title: Investigation of the impact of stool collection methods on the metabolomics analysis/profiles of infant fecal samples

abstract: Metabolomic studies are important to understand microbial metabolism and interaction between the host and the gut microbiome. Although there have been efforts to standardize sample processing in metabolomic studies, infant samples are mostly disregarded. In birth cohort studies, the use of diaper liners is prevalent and its impact on fecal metabolic profile remains untested. In this study, we compared metabolite profiles of fecal samples collected as solid stool and those collected from stool saturated liner. One infants stool sample was collected in triplicate for solid stool and stool saturated liner. Comprehensive metabolomics analysis of the fecal samples was performed using NMR, UPLC and DI-MS. The total number, identities and concentrations of the metabolites were determined and compared between stool sample collection methods (stool vs. liner). The number and identity of metabolites did not differ between collection methods for NMR and DI-MS when excluding metabolites with a coefficient of variation (CV) > 40%. NMR analysis demonstrated lowest bias between collection methods, and lowest technical precision between triplicates of the same method followed by DI-MS then UPLC. Concentrations of many metabolites from stool and stool saturated liner differed significantly as revealed by Bland-Altman plots and t-tests. Overall, a mean bias of 10.2% in the Bland-Altman analysis was acceptable for some metabolites confirming mutual agreement but not for others with a wide range of bias (-97-117%). Consequently, stool and stool-saturated liner could be used interchangeably only for some select metabolite classes e.g. amino acids. Differences between the metabolomic profiles of solid stool samples and stool saturated liner samples for some important molecules e.g., ethanol, fumarate, short chain fatty acids and bile acids, indicate the need for standardization in stool collection method for metabolomic studies performed in infants.


title: Structure of the Shaker Kv channel and mechanism of slow C-type inactivation

abstract:
Voltage-activated potassium (Kv) channels open upon membrane depolarization and proceed to spontaneously inactivate. Inactivation controls neuronal firing rates and serves as a form of short-term memory, and is implicated in various human neurological disorders. Here, we use high-resolution cryo-electron microscopy and computer simulations to determine one of the molecular mechanisms underlying this physiologically crucial process. Structures of the activated Shaker Kv channel and of its W434F mutant in lipid bilayers demonstrate that C-type inactivation entails the dilation of the ion selectivity filter, and the repositioning of neighboring residues known to be functionally critical. Microsecond-scale molecular dynamics trajectories confirm these changes inhibit rapid ion permeation through the channel. This long-sought breakthrough establishes how eukaryotic K+ channels self-regulate their functional state through the plasticity of their selectivity filters.

One-Sentence SummaryStructures of the Shaker Kv channel reveal the mechanism of slow C-type inactivation involves dilation of the selectivity filter.

category: biophysics


title: Ligand binding remodels protein side chain conformational heterogeneity

abstract: While protein conformational heterogeneity plays an important role in many aspects of biological function, including ligand binding, its impact has been difficult to quantify. Macromolecular X-ray diffraction is commonly interpreted with a static structure, but it can provide information on both the anharmonic and harmonic contributions to conformational heterogeneity. Here, through multiconformer modeling of time- and space-averaged electron density, we measure conformational heterogeneity of 743 stringently matched pairs of crystallographic datasets that reflect unbound/apo and ligand-bound/holo states. When comparing the conformational heterogeneity of side chains, we observe that when binding site residues become more rigid upon ligand binding, distant residues tend to become more flexible, especially in non-solvent exposed regions. Among ligand properties, we observe increased protein flexibility as the number of hydrogen bonds decrease and relative hydrophobicity increases. Across a series of 13 inhibitor bound structures of CDK2, we find that conformational heterogeneity is correlated with inhibitor features and identify how conformational changes propagate differences in conformational heterogeneity away from the binding site. Collectively, our findings agree with models emerging from NMR studies suggesting that residual side chain entropy can modulate affinity and point to the need to integrate both static conformational changes and conformational heterogeneity in models of ligand binding.


title: Exposure to the gut microbiota from cigarette smoke-exposed mice exacerbates cigarette smoke extract-induced inflammation in zebrafish larvae

abstract: Cigarette smoke (CS)-induced inflammation leads to a range of diseases including chronic obstructive pulmonary disease and cancer. The gut microbiota is a major modifying environmental factor that determine the severity of cigarette smoke-induced pathology. Microbiomes and metabolites from CS-exposed mice exacerbate lung inflammation via the gut-lung axis of shared mucosal immunity in mice but these systems are expensive to establish and analyse. Zebrafish embryos and larvae have been used to model the effects of cigarette smoking on a range of physiological processes and offer an amenable platform for screening modifiers of cigarette smoke-induced pathologies with key features of low cost and rapid visual readouts. Here we exposed zebrafish larvae to cigarette smoke extract (CSE) and characterised a CSE-induced leukocytic inflammatory phenotype with increased neutrophilic and macrophage inflammation in the gut. The CSE-induced phenotype was exacerbated by co-exposure to microbiota from the faeces of CS-exposed mice, but not control mice. Microbiota could be recovered from the gut of zebrafish and studied in isolation in a screening setting. This demonstrates the utility of the zebrafish-CSE exposure platform for identifying environmental modifiers of cigarette smoking-associated pathology and demonstrates that the CS-exposed mouse gut microbiota potentiates the inflammatory effects of CSE across host species.

category: immunology


title: Cryptococcus neoformans releases proteins during intracellular residence that affect the outcome of the fungal-macrophage interaction

abstract: Cryptococcus neoformans is a facultative intracellular pathogen that can replicate and disseminate in mammalian macrophages. In this study, we analyzed fungal proteins identified in murine macrophage-like cells after infection with C. neoformans. To accomplish this, we developed a protocol to identify proteins released from cryptococcal cells inside macrophage-like cells; we identified 127 proteins of fungal origin in infected macrophage-like cells. Among the proteins identified was urease, a known virulence factor, and others such as transaldolase and phospholipase D, which have catalytic activities that could contribute to virulence. This method provides a straightforward methodology to study host-pathogen interactions. We chose to study further Yor1, a relatively uncharacterized protein belonging to the large family of ATP binding cassette transporter (ABC transporters). These transporters belong to a large and ancient protein family found in all extant phyla. While ABC transporters have an enormous diversity of functions across varied species, in pathogenic fungi they are better studied as drug efflux pumps. Analysis of C. neoformans yor1{Delta} strains revealed defects in non-lytic exocytosis and capsule size, when compared to wild-type strains. We detected no difference in growth rates, cell body size and vesicle secretion. Our results indicate that C. neoformans releases a large suite of proteins during macrophage infection, some of which can modulate fungal virulence and are likely to affect the fungal-macrophage interaction.

