest that MAX-1 attenuates UNC-5 signaling by regulating UNC-5 stability or trafficking. In summary, in the context of growth cone protrusion, MAX-1 inhibits UNC-5, demonstrating the mechanistic insight that can be gained by analyzing growth cones during outgrowth in addition to axon guidance endpoint analysis.


title: C. elegans enteric motor neurons fire synchronized action potentials underlying the defecation motor program

abstract: The C. elegans nervous system was thought to be strictly analog, constituted solely by graded neurons. We recently discovered neuronal action potentials in the sensory neuron AWA; however, the extent to which the C. elegans nervous system relies on analog or digital neural signaling and coding is unclear. Here we report that the enteric motor neurons AVL and DVB fire all-or-none calcium-mediated action potentials that play essential roles in the rhythmic defecation behavior in C. elegans. Both AVL and DVB synchronously fire giant action potentials to faithfully execute all-or-none expulsion following the intestinal pacemaker. AVL fires unusual compound action potentials with each positive calcium-mediated spike followed by a potassium-mediated negative spike. The depolarizing calcium spikes in AVL are mediated by a CaV2 calcium channel UNC-2, while the negative potassium spikes are mediated by a repolarization-activated potassium channel EXP-2. Whole-body behavior tracking and simultaneous neural imaging in free-moving animals suggest that action potentials initiated in AVL in the head propagate along its axon to the tail and activate DVB through the INX-1 gap junction. Synchronized action potential spikes between AVL and DVB, as well as the negative spike and long-lasting afterhyperpolarization in AVL, play an important function in executing expulsion behavior. This work provides the first evidence that in addition to sensory coding, C. elegans motor neurons also use digital coding scheme to perform specific functions including long-distance communication and temporal synchronization, suggesting further, unforeseen electrophysiological diversity remains to be discovered in the C. elegans nervous system.


title: Post-stroke dendritic arbor regrowth - a cortical repair process requiring the actin nucleator Cobl

abstract: Ischemic stroke is a major cause of death and long-term disability. We demonstrate that middle cerebral artery occlusion in mice leads to a strong decline in dendritic arborization of penumbral neurons. These defects were subsequently repaired by an ipsilateral recovery process requiring the actin nucleator Cobl. Ischemic stroke and excitotoxicity, caused by calpain-mediated proteolysis, significantly reduced Cobl levels. In an apparently unique manner among excitotoxicity-affected proteins, this Cobl decline was rapidly restored by increased mRNA expression and Cobl then played a pivotal role in post-stroke dendritic arbor repair in peri-infarct areas. In Cobl KO mice, the dendritic repair window determined to span day 2-4 post-stroke in WT strikingly passed without any dendritic regrowth. Instead, Cobl KO penumbral neurons of the primary motor cortex continued to show the dendritic impairments caused by stroke. Our results thereby highlight a powerful post-stroke recovery process and identified causal molecular mechanisms critical during post-stroke repair.


title: Adar-mediated A-to-I editing is required for establishment of embryonic body axes in zebrafish

abstract: Adenosine deaminases (ADARs) catalyze the deamination of adenosine to inosine, also known as A-to-I editing, in RNA. Although A-to-I editing occurs widely across animals, and is well studied, new biological roles are still being discovered. Here, we study the role of A-to-I editing in early zebrafish development. We demonstrate that Adar, the zebrafish orthologue of mammalian ADAR1, is essential for establishing the antero-posterior and dorso-ventral axes and patterning. Genome-wide editing discovery revealed pervasive editing in maternal and the earliest zygotic transcripts, the majority of which occurred in the 3-UTR. Interestingly, transcripts implicated in gastrulation as well as dorso-ventral and antero-posterior patterning were found to contain multiple editing sites. Adar knockdown or overexpression affected gene expression and global editing patterns at 12 hpf, but not earlier. Our study established that RNA editing by Adar is necessary for the earliest steps of embryonic patterning along the zebrafish antero-posterior and dorso-ventral axes.

category: developmental biology


title: An efficient and robust HPLC method to determine the sialylation levels of human epithelial cells

abstract: Sialyltransferase, an enzyme responsible for attaching sialic acid to the cell surface, is reported to play a key role in cancer, making sialyltransferase a potent therapeutic target in drug development for cancer. Hence, this paper aimed to develop a simple method to detect and quantify sialic acids in cancer cells. An efficient method was developed using a reverse-phase ion-pairing HPLC-UV using triisopropanolamine as the ion-pairing agent with a C18 column. Neu5Ac was successfully eluted with the retention time 6.344 min with a flow rate of 0.4 mL/min. The proposed method was validated appropriately according to the AOAC guidelines (2013). This work demonstrates that the proposed method is not only relatively simple but also cost and time effective compared to pre-existing methods to successfully determine both free and protein-bound Neu5Ac in a complex cancer cell matrix. Furthermore, by applying the proposed method, a statistically significant decrease was observed for both HeLa and HuCCT1 cell lines with the application of deoxycholic acid - a known sialyltransferase inhibitor. Hence, the proposed method may be applicable to evaluate the effectiveness of sialyltransferase inhibitors.


title: The entomopathogenic nematode Steinernema hermaphroditum  is a self-fertilizing hermaphrodite and a genetically tractable system for the study of parasitic and mutualistic symbiosis

abstract: Entomopathogenic nematodes, including Heterorhabditis and Steinernema, are parasitic to insects and contain mutualistically symbiotic bacteria in their intestines (Photorhabdus and Xenorhabdus, respectively) and therefore offer opportunities to study both mutualistic and parasitic symbiosis. The establishment of genetic tools in entomopathogenic nematodes has been impeded by limited genetic tractability, inconsistent growth in vitro, variable cryopreservation, and low mating efficiency. We obtained the recently described Steinernema hermaphroditum strain CS34 and optimized its in vitro growth, with a rapid generation time on a lawn of its native symbiotic bacteria Xenorhabdus griffiniae. We developed a simple and efficient cryopreservation method. Previously, S. hermaphroditum isolated from insect hosts was described as first-generation hermaphroditic and second-generation gonochoristic. We discovered that CS34, when grown in vitro, produced consecutive generations of autonomously reproducing hermaphrodites accompanied by rare males. We performed mutagenesis screens in S. hermaphroditum that produced mutant lines with visible and heritable phenotypes. Genetic analysis of the mutants demonstrated that this species reproduces by self-fertilization rather than parthenogenesis and that its sex is determined chromosomally. Genetic mapping has thus far identified markers on the X chromosome and three of four autosomes. We report that S. hermaphroditum CS34 is the first consistently hermaphroditic entomopathogenic nematode and is suitable for genetic model development to study naturally occurring mutualistic symbiosis and insect parasitism.