category: microbiology


title: Quantitative input-output dynamics of a c-di-GMP signal-transduction cascade in Vibrio cholerae

abstract: Bacterial biofilms are multicellular communities that collectively overcome environmental threats and clinical treatments. To regulate the biofilm lifecycle, bacteria commonly transduce sensory information via the second-messenger molecule cyclic diguanylate (c-di-GMP). Using experimental and modeling approaches, we quantitatively capture c-di-GMP signal transmission via the bifunctional polyamine receptor NspS-MbaA, from ligand binding to output, in the pathogen Vibrio cholerae. Upon binding of norspermidine or spermidine, NspS-MbaA synthesizes or degrades c-di-GMP, respectively, which in turn, drives alterations specifically to biofilm gene expression. A longstanding question is how output specificity is achieved via c-di-GMP, a diffusible molecule that regulates dozens of effectors. We show that NspS-MbaA signals locally to specific effectors, sensitizing V. cholerae to polyamines. However, local signaling is not required for specificity, as changes to global cytoplasmic c-di-GMP levels can selectively regulate biofilm genes. This work establishes the input-output dynamics underlying c-di-GMP signaling, which could be useful for developing bacterial manipulation strategies.


title: Non-target impacts of fungicide disturbance on phyllosphere yeasts in different crop species and management systems

abstract:
O_LIFungicides reduce fungal pathogen populations and are essential to food security. Fungicide disturbance of plant microbiomes has received limited attention. Understanding the impacts of fungicides on crop microbiomes in different cropping systems is vital to minimizing unintended consequences while maintaining their use for plant protection.
C_LIO_LIWe used amplicon sequencing of fungi and prokaryotes in maize and soybean microbiomes before and after foliar fungicide application in leaves and roots from plots under long-term no-till and conventional tillage managements. We examine fungicide disturbance and microbiome resilience across these treatments.
C_LIO_LIFoliar fungicides directly affected phyllosphere fungal communities, but not root fungal communities or prokaryote communities. Impacts on fungal phyllosphere composition and resiliency were management-dependent and lasted more than thirty days. Fungicides lowered pathogen abundance in maize and soybean and decreased the abundance of Tremellomycetes yeasts, especially the Bulleribacidiaceae, including core microbiome members.
C_LIO_LIFungicide application reduced network complexity in the soybean phyllosphere. Bulleribacidiaceae often co-occurred with Sphingomonas and Hymenobacter in control plots, but co-occurrences were altered in fungicide plots. Results indicate that foliar fungicides lower pathogen and non-target fungal abundance and may impact prokaryotes indirectly. No-till management was more resilient following fungicide disturbance and recovery.
C_LI


title: Facile synthesis of crosslinked Cu:ZnS-lignocellulose nanocomposite: a potent antifungal and antisporulant system against the tea pathogen Exobasidium vexans

abstract: The objective of the present study was to synthesize Cu doped ZnS nanocore crosslinked with lignocellulose (represented as Cu:ZnS-lignocellulose nanocomposite) for antifungal action against the devastating tea blister blight pathogen Exobasidium vexans. The characteristic features of the nanocomposite were analyzed via different physicochemical techniques like FTIR, XRD, XPS, SEM, SEM-EDX, Elemental mapping, PCS, and UV-PL studies. The FTIR and XPS investigations revealed the crosslinking between lignocellulose and the Cu:ZnS. The presence of lignocellulose was seen to attribute a potent antifungal efficacy, also enhancing the stability of the resulting nanocomposite in aqueous suspensions. The antifungal efficacy confirmed through disk diffusion and broth dilution assays have a maximum zone of inhibition of 1.75 cm2 and a MIC50 of 0.05 mg/ml against E. vexans. Additionally, the antisporulant activity was evident as the basidiospores failed to germinate in presence of the Cu:ZnS-lignocellulose nanocomposites. This shows potential for stemming the rapid infectivity of E. vexans by achieving disease inhibition at the early stage. Finally, the comparison with two commonly used commercial fungicides (copper oxychloride and fluconazole) demonstrated >10-fold higher antifungal activity for Cu:ZnS-lignocellulose nanocomposites.


title: The Integrator complex regulates microRNA abundance through RISC loading

abstract: MicroRNA (miRNA) homeostasis is crucial for the post-transcriptional regulation of their target genes and miRNA dysregulation has been linked to multiple diseases, including cancer. The molecular mechanisms underlying miRNA biogenesis from processing of primary miRNA transcripts to formation of mature miRNA duplex are well understood1-4. Loading of miRNA duplex into members of the Argonaute (Ago) protein family, representing the core of the RNA-induced silencing complex (RISC), is pivotal to miRNA-mediated gene silencing5-7. The Integrator complex has been previously shown to be an important regulator of RNA maturation, RNA polymerase II pause-release, and premature transcriptional termination8-11. Here, we report that loss of Integrator results in global diminution of mature miRNAs. By incorporating 4-Thiouridine (s4U) in nascent transcripts, we traced miRNA fate from biogenesis to stabilization and identified Integrator to be essential for proper miRNA assembly into RISC. Enhanced UV crosslinking and immunoprecipitation (eCLIP) of Integrator confirms a robust association with mature miRNAs. Indeed, Integrator potentiates Ago2-mediated cleavage of target RNAs. These findings highlight an essential role for Integrator in miRNA abundance and RISC function.

category: molecular biology


title: The GET pathway serves to activate Atg32-mediated mitophagy by ER targeting of the Ppg1-Far complex

abstract: Mitophagy removes defective or superfluous mitochondria via selective autophagy. In yeast, the pro-mitophagic protein Atg32 localizes to the mitochondrial surface and interacts with the scaffold protein Atg11 to promote degradation of mitochondria. Although Atg32-Atg11 interactions are thought to be stabilized by Atg32 phosphorylation, how this posttranslational modification is regulated remains obscure. Here we show that cells lacking the guided entry of tail-anchored proteins (GET) pathway exhibit reduced Atg32 phosphorylation and Atg32-Atg11 interactions, which can be rescued by additional loss of the ER-resident Ppg1-Far complex, a multi-subunit phosphatase negatively acting in mitophagy. In GET-deficient cells, Ppg1-Far is predominantly localized to mitochondria. An artificial ER anchoring of Ppg1-Far in GET-deficient cells significantly ameliorates defects in Atg32-Atg11 interactions and mitophagy. Moreover, disruption of GET and Msp1, an AAA-ATPase that extracts non-mitochondrial proteins localized to the mitochondrial surface, elicits synthetic defects in mitophagy. Collectively, we propose that the GET pathway mediates ER targeting of Ppg1-Far, thereby preventing dysregulated suppression of mitophagy activation.

category: cell biology


title: Comparative analysis of genome-encoded viral sequences reveals the evolutionary history of Flaviviridae.

abstract: The flaviviruses (family Flaviviridae) are a group of positive-strand RNA viruses, many of which pose serious risks to human health on a global scale. Here we use flavivirus-derived DNA sequences, identified in animal genomes, to reconstruct the long-term evolutionary history of the family Flaviviridae. We demonstrate that the family is at least 100 million years old and show that this timing can be integrated with dates inferred from co-phylogenetic analysis and paleontological records to produce a cohesive overview of flavivirus evolution in which the main subgroups originate early in animal evolution and broadly co-diverge with animal phyla. In addition, we show that the arthropod-borne  classical flaviviruses first evolved from tick-specific viruses, and only later adapted to become insect-borne. Our findings demonstrate that the biological properties of flaviviruses have been acquired over many millions of years, implying that broad-scale comparative analysis will reveal fundamental insights into flavivirus biology. Furthermore, we implement a novel approach to computational genomic studies of viruses that can support these efforts by enabling more efficient utilization of evolution-related domain knowledge in virus research.

category: evolutionary biology


title: High Resolution Ancestry Deconvolution for Next Generation Genomic Data

abstract: As genome-wide association studies and genetic risk prediction models are extended to globally diverse and admixed cohorts, ancestry deconvolution has become an increasingly important tool. Also known as local ancestry inference (LAI), this technique identifies the ancestry of each region of an individuals genome, thus permitting downstream analyses to account for genetic effects that vary between ancestries. Since existing LAI methods were developed before the rise of massive, whole genome biobanks, they are computationally burdened by these large next generation datasets. Current LAI algorithms also fail to harness the potential of whole genome sequences, falling well short of the accuracy that such high variant densities can enable. Here we introduce Gnomix, a set of algorithms that address each of these points, achieving higher accuracy and swifter computational performance than any existing LAI method, while also enabling portable models that are particularly useful when training data are not shareable due to privacy or other restrictions. We demonstrate Gnomix (and its swift phase correction counterpart Gnofix) on worldwide whole-genome data from both humans and canids and utilize its high resolution accuracy to identify the location of ancient New World haplotypes in the Xoloitzcuintle, dating back over 100 generations. Code is available at https://github.com/AI-sandbox/gnomix.

category: genomics


title: Symmetry reduction in a hyperpolarization-activated homotetrameric ion channel

abstract: Plants obtain nutrients from the soil via transmembrane transporters and channels in their root hairs, from which ions radially transport in towards the xylem for distribution across the plant body. We determined structures of the hyperpolarization-activated channel AKT1 from Arabidopsis thaliana, which mediates K+ uptake from the soil into plant roots. These structures of AtAKT1 embedded in lipid nanodiscs show that the channel undergoes a reduction of C4 to C2 symmetry, possibly to regulate its electrical activation.


title: Cryo-electron tomography reveals structural insights into the membrane binding and remodeling activity of dynamin-like EHDs

abstract: Dynamin-related Eps15-homology domain containing proteins (EHDs) oligomerize on membrane surfaces into filaments leading to membrane remodeling. EHD crystal structures in an open and a closed conformation were previously reported, but structural information on the membrane-bound EHD oligomeric structure has remained enigmatic. Consequently, mechanistic insight into EHD-mediated membrane remodeling is lacking. Here, by using cryo-electron tomography and subtomogram averaging, we determined the structure of an EHD4 filament on a tubular membrane template at an average resolution of 7.6 [A]. Assembly of EHD4 is mediated via interfaces in the G-domain and the helical domain. The oligomerized EHD4 structure resembles the closed conformation, where the tips of the helical domains protrude into the membrane. The variation in filament geometry and tube radius suggests the AMPPNP-bound filament has a spontaneous curvature of approximately 1/70 nm-1. Combining the available structural and functional data, we propose a model of EHD-mediated membrane remodeling.


title: Exosomal miR-769-5p confers cisplatin resistance and tumorigenesis in gastric cancer

abstract: Cisplatin resistance is the main cause of poor clinical prognosis in patients with gastric cancer (GC). Yet, the exact mechanism of cisplatin resistance remains unclear. Recent studies have suggested that exocrine miRNAs found in the tumor microenvironment participates in tumor metastasis and drug resistance. In this study, we discovered that cisplatin-resistant GC cells communicate with the tumor microenvironment by secreting microvesicles. The biologically active miR-769-5p can be integrated into exosomes and delivered to sensitive cells, thereby spreading cisplatin resistance. Mi769-5p was upregulated in GC tissues and enriched in the serum exosomes of cisplatin-resistant patients. Mechanistically, miR-769-5p promotes cisplatin resistance by targeting CASP9 so as to inhibit the downstream caspase pathway and promote the degradation of the apoptosis-related protein p53 through the ubiquitin-proteasome pathway. Targeting miR-769 with its antagonist to treat cisplatin-resistant GC cells can restore the cisplatin response, confirming that exosomal miR-769-5p can be a key regulator of cisplatin resistance in GC. Therefore, exosomal miR-769-5p derived from drug-resistant cells can be used as a potential therapeutic predictor of anti-tumor chemotherapy to enhance the effect of anti-cancer chemotherapy, which provides a new treatment option for GC.