title: Structural remodeling of SARS-CoV-2 spike protein glycans reveals the regulatory roles in receptor binding affinity

abstract:
Glycans of the SARS-CoV-2 spike protein are speculated to play functional roles in the infection processes as they extensively cover the protein surface and are highly conserved across the variants. To date, the spike protein has become the principal target for vaccine and therapeutic development while the exact effects of its glycosylation remain elusive. Experimental reports have described the heterogeneity of the spike protein glycosylation profile. Subsequent molecular simulation studies provided a knowledge basis of the glycan functions. However, there are no studies to date on the role of discrete glycoforms on the spike protein pathobiology. Building an understanding of its role in SARS-CoV-2 is important as we continue to develop effective medicines and vaccines to combat the disease. Herein, we used designed combinations of glycoengineering enzymes to simplify and control the glycosylation profile of the spike protein receptor-binding domain (RBD). Measurements of the receptor binding affinity revealed the regulatory effects of the RBD glycans. Remarkably, opposite effects were observed from differently remodeled glycans, which presents a potential strategy for modulating the spike protein behaviors through glycoengineering. Moreover, we found that the reported anti-SARS-CoV-(2) antibody, S309, neutralizes the impact of different RBD glycoforms on the receptor binding affinity. Overall, this work reports the regulatory roles that glycosylation plays in the interaction between the viral spike protein and host receptor, providing new insights into the nature of SARS-CoV-2. Beyond this study, enzymatic remodeling of glycosylation offers the opportunity to understand the fundamental role of specific glycoforms on glycoconjugates across molecular biology.

Covert art LegendsThe glycosylation of the SARS-CoV-2 spike protein receptor-binding domain has regulatory effects on the receptor binding affinity. Sialylation or not determines the "stabilizing" or "destabilizing" effect of the glycans. (Protein structure model is adapted from Protein Data Bank: 6moj. The original model does not contain the glycan structure.)

SignificanceGlycans extensively cover the surface of SARS-CoV-2 spike (S) protein but the relationships between the glycan structures and the protein pathological behaviors remain elusive. Herein, we simplified and harmonized the glycan structures in the S protein receptor-binding domain and reported their regulatory roles in human receptor interaction. Opposite regulatory effects were observed and were determined by discrete glycan structures, which can be neutralized by the reported S309 antibody binding to the S protein. This report provides new insight into the mechanism of SARS-CoV-2 S protein infection as well as S309 neutralization.


title: Synovial Fluid Mitochondrial DNA Concentration Reflects the Degree of Cartilage Damage After Naturally Occurring Articular Injury

abstract:
Posttraumatic osteoarthritis (PTOA) is a debilitating sequela to joint injury with no current therapeutics that can slow its progression. Early intervention, prior to the development of degenerative joint changes, has the potential for greater therapeutic success but requires early detection of joint injury. In other tissue types, trauma is associated with the extracellular release of mitochondrial DNA (mtDNA), which serves as a mitochondria-specific Damage Associated Molecular Pattern (mDAMP) to perpetuate inflammation. We demonstrated that chondrocytes release mtDNA following cellular stress and that mtDNA is increased in equine synovial fluid following experimental and naturally occurring mechanical injury to the joint surface. Moreover, we found a strong correlation between the degree of cartilage damage and mtDNA concentration. Finally, impact-induced mtDNA release was mitigated by mitoprotective treatment. These data suggest synovial fluid mtDNA may represent a sensitive marker of early articular injury, prior to the onset of changes on standard diagnostic imaging modalities.

One Sentence SummarySynovial fluid mitochondrial DNA increases after articular injury, is associated with the degree of cartilage damage, and is mitigated by mitoprotective treatment.


title: Single-cell chromatin and transcriptome dynamics of Synovial Fibroblasts transitioning from homeostasis to pathology in modelled TNF-driven arthritis

abstract: Synovial fibroblasts (SFs) are specialized cells of the synovium that provide nutrients and lubricants for the maintenance of proper function of diarthrodial joints. Chronic TNF signals are known to trigger activation of SFs and orchestration of arthritic pathology via proinflammatory effector functions, secretion of cartilage degrading proteases and promotion of osteolysis. We performed single-cell (sc) profiling of SFs transcriptome by RNA-sequencing (scRNA-seq) and of chromatin accessibility by scATAC-seq in normal mouse SFs and SFs derived from early and advanced TNF-driven arthritic disease. We describe here distinct subsets of SFs in the homeostatic synovium, serving diverse functions such as chondro- and osteogenesis, tissue repair and immune regulation. Strikingly, development of spontaneous arthritis by transgenic TNF overexpression primes the emergence of distinct pathology-associated SF subtypes. We reveal 7 constitutive and 2 disease-specific SF subtypes. The latter emerge in the early stage, expand in late disease and are localized in areas at the interface between the invasive pannus and the articular bone. The associated transcription profiles are characterized by enhanced inflammatory responses, promigratory behaviour, neovascularization and collagen metabolic processes. Temporal reconstruction of transcriptomic events indicated which specific sublining cells may function as progenitors at the root of trajectories leading to intermediate subpopulations and culminating to a destructive lining inflammatory identity. Integrated analysis of chromatin accessibility and transcription changes revealed key transcription factors such as Bach and Runx1 to drive arthritogenesis. Parallel analysis of human arthritic SF data showed highly conserved core regulatory and transcriptional programs between the two species. Therefore, our study dissects the dynamic SF landscape in TNF-mediated arthritis and sets the stage for future investigations that might address the functions of specific SF subpopulations to understand joint pathophysiology and combat chronic inflammatory and destructive arthritic diseases.


title: Elys deficiency constrains Kras-driven tumour burden by amplifying oncogenic stress

abstract: The nucleoporin ELYS, encoded by AHCTF1, is a large multifunctional protein with essential roles in nuclear pore assembly and mitosis. Using a zebrafish model of hepatocellular carcinoma, in which the expression of an inducible mutant kras transgene (krasG12V) drives hepatocyte-specific hyperplasia and liver enlargement, we show that reducing ahctf1 gene dosage by 50% markedly shrinks tumour burden, while non-hyperplastic tissues are unaffected. We demonstrate that ahctf1 heterozygosity impairs nuclear pore formation, mitotic spindle assembly and chromosome segregation, leading to DNA damage and activation of TP53-dependent and independent mechanisms of cell death and cell cycle arrest. This selective vulnerability of cancer cells to mild disruption of Elys function uncovers a novel synthetic lethal interaction between ahctf1 and kras mutations that could be exploited therapeutically. Heterozygous expression of both ahctf1 and ranbp2, or treatment of heterozygous ahctf1 larvae with the nucleocytoplasmic transport inhibitor, Selinexor, completely blocked krasG12V-driven hepatocyte hyperplasia, revealing promising avenues for combinatorial treatments.