category: cancer biology


title: Proton export drives the Warburg Effect

abstract: Aggressive cancers commonly ferment glucose to lactic acid at high rates, even in the presence of oxygen. This is known as aerobic glycolysis, or the "Warburg Effect". It is widely assumed that this is a consequence of the upregulation of glycolytic enzymes. Oncogenic drivers can increase the expression of most proteins in the glycolytic pathway, including the terminal step of exporting H+ equivalents from the cytoplasm. Proton exporters maintain an alkaline cytoplasmic pH, which can enhance all glycolytic enzyme activities, even in the absence of oncogene-related expression changes. Based on this observation, we hypothesized that increased uptake and fermentative metabolism of glucose could be driven by the expulsion of H+ equivalents from the cell. To test this hypothesis, we stably transfected lowly-glycolytic MCF-7, U2-OS, and glycolytic HEK293 cells to express proton exporting systems: either PMA1 (yeast H+-ATPase) or CAIX (carbonic anhydrase 9). The expression of either exporter in vitro enhanced aerobic glycolysis as measured by glucose consumption, lactate production, and extracellular acidification rate. This resulted in an increased intracellular pH, and metabolomic analyses indicated that this was associated with an increased flux of all glycolytic enzymes upstream of pyruvate kinase. These cells also demonstrated increased migratory and invasive phenotypes in vitro, and these were recapitulated in vivo by more aggressive behavior, whereby the acid-producing cells formed higher grade tumors with higher rates of metastases. Neutralizing tumor acidity with oral buffers reduced the metastatic burden. Therefore, cancer cells with increased H+ export increase intracellular alkalization, even without oncogenic driver mutations, and this is sufficient to alter cancer metabolism towards a Warburg phenotype.


title: GPER-dependent estrogen signaling increases cardiac GCN5L1 expression and MCAD activity

abstract: Reversible lysine acetylation regulates the activity of cardiac metabolic enzymes, including those controlling fuel substrate metabolism. Mitochondrial-targeted GCN5L1 and SIRT3 have been shown to regulate the acetylation status of mitochondrial enzymes, which results in alterations to the relative oxidation rates of fatty acids, glucose, and other fuels for contractile activity. However, the role that lysine acetylation plays in driving metabolic differences between male and female hearts is not currently known. In this study, we report that estrogens induce the expression of GCN5L1 via GPER agonism in cardiac cells, which increases the enzymatic activity and acetylation status of the fatty acid oxidation enzyme medium chain acyl-CoA dehydrogenase (MCAD).

category: physiology


title: miR2105 regulates ABA biosynthesis via OsbZIP86-OsNCED3 module to contribute to drought tolerance in rice

abstract:
Induced abscisic acid (ABA) biosynthesis plays an important role in plant tolerance to abiotic stresses, including drought, cold and salinity. However, regulation pathway of the ABA biosynthesis in response to stresses is unclear. Here, we identified a rice miRNA, osa-miR2105 (miR2105), which plays a crucial role in ABA biosynthesis under drought stress. Analysis of expression, transgenic rice and cleavage site showed that OsbZIP86 is a target gene of miR2105. Subcellular localization and luciferase activity assays showed that OsbZIP86 is a nuclear transcription factor. In vivo and in vitro analyses showed that OsbZIP86 directly binds to the promoter of OsNCED3, and interacts with OsSAPK10, resulting in enhanced-expression of OsNCED3. Transgenic rice plants with knock-down of miR2105 or overexpression of OsbZIP86 showed higher ABA content, more tolerance to drought, a lower rate of water loss, more stomatal closure than wild type rice ZH11 under drought stress. These rice plants showed no penalty with respect to agronomic traits under normal conditions. By contrast, transgenic rice plants with miR2105 overexpression, OsbZIP86 downregulation, or OsbZIP86 knockout displayed less tolerance to drought stress and other phenotypes. Collectively, our results show that a regulatory network of  miR2105-OsSAPK10/OsbZIP86-OsNCED3 control ABA biosynthesis in response to drought stress.

One-sentence summary miR2105-OsbZIP86-OsNCED3 module plays crucial role in mediating ABA biosynthesis to contribute to drought tolerance with no penalty with respect to agronomic traits under normal conditions.


title: Hematopoietic recovery after transplantation is primarily derived from the stochastic contribution of hematopoietic stem cells

abstract: Reconstitution after hematopoietic stem cell (HSC) transplantation is assumed to occur in two distinct phases: initial recovery mediated by short-term progenitors and long-term repopulation by multipotent HSCs which do not contribute to hematopoietic reconstitution during the first 6-9 months. We have previously reported the transplantation and exclusive engraftment of the HSC-enriched CD34+CD45RA-CD90+ phenotype in a nonhuman primate model. Here, we closely followed the clonal diversity and kinetics in these animals. Enhanced sampling and high density clonal tracking within the first 3 month revealed that multipotent HSCs actively contributed to the early phases of neutrophil recovery and became the dominant source for blood cells as early as 50 days after transplant. Longitudinal changes in clonal diversity supported a stochastic engraftment of HSCs with the majority of HSCs clones vanishing early during neutrophil recovery and a smaller fraction of HSC clones expanding into bigger pools to support long-term hematopoiesis. In contrast to the bi-phasic model, we propose that hematopoietic recovery after myeloablation and transplantation is primarily derived from HSCs in a stochastic manner rather than in two phases by independent cell populations.