title: Transient DUX4 expression induces blastomere-like expression program that is marked by SLC34A2

abstract: DUX4 has recently been recognized as a key regulator in human embryonic genome activation (EGA). The exact role of DUX4 in human embryo is still elusive, partly due to the cytotoxicity of persistent DUX4 expression in cellular models. We report here that a transient DUX4 expression in human embryonic stem cells (hESCs) retains cell viability while inducing an EGA-like expression program in a subpopulation of the cells. These cells showed resemblance to 8-cell stage blastomeres and were thus named induced blastomere-like (iBM) cells. Trajectory inference from the single-cell RNA-seq data suggested that the expression profile of these cells progressed in a manner similar to the morula to blastocyst transition in human embryo. Finally, viable iBM cells could be enriched using an antibody against NaPi2b (SLC34A2), paving the way for further experimental approaches. The iBM cells can become a powerful tool to model transcriptional dynamics and regulation during early human embryogenesis.


title: Crossover patterning through kinase-regulated condensation and coarsening of recombination nodules

abstract: Meiotic recombination is highly regulated to ensure precise segregation of homologous chromosomes. Evidence from diverse organisms indicates that the synaptonemal complex (SC), which assembles between paired chromosomes, plays essential roles in crossover formation and patterning. Several additional "pro-crossover" proteins are also required for recombination intermediates to become crossovers. These typically form multiple foci or recombination nodules along SCs, and later accumulate at fewer, widely spaced sites. Here we report that in C. elegans CDK-2 is required to stabilize all crossover intermediates and stabilizes interactions among pro-crossover factors by phosphorylating MSH-5. Additionally, we show that the conserved RING domain proteins ZHP-3/4 diffuse along the SC and remain dynamic following their accumulation at recombination sites. Based on these and previous findings we propose a model in which recombination nodules arise through spatially restricted biomolecular condensation and then undergo a regulated coarsening process, resulting in crossover interference.


title: Extrinsic KRAS signaling shapes the pancreatic microenvironment through fibroblast reprogramming.

abstract: Oncogenic KRAS is the hallmark mutation of human pancreatic cancer and a driver of tumorigenesis in genetically engineered mouse models of the disease. While the tumor cell-intrinsic effects of oncogenic Kras expression have been widely studied, its role in regulating the extensive pancreatic tumor microenvironment is less understood. Using a genetically engineered mouse model of inducible and reversible oncogenic Kras expression and a combination of approaches that include mass cytometry and single cell RNA sequencing, we have discovered that non-cell autonomous (i.e., extrinsic) oncogenic KRAS signaling reprograms pancreatic fibroblasts, activating an inflammatory gene expression program. As a result, fibroblasts become a hub of extracellular signaling, mediating the polarization and function of pro-tumorigenic myeloid cells while also preventing tissue repair. Our study provides fundamental new knowledge on the mechanisms underlying the formation of the fibroinflammatory stroma in pancreatic cancer and highlights stromal pathways with the potential to be exploited therapeutically.


title: Does genetic differentiation underlie behavioral divergence in response tomigration barriers in sticklebacks? A common garden experiment

abstract:
Water management measures in the 1970s in the Netherlands have produced a large number of  resident populations of three-spined sticklebacks that are no longer able to migrate to the sea. This may be viewed as a replicated field experiment, allowing us to study how the resident populations are coping with human-induced barriers to migration. We have previously shown that residents are smaller, bolder, more exploratory, more active, more aggressive, exhibited lower shoaling and lower migratory tendencies compared to their ancestral  migrant counterparts. However, it is not clear if these differences in wild-caught residents and migrants reflect genetic differentiation, rather than different developmental conditions. To investigate this, we raised offspring of four crosses (migrant [male] x migrant [female], resident [male] x resident [female], migrant [male] x resident [female], resident [male] x migrant [female]) under similar controlled conditions and tested for differences in morphology and behavior as adults. We found that lab-raised resident sticklebacks exhibited lower shoaling and migratory tendencies as compared to lab-raised migrants, retaining the differences in their wild-caught parents. This indicates genetic differentiation of these traits. For all other traits, the lab-raised sticklebacks of the various crosses did not differ significantly, suggesting that the earlier-found contrast between wild-caught fish reflect differences in their environment. Our study shows that barriers to migration can lead to rapid differentiation in behavioral tendencies over contemporary timescales ([~]50 generations), and that part of these differences reflects genetic differentiation.

Significance statementMany organisms face changes to their habitats due to human activities. Much research is therefore dedicated to the question whether and how organisms are able to adapt to novel conditions. We address this question in three-spined sticklebacks, where water management measures cut off some populations, prohibiting their seasonal migration to the North Sea. In a previous study, we showed that wild-caught  resident fish exhibited markedly different behavior than migrants. To disentangle whether these differences reflect genetic differentiation or differences in the conditions under which the wild-caught fish grew up, we conducted crosses, raising the F1 offspring under identical conditions. As their wild-caught parents, the F1 of resident x resident crosses exhibited lower migratory and shoaling tendencies than the F1 of migrant x migrant crosses, while the F1 of hybrid crosses were intermediate. This suggests that [~]50 years of isolation are sufficient to induce behaviorally relevant genetic differentiation.

category: evolutionary biology


title: Robust, Universal Tree Balance Indices

abstract: Balance indices that quantify the symmetry of branching events and the compactness of trees are widely used to compare evolutionary processes or tree-generating algorithms. Yet existing indices have important shortcomings, including that they are unsuited to the tree types commonly used to describe the evolution of tumours, microbial populations, and cell lines. The contributions of this article are twofold. First, we define a new class of robust, universal tree balance indices. These indices take a form similar to Colless index but account for node sizes, are defined for trees with any degree distribution, and enable more meaningful comparison of trees with different numbers of leaves. Second, we show that for bifurcating and all other full m-ary cladograms (in which every internal node has the same out-degree), one such Colless-like index is equivalent to the normalised reciprocal of Sackins index. Hence we both unify and generalise the two most popular existing tree balance indices. Our indices are intrinsically normalised and can be computed in linear time. We conclude that these more widely applicable indices have potential to supersede those in current use.