title: A bipartite chromatophore transit peptide and N-terminal processing of protein in the Paulinella chromatophore

abstract:
The cercozoan amoeba Paulinella chromatophora contains photosynthetic organelles - termed chromatophores - that evolved from a cyanobacterium [~]100 million years ago, independently from plastids in plants and algae. Despite its more recent origin, at least one third of the chromatophore proteome consists of nucleus-encoded proteins that are imported by an unknown mechanism across the chromatophore double envelope membranes. Chromatophore-targeted proteins fall into two classes. Proteins exceeding 250 amino acids carry a conserved N-terminal sequence extension, termed the  chromatophore transit peptide (crTP), that is presumably involved in guiding these proteins into the chromatophore. Short imported proteins do not carry discernable targeting signals. To explore whether the import of protein is accompanied by their N-terminal processing, here we used a mass spectrometry-based approach to determine protein N-termini in Paulinella chromatophora and identified N-termini of 208 chromatophore-localized proteins. Our study revealed extensive N-terminal modifications by acetylation and proteolytic processing in both, the nucleus and chromatophore-encoded fraction of the chromatophore proteome. Mature N-termini of 37 crTP-carrying proteins were identified, of which 30 were cleaved in a common processing region. Our results imply that the crTP mediates trafficking through the Golgi, is bipartite and surprisingly only the N-terminal third ( part 1) becomes cleaved upon import, whereas the rest ( part 2) remains at the mature proteins. In contrast, short imported proteins remain largely unprocessed. Finally, this work sheds light on N-terminal processing of proteins encoded in an evolutionary-early-stage photosynthetic organelle and suggests host-derived post-translationally acting factors involved in dynamic regulation of the chromatophore-encoded chromatophore proteome.

One sentence summaryProteins targeted to the evolutionary-early-stage photosynthetic organelle of Paulinella carry a bipartite N-terminal targeting sequence that is only partially removed upon protein import.


title: DNA motif analysis of shear stress responsive endothelial enhancers reveals differential association of KLF and ETV/ETS binding sites with gained and lost enhancers

abstract: Endothelial cells (EC) lining blood vessels are exposed to mechanical forces, such as shear stress exerted by the flowing blood. These forces control many aspects of EC biology, including vascular tone, cell migration and proliferation in addition to cell size and shape. Despite a good understanding of the genes and signaling pathways responding to shear stress, our insights into the transcriptional regulation of these responses is much more limited. In particular, we do not know the different sets of regulatory elements (enhancers) that might control increases or decreases in gene expression. Here, we set out to study changes in the chromatin landscape of human umbilical vein endothelial cells (HUVEC) exposed to laminar shear stress. To do so, we performed ChIP-Seq for H3K27 acetylation, indicative of active enhancer elements and ATAC-Seq to mark regions of open chromatin in addition to RNA-Seq on HUVEC exposed to 6 hours of laminar shear stress. Our results show a correlation of gained and lost enhancers with up- and downregulated genes, respectively. DNA motif analysis revealed an over-representation of KLF transcription factor (TF) binding sites in gained enhancers, while lost enhancers contained more ETV/ETS motifs. We validated a subset of flow responsive enhancers using luciferase-based reporter constructs and CRISPR-Cas9 mediated genome editing. Lastly, we characterized shear stress responsive genes in ECs of zebrafish embryos using RNA-Seq. Together, our results reveal the presence of shear stress responsive DNA regulatory elements and lay the groundwork for the future exploration of these elements and the TFs binding to them in controlling EC biology.


title: Role of Disulfide Bonds in Membrane Partitioning of a Viral Peptide

abstract: The importance of disulfide bond in mediating viral peptide entry into host cells is well known. In the present work, we elucidate the role of disulfide (SS) bond in partitioning mechanism of membrane active Hepatitis A Virus-2B (HAV-2B) peptide, which harbours three cysteine residues promoting formation of multiple SS-bonded states. The inclusion of SS-bond not only results in a compact conformation but also induces distorted -helical hairpin geometry in comparison to SS-free state, resulting in reduced hydrophobic exposure. Owing to this, the partitioning of HAV-2B peptide is completely or partly abolished. In a way, the disulfide bond regulates the partitioning of HAV-2B peptide, such that the membrane remodelling effects of this viral peptide are significantly reduced. The current findings may have potential implications in drug designing, targeting the HAV-2B protein by promoting disulfide bond formation within its membrane active region.


title: LM-GVP: A Generalizable Deep Learning Framework for Protein Property Prediction from Sequence and Structure

abstract: Proteins perform many essential functions in biological systems and can be successfully developed as bio-therapeutics. It is invaluable to be able to predict their properties based on a proposed sequence and structure. In this study, we developed a novel generalizable deep learning framework, LM-GVP, composed of a protein Language Model (LM) and Graph Neural Network (GNN) to leverage information from both 1D amino acid sequences and 3D structures of proteins. Our approach outperformed the state-of-the-art protein LMs on a variety of property prediction tasks including fluorescence, protease stability, and protein functions from Gene Ontology (GO). We also illustrated insights into how a GNN prediction head can guide the protein LM to better leverage structural information. We envision that our deep learning framework will be generalizable to many protein property prediction problems to greatly accelerate protein engineering and drug development.

category: synthetic biology


title: Identification of topoisomerase as a precision-medicine target in chromatin reader SP140-driven Crohns disease

abstract: How mis-regulated chromatin directly impacts human immunological disease is poorly understood. Speckled Protein 140 (SP140) is an immune-restricted PHD and bromodomain-containing chromatin  reader whose loss-of-function associates with Crohns disease (CD), multiple sclerosis (MS) and chronic lymphocytic leukemia (CLL). However, mechanisms underlying SP140-driven pathogenicity and therapeutic approaches that rescue SP140 remain unexplored. Using a global proteomic strategy, we identified SP140 as a repressor of topoisomerases (TOP) that maintains heterochromatin and immune cell fate. In humans and mice, SP140 loss resulted in unleashed TOP activity, genome instability, severely compromised lineage-defining and microbe-inducible innate transcriptional programs and defective bacterial killing. Pharmacological inhibition of TOP1 or TOP2 rescued these defects. Furthermore, exacerbated colitis was restored with TOP1 or TOP2 inhibitors in Sp140-/- mice, but not wild-type mice, in vivo. Collectively, we identify SP140 as a repressor of topoisomerases and reveal repurposing of TOP inhibition as a precision strategy for reversing SP140-driven immune disease.


title: YfiB and the tripartite signaling system YfiBNR is involved in virulence of the gut invasive pathogen Shigella flexneri.