title: Conserved genomic landscapes of differentiation across Populus speciation continuum

abstract: Investigating genome-wide variation patterns along a speciation continuum is of central importance to understand the evolutionary processes contributing to lineage diversification. To identify which forces have shaped the genomic landscapes in Populus, we resequenced 201 whole-genomes from eight closely related species, with pairs of species at different stages along the speciation continuum. Using population structure and identity by descent analyses, we show extensive introgression between some species pairs, especially those with parapatric distributions. Inference of historical changes in effective population sizes support species-specific demographic trajectories, including recent population expansions in species characterized by broad present-day distributions. We observe highly conserved genomic landscapes, either focusing on within-species (genetic diversity:{pi} and recombination rate:{rho} ) or among-species variation (relative divergence: FST and absolute divergence: DXY). Independent of the stage across the divergence continuum, we recovered positive correlations between the pair{pi} and{rho} and the pair DXY and{rho} across all species pairs, which is consistent with a substantial contribution of linked selection in shaping these genomic landscapes. However, the positive correlations between{pi} and DXY became weaker as the overall divergence level (da) increased, suggesting that background selection is not the only factor at play. Positive correlations between FST and DXY in all species pairs, regardless of the rate of gene flow, also indicates the high FST could be due to divergent sorting of ancient polymorphism before speciation. Our study showcases the importance of investigating genomic patterns on multiple species across the speciation continuum to better understand the genomic landscapes of diversity and differentiation.


title: Rapid gene evolution in an ancient post-transcriptional and translationalregulatory system compensates for meiotic X chromosomal inactivation

abstract: It is conventionally assumed that conserved pathways evolve slowly with little participation of gene evolution. Nevertheless, it has been recently observed that young genes can take over fundamental functions in essential biological processes, for example, development and reproduction. It is unclear how newly duplicated genes are integrated into ancestral networks and reshape the conserved pathways of important functions. Here, we investigated origination and function of two autosomal genes that evolved recently in Drosophila: Poseidon and Zeus, which were created by RNA-based duplications from the X-linked CAF40, a subunit of the conserved CCR4-NOT deadenylase complex involved in post-transcriptional and translational regulation. Knockdown and knockout assays show that the two genes quickly evolved critically important functions in viability and male fertility. Moreover, our transcriptome analysis demonstrates that the three genes have a broad and distinct effect in the expression of hundreds of genes, with almost half of the differentially expressed genes being perturbed exclusively by one paralog, but not the others. Co-immunoprecipitation and tethering assays show that the CAF40 paralog Poseidon maintains the ability to interact with the CCR4-NOT deadenylase complex and might act in post-transcriptional mRNA regulation. The rapid gene evolution in the ancient post-transcriptional and translational regulatory system may be driven by evolution of sex chromosomes to compensate for the meiotic X chromosomal inactivation (MXCI) in Drosophila.


title: Population-level survey of loss-of-function mutations revealed that background dependent fitness genes are rare and functionally related in yeast

abstract: In natural populations, the same mutation can lead to different phenotypic outcomes due to the genetic variation that exists among individuals. Such genetic background effects are commonly observed, including in the context of many human diseases. However, systematic characterization of these effects at the species level is still lacking to date. Here, we sought to comprehensively survey background-dependent traits associated with gene loss-of-function (LoF) mutations in 39 natural isolates of Saccharomyces cerevisiae using a transposon saturation strategy. By analyzing the modeled fitness variability of a total of 4,469 genes, we found that 15% of them, when impacted by a LoF mutation, exhibited a significant gain- or loss-of-fitness phenotype in certain natural isolates compared to the reference strain S288C. Out of these 632 genetic background-dependent fitness genes identified, a total of 2/3 show a continuous variation across the population while 1/3 are specific to a single genetic background. Genes related to mitochondrial function are significantly overrepresented in the set of genes showing a continuous variation and display a potential functional rewiring with other genes involved in transcription and chromatin remodeling as well as in nuclear-cytoplasmic transport. Such rewiring effects are likely modulated by both the genetic background and the environment. While background-specific cases are rare and span diverse cellular processes, they can be functionally related at the individual level. All background-dependent fitness genes tend to have an intermediate connectivity in the global genetic interaction network and have shown relaxed selection pressure at the population level, highlighting their potential evolutionary characteristics.


title: The impact of genetic modifiers on variation in germline mutation rates within and among human populations

abstract: Mutation rates and spectra differ among human populations. Here, we examine whether this variation could be explained by evolution at mutation modifiers. To this end, we consider genetic modifier sites at which mutations, "mutator alleles", increase genome-wide mutation rates and model their evolution under purifying selection due to the additional deleterious mutations that they cause, genetic drift, and demographic processes. We solve the model analytically for a constant population size and characterize how evolution at modifier sites impacts variation in mutation rates within and among populations. We then use simulations to study the effects of modifier sites under a plausible demographic model for Africans and Europeans. When comparing populations that evolve independently, weakly selected modifier sites (2Nes {approx} 1), which evolve slowly, contribute the most to variation in mutation rates. In contrast, when populations recently split from a common ancestral population, strongly selected modifier sites (2Nes >> 1), which evolve rapidly, contribute the most to variation between them. Moreover, a modest number of modifier sites (e.g., 10 per mutation type in the standard classification into 96 types) subject to moderate to strong selection (2Nes > 1) could account for the variation in mutation rates observed among human populations. If such modifier sites indeed underlie differences among populations, they should also cause variation in mutation rates within populations and their effects should be detectable in pedigree studies.


title: Sex differences in deleterious genetic variants in a haplodiploid social insect

abstract: Deleterious variants are selected against but can linger in populations at low frequencies for long periods of time, decreasing fitness and contributing to disease burden in humans and other species. Deleterious variants occur at low frequency but distinguishing deleterious variants from low frequency neutral variation is challenging based on population genetics data. As a result, we have little sense of the number and identity of deleterious variants in wild populations. For haplodiploid species, it has been hypothesized that deleterious alleles will be directly exposed to selection in haploid males, but selection can be masked in diploid females due to partial or complete dominance, resulting in more efficient purging of deleterious mutations in males. Therefore, comparisons of the differences between haploid and diploid genomes from the same population may be a useful method for inferring rare deleterious variants. This study provides the first formal test of this hypothesis. Using wild populations of Northern paper wasps (Polistes fuscatus), we find that males have fewer overall variants, and specifically fewer missense and nonsense variants, than females from the same population. Allele frequency differences are especially pronounced for rare missense and nonsense mutations and these differences lead to a lower genetic load in males than females. Based on these data we estimate that a large number of highly deleterious mutations are segregating in the paper wasp population. Stronger selection against deleterious alleles in haploid males may have implications for adaptation in other haplodiploid insects and provides evidence that wild populations harbor abundant deleterious variants.


title: Expression variations in Ectodysplasin-A gene (eda) may contribute to morphological divergence of scales in Haplochromine cichlids

abstract:
BackgroundElasmoid scales are one of the most common dermal appendages and can be found in almost all species of bony fish differing greatly in their shape. Whilst the genetic underpinnings behind elasmoid scale development have been investigated, not much is known about the mechanisms involved in the shaping of scales. To investigate the links between gene expression differences and morphological divergence, we inferred shape variation of scales from two different areas of the body (anterior and posterior) stemming from ten haplochromine cichlid species from different origins (Lake Tanganyika, Lake Malawi, Lake Victoria and riverine). Additionally, we investigated transcriptional differences of a set of genes known to be involved in scale development and morphogenesis in fish.

ResultsWe found that scales from the anterior and posterior part of the body strongly differ in their overall shape, and a separate look on scales from each body part revealed similar trajectories of shape differences considering the lake origin of single investigated species. Above all, nine as well as 11 out of 16 target genes showed expression differences between the lakes for the anterior and posterior dataset, respectively. Whereas in posterior scales four genes (dlx5, eda, rankl and shh) revealed significant correlations between expression and morphological differentiation, in anterior scales only one gene (eda) showed such a correlation. Furthermore, eda displayed the most significant expression difference between species of Lake Tanganyika and species of the other two younger lakes. Finally, we found genetic differences in downstream regions of eda gene (e.g. in the eda-tnfsf13b inter-genic region) that are associated with observed expression differences. This is reminiscent of a genetic difference in the eda-tnfsf13b inter-genic region which leads to gain or loss of armour plates in stickleback.

ConclusionThese findings provide evidence for cross-species transcriptional differences of an important morphogenetic factor, eda, which is involved in formation of ectodermal appendages. These expression differences appeared to be associated with morphological differences observed in the scales of haplochromine cichlids indicating potential role of eda mediated signal in divergent scale morphogenesis in fish.


title: Adaptation in outbred sexual yeast is repeatable, polygenic, and favors rare haplotypes

abstract: We describe the results of a 200 generation Evolve and Resequence (E&R) study initiated from an outbred dipliod recombined synthetic base population derived from 18 genetically diverse founders. Replicate populations were maintained at large effective population sizes (>105 individuals), exposed to several different chemical challenges over 12 weeks of evolution, and whole-genome resequenced. Weekly forced outcrossing implies a per gene per cell-division recombination rate higher than that achieved in Drosophila E&R studies. In 55 sexual populations we observe large fitness gains and highly repeatable patterns of genome-wide haplotype change within each chemical challenge. There was little evidence for pervasive pleiotropy, as evidenced by patterns of haplotype change between drug treatments. Within treatment adaptation appears highly polygenic with almost the entire genome showing significant consistent haplotype change. Finally, adaptation was almost always associated with only one of the 18 founder alleles, suggesting selection primarily acts on rare variants private to a founder or haplotype blocks harboring multiple mutations. This observation contradicts the notion that adaptation is often due to subtle frequency shifts at intermediate frequency variants.


title: The transcriptomic signature of obligate parthenogenesis

abstract: Investigating the origin of parthenogenesis through interspecific hybridization can provide insight into how meiosis may be altered by genetic incompatibilities, which is fundamental for our understanding of the formation of reproductive barriers. Yet the genetic mechanisms giving rise to obligate parthenogenesis in eukaryotes remain understudied. In the microcrustacean Daphnia pulex species complex, obligately parthenogenetic (OP) isolates emerged as backcrosses of two cyclically parthenogenetic (CP) parental species, D. pulex and D. pulicaria, two closely related but ecologically distinct species. We examine the genome-wide expression in OP females at the early resting egg production stage, a life-history stage distinguishing OP and CP reproductive strategies, in comparison to CP females of the same stage from the two parental species. Our analyses of the expression data reveal that misregulated genes (underdominant and overdominant genes) are abundant in OP isolates, suggesting widespread regulatory incompatibilities between the parental species. More importantly, underdominant genes (i.e., genes with expression lower than both parentals) in the OP isolates are enriched in meiosis and cell-cycle pathways, indicating an important role of underdominance in the origin of obligate parthenogenesis. Furthermore, metabolic and biosynthesis pathways enriched with overdominant genes (i.e., expression higher than both parentals) are another genomic signature of OP isolates.


title: Effective Ensemble of Deep Neural Networks Predicts Neural Responses to Naturalistic Videos

abstract: This report provides a review of our submissions to the Algonauts Challenge 2021. In this challenge, neural responses in the visual cortex were recorded using functional neuroimaging when participants were watching naturalistic videos. The goal of the challenge is to develop voxel-wise encoding models which predict such neural signals based on the input videos. Here we built an ensemble of models that extract representations based on the input videos from 4 perspectives: image streams, motion, edges, and audio. We showed that adding new modules into the ensemble consistently improved our prediction performance. Our methods achieved state-of-the-art performance on both the mini track and the full track tasks.


title: Long-term precision editing of neural circuits using engineered gap junction hemichannels

abstract: The coordination of activity between brain cells is a key determinant of neural circuit function; nevertheless, approaches that selectively regulate communication between two distinct cellular components of a circuit, while leaving the activity of the presynaptic brain cell undisturbed remain sparse. To address this gap, we developed a novel class of electrical synapses by selectively engineering two connexin proteins found in Morone americana (white perch fish): connexin34.7 (Cx34.7) and connexin35 (Cx35). By iteratively exploiting protein mutagenesis, a novel in vitro assay of connexin docking, and computational modeling of connexin hemichannel interactions, we uncovered the pattern of structural motifs that broadly determine connexin hemichannel docking. We then utilized this knowledge to design Cx34.7 and Cx35 hemichannels that dock with each other, but not with themselves nor with other major connexins expressed in the mammalian central nervous system. We validated these hemichannels in vivo using C. elegans and mice, demonstrating that they can facilitate communication across neural circuits composed of pairs of genetically distinct cell types and modify behavior accordingly. Thus, we establish a potentially translational approach,  Long-term integration of Circuits using connexins (LinCx), for context-precise circuit-editing with unprecedented spatiotemporal specificity in mammals.