abstract: The intracellular pathogen Shigella flexneri, which is the causative agent of bacillary dysentery, significantly influences the worldwide implication of diarrheal infections, consequentially causing about 1.1 million deaths each year. Due to a non-availability of an authorized vaccine and the upsurge of multidrug resistance amongst Shigella strains, there has been a huge demand for further genetic analyses which could help in the advancement of new/improved drugs, and finding vaccine candidates against the pathogen. Whilst many features about the invasion of colonic cells by Shigella have been identified, fundamental gaps in information concerning in what way the bacteria transit, survive, and control gene expression, remain. The present study aims to illustrate the role of yfiB gene in Shigella virulence, which is a part of the periplasmic YfiBNR tripartite signaling system. This system is involved in the regulation of cyclic-di-GMP levels inside the bacterial cells, which is a vital messenger molecule impacting varied cellular processes such as biofilm formation, cytotoxicity, motility, synthesis of exopolysaccharide, and other virulence mechanisms like adhesion and invasion of the bacteria. Through a combination of genetic, biochemical, and virulence assays, we show how knocking out the yfiB gene can disrupt the entire YfiBNR system and affect biofilm formation, bacterial invasion, host-surface attachment, and the overall virulence of Shigella. This study eventually improves our understanding of the in-vivo persistence and survival of Shigella, brings light to the c-di-GMP lead regulation of Shigella virulence, and provides a prospective new target to design anti-infective drugs and vaccines against S. flexneri and other bacterial pathogens.


title: YfiB: An Outer Membrane Protein Involved in the Virulence of Shigella flexneri


title: A uniform benchmark for testing ssrA-derived degrons in the Escherichia coli ClpXP pathway

abstract:
The ssrA degron is commonly used in fusion proteins to control protein stability in bacteria or as an interaction module. These applications often rely on the modular activities of the ssrA tag in binding to the SspB adaptor and in engaging the ClpXP protease. However, a comparison of these activities for a substantial standard set of degron variants has not been conducted previously, which may hinder developments of new variants optimized exclusively for one application. Here, we strive to establish a benchmark that will facilitate the comparison of ssrA variants under uniform conditions. In our workflow, we included methods for expression and purification of ClpX, ClpP, SspB and eGFP-degrons, assays of ClpX ATPase activity, of eGFP-degron binding to SspB and for measuring eGFP-degron degradation in vitro and in vivo. Using uniform, precise and sensitive methods under the same conditions on a range of eGFP-degrons allowed us to determine subtle differences in their properties that can affect their potential applications. Our findings can serve as a reference and a resource for developing targeted protein degradation approaches.

SUMMARYThis work lays standards for assays used to compare engagement of SspB and ClpXP by a set of ssrA-derived degrons that can be used to fine-tune tools for protein stability control.

category: biochemistry


title: Zika virus causes placental pyroptosis and associated adverse fetal outcomes by activating GSDME

abstract:
Zika virus (ZIKV) can be transmitted from mother to fetus during pregnancy, causing adverse fetal outcomes. Several studies have indicated that ZIKV can damage the fetal brain directly; however, whether the ZIKV-induced maternal placental injury contribute to adverse fetal outcomes are sparsely defined. Here, we discovered that ZIKV causes the pyroptosis of placental cells by activating the executor Gasdermin E (GSDME) in vitro and in vivo. Mechanistically, caspase-8 undergoes activation upon the recognition of 5 untranslated region of viral RNA by RIG-I, followed by the stimulation of caspase-3 to ultimately escalate the GSDME cleavage. Further analyses revealed that the ablation of GSDME in ZIKV-infected pregnant mice attenuates placental pyroptosis, which consequently confers protection against adverse fetal outcomes. In conclusion, our study unveils a novel mechanism of ZIKV-induced adverse fetal outcomes via causing placental cell pyroptosis, which could be employed for developing new therapies for ZIKV-associated diseases.

Significance statementSeveral studies have elucidated the link between ZIKV infection and congenital ZIKV syndroms (CZS), but the pathogenesis yet needs further study. Here, we reported a novel pathogenic mechanism of ZIKV which leads to pyroptosis of placental cells through activating the pyroptotic executor GSDME, rather than GSDMD. Upon ZIKV infection, GSDME-mediated pyroptosis damages the structure and function of the placenta, thereby affecting the development of the fetus and contributing to the adverse fetal outcomes. Our study highlights the importance of pyroptotic executor GSDME in regulate ZIKV pathogenicity and further confirms that placental injury caused by ZIKV infection is a key factor for CZS.


title: Galvanic current activates the NLRP3 inflammasome to promote type I collagen production in tendon

abstract: The NLRP3 inflammasome coordinates inflammation in response to different pathogen- and damage-associated molecular patterns, being implicated in different infectious, chronic inflammatory, metabolic and degenerative diseases. In chronic tendinopathies lesions, different non-resolving mechanisms produce a degenerative condition that impairs tissue healing, complicating their clinical management. Percutaneous needle electrolysis consist in the application of a galvanic current and is emerging as a novel treatment for tendinopathies. Here we found that galvanic current activates the NLRP3 inflammasome and the induction of an inflammatory response promoting a collagen-mediated regeneration of the tendon. This study establish the molecular mechanism of percutaneous electrolysis for the treatment of chronic lesions and the beneficial effects of an induced inflammasome-related response.


title: Polygenic signals of sexually antagonistic selection in contemporary human genomes

abstract: Mutations that increase fitness in one sex may decrease fitness in the other. Such "sexually antagonistic" (SA) genetic variants can constrain adaptation and increase variability for fitness components (e.g., survival, fertility, and disease susceptibility). However, detecting SA selection in genomes is immensely challenging, as it requires prohibitively large datasets that combine genomic sequences with individual fitness measurements. Here, we use genotypic and reproductive success data from [~]250,000 UK Biobank individuals to comprehensively assess the extent of SA genetic variation in humans. We first develop new theoretical models for signals of SA selection spanning a full generational life cycle--including SA polymorphisms affecting survival, reproductive success and overall fitness. Comparing our models with UK Biobank data, we uncover multiple empirical signals of polygenic SA selection, including sex-differential effects of genetic variants on each fitness component, and positive correlations between sex-differential effects and minor allele frequencies. We show that these signals cannot be explained by simple models of sex differences in purifying selection, or by potential confounders such as population structure and sequence mapping errors. We further show that candidate SA sites disproportionately affect functional genomic regions, including polymorphisms associated with quantitative traits and disease. Finally, we examine historical evolutionary processes affecting candidate SA sites, which are consistent with the drift-dominated dynamics predicted by previous theory. Overall, our results support SA genomic variation in humans and highlight its broader functional and evolutionary consequences.