title: Long-term precision editing of neural circuits in mammals using engineered gap junction hemichannels


title: Epitopedia: identifying molecular mimicry of known immune epitopes

abstract: Upon infection, foreign antigenic proteins stimulate the hosts immune system to produce antibodies targeting the pathogen. These antibodies bind to regions on the antibody called epitopes. Structural similarity (molecular mimicry) of epitopes between an infecting pathogen and host proteins or other pathogenic proteins the host has previously encountered can impact the host immune response to the pathogen and may lead to cross-reactive antibodies. The ability to identify potential regions of molecular mimicry in a pathogen can illuminate immune effects which are especially important to pathogen treatment and vaccine design. Here we present Epitopedia, a software pipeline that facilitates the identification of regions that may exhibit potential three-dimensional molecular mimicry between an antigenic pathogen protein and known immune epitopes as catalogued by the immune epitope database (IEDB). Epitopedia is open-source software released under the MIT license and is freely available on GitHub, including a Docker container with all other software dependencies preinstalled. We performed an analysis describing how various secondary structure states, identity between pentapeptide pairs, and identity between the parent sequences of pentapeptide pairs affects RMSD. We found that pentapeptides pairs in a helical conformation had considerably lower RMSD values than those in Extended or Coil conformations. We also found that RMSD is significantly increased when pentapeptide pairs are from non-homologous sequences.


title: Epitopedia: identifying molecular mimicry between pathogens and known immune epitopes


title: From Alpha to Zeta: Identifying variants and subtypes of SARS-CoV-2 via clustering

abstract:
The availability of millions of SARS-CoV-2 sequences in public databases such as GISAID and EMBL-EBI (UK) allows a detailed study of the evolution, genomic diversity and dynamics of a virus like never before. Here we identify novel variants and sub-types of SARS-CoV-2 by clustering sequences in adapting methods originally designed for haplotyping intra-host viral populations. We asses our results using clustering entropy -- the first time it has been used in this context.

Our clustering approach reaches lower entropies compared to other methods, and we are able to boost this even further through gap filling and Monte Carlo based entropy minimization. Moreover, our method clearly identifies the well-known Alpha variant in the UK and GISAID datasets, but is also able to detect the much less represented (< 1% of the sequences) Beta (South Africa), Epsilon (California), Gamma and Zeta (Brazil) variants in the GISAID dataset. Finally, we show that each variant identified has high selective fitness, based on the growth rate of its cluster over time. This demonstrates that our clustering approach is a viable alternative for detecting even rare subtypes in very large datasets.


title: Microbial dark carbon fixation fueled by nitrate enrichment

abstract:
Anthropogenic nitrate amendment to coastal marine sediments can increase rates of heterotrophic mineralization and autotrophic dark carbon fixation (DCF). DCF may be favored in sediments where organic matter is biologically unavailable, leading to a microbial community supported by chemoautotrophy. Niche partitioning among DCF communities and adaptations for nitrate metabolism in coastal marine sediments remain poorly characterized, especially within salt marshes. We used genome-resolved metagenomics, phylogenetics, and comparative genomics to characterize the potential niche space, phylogenetic relationships, and adaptations important to microbial communities within nitrate enriched sediment. We found that nitrate enrichment of sediment from discrete depths between 0-25 cm supported both heterotrophs and chemoautotrophs that use sulfur oxidizing denitrification to drive the Calvin-Benson-Bassham (CBB) or reductive TCA (rTCA) DCF pathways. Phylogenetic reconstruction indicated that the nitrate enriched community represented a small fraction of the phylogenetic diversity contained in coastal marine environmental genomes, while pangenomics revealed close evolutionary and functional relationships with DCF microbes in other oligotrophic environments. These results indicate that DCF can support coastal marine microbial communities and should be carefully considered when estimating the impact of nitrate on carbon cycling in these critical habitats.

ImportanceSalt marshes store carbon at one of the fastest rates of any blue carbon system and buffer coastal marine waters from eutrophication. Dark carbon fixation (DCF) conducted by microbes within the sediment can influence the carbon storage capacity, but little is known about the ecology or genomic potential of these organisms. Our study identifies a potential niche space for several functionally distinct groups of chemoautotrophs which primarily use sulfur oxidizing denitrification to fuel DCF under high nitrate concentrations. These findings fill an important gap in our understanding of microbial contributions to carbon storage within salt marsh sediments and how this critical blue carbon system responds to anthropogenic nitrate enrichment.

category: ecology


title: Large scale function-based genome prospecting for industrial traits applied to 1,3-propanediol production.

abstract: Due to the success of next-generation sequencing, there has been a vast build-up of microbial genomes in the public repositories. FAIR genome prospecting of this huge genomic potential for biotechnological benefiting, require new efficient and flexible methods. In this study, Semantic Web technologies are applied to develop a function-based genome mining approach that follows a knowledge and discovery in database (KDD) protocol. Focusing on the industrial important trait of 1,3-propanediol (1,3-PD) production 187 new candidate species were identified. Furthermore, the genetic architecture of the particular trait was resolved, and persistent domains identified.


title: From Text-based Genome, Population Variations, and Transcriptome Datafiles to SQLite Database and Web Application: A Bioinformatical Study on Alfalfa

abstract: In this study, a web database application with the Flask framework was developed to implement three types of queries and visualize the results over a bioinformatical dataset from Alfalfa (Medicago sativa). A backend SQLite database was constructed from genome FASTA, population variations, transcriptome, and annotation files with extensions ".fasta", ".gff", "vcf", ".annotate", etc. Further, a supplementary command-line-based Java application was also developed for faster access to the database without direct SQL programming. Overall, Python, Java, and HTML were the main programming languages used in this application. Those scripts and the development procedures are valuable for bioinformaticians to build online databases from similar raw datasets of other species.


title: A murine model with JAK2V617F expression in both hematopoietic cells and vascular endothelial cells recapitulates the key features of human myeloproliferative neoplasm

abstract:
The myeloproliferative neoplasms (MPNs) are characterized by an expansion of the neoplastic hematopoietic stem/progenitor cells (HSPC) and an increased risk of cardiovascular complications. The acquired kinase mutation JAK2V617F is present in hematopoietic cells in a majority of patients with MPNs. Vascular endothelial cells (ECs) carrying the JAK2V617F mutation can also be detected in patients with MPNs. In this study, we show that a murine model with both JAK2V617F-bearing hematopoietic cells and JAK2V617F-bearing vascular ECs recapitulated all the key features of the human MPN disease, which include disease transformation from essential thrombocythemia to myelofibrosis, extramedullary splenic hematopoiesis, and spontaneous cardiovascular complications. During aging and MPN disease progression, there was a loss of both HSPC number and HSPC function in the marrow while the neoplastic hematopoiesis was relatively maintained in the spleen, mimicking the advanced phases of human MPN disease. Different vascular niche of the marrow and spleen could contribute to the different JAK2V617F mutant stem cell functions we have observed in this JAK2V617F-positive murine model. Compared to other MPN murine models reported so far, our studies demonstrate that endothelial dysfunction plays an important role in both the hematologic and cardiovascular abnormalities of MPN.

Key PointsO_LIA murine model in which JAK2V617F is expressed in both hematopoietic cells and ECs recapitulated the key features of the human MPN disease
C_LIO_LIDifferent vascular niche of the marrow and spleen could contribute to different JAK2V617F HSC functions during MPN disease progression
C_LI


title: JAK2V617F mutant megakaryocytes contribute to hematopoietic aging in a murine model of myeloproliferative neoplasm

abstract:
Megakaryocytes (MKs) is an important component of the hematopoietic niche. Abnormal MK hyperplasia is a hallmark feature of myeloproliferative neoplasms (MPNs). The JAK2V617F mutation is present in hematopoietic cells in a majority of patients with MPNs. Using a murine model of MPN in which the human JAK2V617F gene is expressed specifically in the MK lineage, we show that the JAK2V617F-bearing MKs promote hematopoietic stem cell (HSC) aging, manifesting as myeloid-skewed hematopoiesis with an expansion of CD41+ HSCs, a reduced engraftment and self-renewal capacity, and a reduced differentiation capacity. HSCs from 2yr old mice with JAK2V617F-bearing MKs were more proliferative and less quiescent than HSCs from age-matched control mice. Examination of the marrow hematopoietic niche reveals that the JAK2V617F-bearing MKs not only have decreased direct interactions with hematopoietic stem/progenitor cells during aging, but also suppress the vascular niche function during aging. Unbiased RNA expression profiling reveals that HSC aging has a profound effect on MK transcriptomic profiles, while targeted cytokine array shows that the JAK2V617F-bearing MKs can alter the hematopoietic niche through increased levels of pro-inflammatory and anti-angiogenic factors. Therefore, as a hematopoietic niche cell, MKs represent an important connection between the extrinsic and intrinsic mechanisms for HSC aging.

Significance StatementThe relative contribution of intrinsic and extrinsic mechanisms to HSC aging remains debated. We find that JAK2V617F mutant MKs can accelerate hematopoietic aging both directly (via decreased MK-HSC interaction) and indirectly (via suppressing vascular niche function). We also show that HSC aging has a profound effect on MK function. Our data suggest that, as a hematopoietic niche cell, MKs represent an important connection between HSC-intrinsic and HSC-extrinsic aging mechanisms.


title: A vimentin-targeting oral compound with host-directed antiviral and anti-inflammatory actions addresses multiple features of COVID-19 and related diseases

abstract:
Damage in COVID-19 results from both the SARS-CoV-2 virus and its triggered overreactive host immune responses. Therapeutic agents that focus solely on reducing viral load or hyperinflammation fail to provide satisfying outcomes in all cases. Although viral and cellular factors have been extensively profiled to identify potential anti-COVID targets, new drugs with significant efficacy remain to be developed. Here, we report the potent preclinical efficacy of ALD-R491, a vimentin-targeting small molecule compound, in treating COVID-19 through its host-directed antiviral and anti-inflammatory actions. We found that by altering the physical properties of vimentin filaments, ALD-491 affected general cellular processes as well as specific cellular functions relevant to SARS-CoV-2 infection. Specifically, ALD-R491 reduced endocytosis, endosomal trafficking, and exosomal release, thus impeding the entry and egress of the virus; increased the microcidal capacity of macrophages, thus facilitating the pathogen clearance; and enhanced the activity of regulatory T cells, therefore suppressing the overreactive immune responses. In cultured cells, ALD-R491 potently inhibited the SARS-CoV-2 spike protein and human ACE2-mediated pseudoviral infection. In aged mice with ongoing, productive SARS-CoV-2 infection, ALD-R491 reduced disease symptoms as well as lung damage. In rats, ALD-R491 also reduced bleomycin-induced lung injury and fibrosis. Our results indicate a unique mechanism and significant therapeutic potential for ALD-R491 against COVID-19. We anticipate that ALD-R491, an oral, fast-acting, and non-toxic agent targeting the cellular protein with multipart actions, will be convenient, safe, and broadly effective, regardless of viral mutations, for patients with early- or late-stage disease, post-COVID complications and other related diseases.

IMPORTANCEWith the Delta variant currently fueling a resurgence of new infections in the fully-vaccinated population, developing an effective therapeutic drug is especially critical and urgent in fighting COVID-19. In contrast to the many efforts to repurpose existing drugs or address only one aspect of COVID-19, we are developing a novel agent with first-in-class mechanism-of-actions that address both the viral infection and the overactive immune system in the pathogenesis of the disease. Unlike virus-directed therapeutics that may lose efficacy due to viral mutations and immunosuppressants that require ideal timing to be effective, this agent, with its unique host-directed antiviral and anti-inflammatory actions, can work against all variants of the virus, be effective during all stages of the disease, and even resolve post-disease damage and complications. A further development of the compound will provide an important tool in the fight against COVID-19, its complications, as well as future outbreaks of new viruses.


title: Mechanisms of ventricular arrhythmias elicited by coexistence of multiple electrophysiological remodeling in ischemia: a simulation study

abstract:
Myocardial ischemia, injury and infarction (MI) are the three stages of acute coronary syndrome (ACS). In the past two decades, a great number of studies focused on myocardial ischemia and MI individually, and showed that the occurrence of reentrant arrhythmias is often associated with myocardial ischemia or MI. However, arrhythmogenic mechanisms in the tissue with various degrees of remodeling in the ischemic heart have not been fully understood. In this study, biophysical detailed single-cell models of ischemia 1a, 1b, and MI were developed to mimic the electrophysiological remodeling at different stages of ACS. 2D tissue models with different distributions of ischemia and MI were constructed to investigate the mechanisms of initiation of reentrant waves during the progression of ischemia. Simulation results in 2D tissues showed that the vulnerable window (VW) in the tissue with multiple pathological conditions were determined by the VWs in the tissues with a single pathological condition. In different pathological conditions, action potential duration (APD) and the conduction velocity (CV) change differently. In the tissue with multiple pathological conditions, when the borders of different pathological conditions were perpendicular to the excitation wavefront, reentrant waves were mainly induced by the spatial heterogeneity of refractory periods due to the interaction of APD and CV along the wavefront. When the borders were parallel to the wavefront, the increased excitation threshold of MI region as well as the impaired excitability of ischemia region were the primary reason for the generation of reentry. Finally, the reentrant wave was observed in a 3D model with a scar reconstructed from MRI images of a MI patient. These comprehensive findings provide novel insights for understanding the arrhythmic risk during the progression of myocardial ischemia and highlight the importance of the multiple pathological stage in designing medical therapies for arrhythmia in ischemia.