title: Signatures of visuospatial information foraging during learning of complex environments

abstract: Many real world environments contain latent features that animals must learn in order to accomplish their goals. Animals often learn these environments over sequences of choices, but the behavioral mechanisms underlying this complex cognitive competence remain poorly characterized. We investigate this sophisticated behavior in two primate species, humans and macaque monkeys, with a task in which subjects searched for shapes hidden on a grid. Both primate species were adept learners, quickly learning the subset of shapes out of the numerous possibilities. Subjects tended to select tiles earlier in trials that were informative in the past about the hidden shape than those that were rewarding. In addition, we found a surprising signature of foraging behavior over sequences of choices during trials, with human subjects searching local areas of the board until information dropped below the average across all choices, at which time they jumped to a different part of the board. This pattern of choices was not evident for rewards in humans. In contrast, the sequences of choices of monkeys were equally well-described as information or reward foraging. Finally, the rate at which humans learned shapes could be predicted by how well their choice sequences matched foraging behavior. These findings suggest that humans are more tuned to the search for information than reward than monkeys and that foraging competence predicts the capacity to learn complex environments.

category: animal behavior and cognition


title: Locally adapted oak populations along an elevation gradient display different molecular strategies to regulate bud phenology.

abstract:
Research conductedWith the ongoing global warming, there are serious concerns about the persistence of locally adapted populations. Indeed, with the raising of temperature, the phenological cycle of tree species may be strongly affected since higher winter temperatures may have a negative impact on endodormancy release if chilling requirements are not fulfilled during winter and late frost in spring may expose trees if buds flush too early. Thus, Environmental gradients (showing continuous variations of environmental conditions) constitute a design of choice to analyze the effect of winter dormancy in locally adapted population.

MethodsIn the present study, we used an elevation gradient in the Pyrenees to explore the gene expression network involved in dormancy regulation in natural populations of sessile oak locally adapted to temperature. Terminal buds were harvested during dormancy induction and release at different elevations. Then, gene expression was quantified using RNAseq and we used a likelihood ratio test to identify genes displaying significant dormancy, elevation or dormancy-by-elevation interaction effects.

Key resultsOur results highlight molecular processes in locally adapted populations along this elevation cline, and made it possible to identify key dormancy-by-elevation responsive genes revealing that locally adapted populations have evolved distinct molecular strategies to adapt their bud phenology in response to environmental variation (i.e. temperature).

category: plant biology


title: Sugar beet cold-induced PMT5a and STP13 carriers are poised for taproot proton-driven plasma membrane sucrose and glucose import

abstract:
O_LIAs the major sugar-producing crop in the northern hemisphere, sugar beet taproots store sucrose at a concentration of about 20 %. While the vacuolar sucrose loader TST has already been identified in the taproot, sugar transporters mediating sucrose uptake across the plasma membrane of taproot parenchyma cells remained unknown.
C_LIO_LIWe electrophysiologically examined taproots for proton-coupled sugar uptake and identified potentially involved transporters by transcriptomic profiling. After cloning, the transporter features were studied in the heterologous Xenopus laevis oocyte expression system using the two-electrode voltage clamp technique. Insights into the structure were gained by 3D homology modeling.
C_LIO_LIAs with glucose, sucrose stimulation of taproot parenchyma cells caused inward H+-fluxes and plasma membrane depolarization, indicating a sugar/proton symport mechanism. As one potential candidate for sugar uploading, the BvPMT5a was characterized as a H+-driven low-affinity glucose transporter, which does not transport sucrose. BvSTP13 operated as a high-affinity H+/sugar symporter, transporting glucose and to some extent sucrose due to a binding cleft plasticity. Both transporter genes were upregulated upon cold exposure, with the transport capacity of BvSTP13 being more cold-resistant than BvPMT5a.
C_LIO_LIIdentification of BvPMT5a and BvSTP13 as taproot sugar transporters could improve breeding of cold-tolerant sugar beet to provide a sustainable energy crop.
C_LI


title: Using all gene families vastly expands data available for phylogenomic inference in primates

abstract: Traditionally, single-copy orthologs have been the gold standard in phylogenomics. Most phylogenomic studies identify putative single-copy orthologs by using clustering approaches and retaining families with a single sequence from each species. However, this approach can severely limit the amount of data available by excluding larger families. Recent methodological advances have suggested several ways to include data from larger families. For instance, tree-based decomposition methods facilitate the extraction of orthologs from large families. Additionally, several popular methods for species tree inference appear to be robust to the inclusion of paralogs, and hence could use all of the data from larger families. Here, we explore the effects of using all families for phylogenetic inference using genomes from 26 primate species. We compare single-copy families, orthologs extracted using tree-based decomposition approaches, and all families with all data (i.e., including orthologs and paralogs). We explore several species tree inference methods, finding that across all nodes of the tree except one, identical trees are returned across nearly all datasets and methods. As in previous studies, the relationships among Platyrrhini remain contentious; however, the tree inference methods matter more than the dataset used. We also assess the effects of each dataset on branch length estimates, measures of phylogenetic uncertainty and concordance, and in detecting introgression. Our results demonstrate that using data from larger gene families drastically increases the number of genes available for phylogenetic inference and leads to consistent estimates of branch lengths, nodal certainty and concordance, and inferences of introgression.


title: Over-expression of the brassinosteroid gene TaDWF4 increases wheat productivity under low and sufficient nitrogen through enhanced carbon assimilation

abstract: There is a strong pressure to reduce nitrogen (N) fertiliser inputs while maintaining or increasing current cereal crop yields. Brassinosteroids, (BR), are a group of phytohormones essential for plant growth and development, that have been demonstrated to regulate several agronomic traits. DWF4 encodes a cytochrome P450 that catalyses a rate-limiting step in BR synthesis. We show that overexpression of the dominant shoot expressed homoeologue TaDWF4-B in wheat can increase plant productivity by up to 105% under a range of N levels on marginal soils, resulting in increased N use efficiency (NUE). We show that a two to four-fold increase in TaDWF4 transcript levels enhances the responsiveness of genes regulated by N. The productivity increases seen were primarily due to the maintenance of photosystem II operating efficiency and carbon assimilation in plants when grown under limiting N conditions and not an overall increase in photosynthesis capacity. The increased biomass production and yield per plant in TaDWF4 OE lines could be linked to modified carbon partitioning and changes in expression pattern of the growth regulator Target Of Rapamycin, offering a route towards breeding for sustained yield and lower N inputs.