Author summaryAbnormal initiation or conduction of electrical impulses may lead to cardiac arrhythmias, which are very important cause of sudden and early death in developed countries. In many cases, cardiac arrhythmias are accompanied by ventricular fibrillation sustained by re-entry. The occurrence of reentrant arrhythmias is often associated with acute coronary syndrome, including three phases of myocardial ischemia 1a, 1b, and infarction. Previous studies have made lots of efforts to unravel the mechanisms of the initiation and maintenance of reentry waves during myocardial ischemia or infarction. However, the mechanisms of initiation of reentrant waves in the tissue with multiple ischemic remodeling are not fully understood. Multi-scale computational models at the cellular, tissue, and organ levels were developed in this study. The main finding in this study is that reentrant waves in multiple pathological tissues were mainly induced by the spatial heterogeneity of refractory periods, when the borders of different pathological conditions were perpendicular to the wavefront of the excitation wave. In addition, the increased excitation threshold of MI region as well as the impaired excitability of ischemia region can induce the generation of reentry, when the borders were parallel to the excitation wavefront. This provides insights into mechanisms of ventricular arrhythmias elicited by coexistence of multiple electrophysiological remodeling in ischemia.

category: physiology


title: Biological research in Papua New Guinea: a publication activity analysis

abstract: The Nature Index and the Web of Science databases are used to analyse the publication patterns in life sciences in Papua New Guinea, aiming to identify the ways of improving the countrys research output.

category: scientific communication and education


title: Mitochondrial dysfunction compromises ciliary homeostasis in astrocytes

abstract: Astrocytes, often considered as secondary responders to neurodegenerative processes, are emerging as primary drivers of brain disease. The underlying pathogenic mechanisms are, however, insufficiently understood. Here we show that pathogenesis of mitochondrial spongiotic encephalopathy, a severe manifestation of mitochondrial brain diseases, involves abnormal maintenance of the astrocytic primary cilium, a major signaling organelle of a cell. We show that progressive respiratory chain deficiency in astrocytes activates FOXJ1 and RFX transcription factors and master regulators of motile ciliogenesis. Consequently, a wide aberrant nuclear expression program with FOXJ1 and RFX target genes, encoding motile cilia components, is induced in astrocytes. While the affected astrocytes still retain a single cilium, these organelles elongate and become remarkably distorted. Multiciliated ventricle-lining ependymal cells show no overt cilia morphology defects despite similar mitochondrial dysfunction. We propose that the chronic activation of the integrated mitochondrial stress response (ISRmt), specifically induced in astrocytes, drives anabolic metabolism and promotes ciliary growth. Collectively, our evidence indicate that 1) an active signaling axis exists between astrocyte mitochondria and primary cilia; 2) ciliary signaling is part of ISRmt in astrocytes; 3) metabolic ciliopathy is a novel pathomechanism for mitochondria-related neurodegenerative diseases.


title: The effects of the NMDAR co-agonist D-serine on the structure and function of the optic tectum

abstract:
The N-methyl-D-aspartate type glutamate receptor (NMDAR) is a molecular coincidence detector which converts correlated patterns of neuronal activity into cues for the structural and functional refinement of developing circuits in the brain. D-serine is an endogenous co-agonist of the NMDAR. In this study, we investigated the effects of potent enhancement of NMDAR-mediated currents by chronic administration of saturating levels of D-serine on the developing Xenopus retinotectal circuit. Chronic exposure to the NMDAR co-agonist D-serine resulted in structural and functional changes to the optic tectum. D-serine administration affected synaptogenesis and dendritic morphology in recently differentiated tectal neurons, resulting in increased arbor compaction, reduced branch dynamics, and higher synapse density. These effects were not observed in more mature neurons. Calcium imaging to examine retinotopic map organization revealed that tectal neurons of animals raised in D-serine had sharper visual receptive fields. These findings suggest that the availability of endogenous NMDAR co-agonists like D-serine at glutamatergic synapses may regulate the refinement of circuits in the developing brain.

Significance statementN-methyl-D-aspartate receptors (NMDAR) are implicated in activity-dependent circuit plasticity. We used administration of the NMDAR co-agonist D-serine to further examine the role of the NMDAR in circuit development in vivo. D-serine stabilised dendritic arbors specifically of recently differentiated neurons, promoted synaptogenesis, and led to sharper retinotopic receptive fields in the optic tectum. Together, these results support the idea that signaling in response to synaptic current through NMDARs promotes the maturation of developing brain circuits.


title: Increased 20-HETE signaling suppresses neurovascular coupling after ischemic stroke in regions beyond the infarct

abstract: Neurovascular coupling, the process by which neuronal activity elicits increases in the local blood supply, is impaired in stroke patients in brain regions outside the infarct. Such impairment may contribute to neurological deterioration over time, but its mechanism is unknown. Using the middle cerebral artery occlusion (MCAO) model of stroke, we show that neuronal activity-evoked capillary dilation is reduced by [~]75% in the intact cortical tissue outside the infarct border. This decrease in capillary responsiveness was not explained by a decrease in local neuronal activity or a loss of vascular contractility. Inhibiting synthesis of the vasoconstrictive molecule 20-HETE, either by inhibiting its synthetic enzyme CYP450 {omega}-hydroxylases or by increasing nitric oxide (NO), which is a natural inhibitor of {omega}-hydroxylases, rescued activity-evoked capillary dilation. The capillary dilation unmasked by inhibiting 20-HETE was dependent on PGE2 activation of EP4 receptors, a vasodilatory pathway previously identified in healthy animals. Cortical 20-HETE levels were increased following MCAO, in agreement with data from stroke patients. Inhibition of {omega}-hydroxylases normalized 20-HETE levels in vivo and increased cerebral blood flow in the peri-infarct cortex. These data identify 20-HETE-dependent vasoconstriction as a mechanism underlying neurovascular coupling impairment after stroke. Our results suggest that the brains energy supply may be significantly reduced after stroke in regions previously believed to be asymptomatic and that {omega}-hydroxylase inhibition may restore healthy neurovascular coupling post-stroke.