title: Exploring neural correlates of behavioral and academic resilience among children in poverty

abstract: Children in poverty must contend with systems that do not meet their needs. We explored what, at a neural level, helps explain childrens resilience in these contexts. Lower coupling between lateral frontoparietal network (LFPN) and default mode network (DMN)--linked, respectively, to externally- and internally-directed thought--has previously been associated with better cognitive performance. However, we recently found the opposite pattern for children in poverty. Here, we probed ecologically-valid assessments of performance. In a pre-registered study, we investigated trajectories of network coupling over ages 9-13 and their relation to school grades and attention problems. We analyzed longitudinal data from ABCD Study (N=8366 children at baseline; 1303 below poverty). The link between cognitive performance and grades was weaker for children in poverty, highlighting the importance of ecologically-valid measures. As predicted, higher LFPN-DMN connectivity was linked to worse grades and attentional problems for children living above poverty, while children below poverty showed opposite tendencies. This interaction between LFPN-DMN connectivity and poverty related to childrens grades two years later; however, it was attenuated when controlling for baseline grades and was not related to attention longitudinally. Together, these findings suggest network connectivity is differentially related to performance in real-world settings for children above and below poverty.

category: neuroscience


title: Density-independent processes decouple component and ensemble density feedbacks

abstract: Analysis of long-term trends in abundance provide insights into population dynamics. Population growth rates are the emergent interplay of fertility, survival, and dispersal, but the density feedbacks on some vital rates (component) can be decoupled from density feedback on population growth rates (ensemble). However, the mechanisms responsible for this decoupling are poorly understood. We simulated component density feedbacks on survival in age-structured populations of long-living vertebrates and quantified how imposed nonstationarity (density-independent mortality and variation in carrying-capacity) modified the ensemble feedback signal estimated from logistic-growth models to the simulated abundance time series. The statistical detection of ensemble density feedback was largely unaffected by density-independent processes, but catastrophic and proportional mortality eroded the effect of density-dependent survival on ensemble-feedback strength more strongly than variation in carrying capacity. Thus, phenomenological models offer a robust approach to capture density feedbacks from nonstationary census data when density-independent mortality is low.

category: ecology


title: The origin and evolution of pollen transport in bees (Hymenoptera: Anthophila)

abstract: The ability to transport pollen from flowers back to the nest represents a key innovation in the evolution of bees from predatory wasp ancestors. Currently, the origin and evolution of pollen transport remains unsettled. Older hypotheses proposed that crop transport was the original mode of pollen transport, but more recent molecular phylogenies have cast doubt on that view. Instead, more recent hypotheses contend that external transport of dry pollen is ancestral in bees. Here, I propose a new hypothesis to explain the origin and subsequent evolution of pollen transport in bees. I propose that pollen transport arose from adult pollen-feeding behavior and that internal transport of pollen is ancestral in bees. This then led to the evolution of external moist transport, which first required a transition step whereby pollen is temporarily accumulated on the venter on a patch of specialized hairs. Finally, external glazed and dry transport evolved from external moist pollen transport, and the evolution of dry transport led to changes in the location of scopae from the original location on the hind tibia and basitarsus. I illustrate many of these hypothetical evolutionary steps using modern-day bee behavior as an example, with a particular focus on the bee Perdita tortifoliae. Examination of the evolution of pollen transport of pollen wasps (subfamily Masarinae) reveals that they have undergone a parallel evolutionary change. Overall, I lay out a broad hypothetical framework to explain the origin and subsequent evolution of pollen transport in bees. This marks a return to the earlier hypothesis that crop transport is ancestral, and it also represents the first in-depth hypothesis to explain how external transport of moistened pollen could have evolved. The evolutionary history of bees has many implications for the biology of bees in the present day, and I lay out a number of predictions that could help confirm or refute my hypotheses.


title: Harvesting electrical current from intact plant leaves

abstract:
Efforts to replace fossil fuels with renewable energy technologies, especially solar energy conversion, continue to improve the potential to produce useful amounts of energy without significant pollution. Utilization of photosynthetic organisms in bio-photo electrochemical cells (BPECs) are a potentially important source of clean energy. Here, we show that it is possible to harvest photocurrent directly from unprocessed plant tissues in specialized BPECs. The source of electrons are shown to originate from the Photosystem II water-oxidation reaction that results in oxygen evolution. In addition to terrestrial and crop plants, we further demonstrate the ability of the desert plant Corpuscularia lehmannii to produce bias-free photocurrent without the addition of an external electrolyte. Finally, we show the use of pond-grown water lilies to generate photocurrent. Different leaves produce photocurrent densities in the range of [~] 1 - 10 mA / cm2 which is significantly higher than microorganism-based BPECs. The relatively high photocurrent and the simplicity of the plants BPEC may pave the way toward the establishment of first applicative photosynthetic based energy technologies.

Broader ContextIt is no secret human society is experiencing an energy and environmental crisis due to our reliance on fossil fuels. In order to promote alternative, cleaner, and more sustainable approaches to energy production, we wish to explore the possibility of using natures method of solar energy conversion in the simplest, least polluting, most sustainable fashion possible. Photosynthesis provides a remarkable example of molecular system for solar energy conversion to storable fuels. Many studies have strived to merge natural photosynthesis (as isolated complexes, isolated membranes, or intact microorganisms) with a variety of electrochemical harvesting technologies. In this paper we show that we can directly couple the power of water oxidation by Photosystem II in intact plants to bio-electrochemical cells without the need to perform expensive, complicated, and polluting isolation. We show that current harvesting (up to current densities of 10 mA / cm2) can be performed using plants of different types: plants of agricultural importance, succulents with internal water-based reservoirs and aquatic plants, used in situ in their growth ponds. We also show that with minimal external bias, hydrogen can be obtained, to be used as a clean fuel. We believe that these results can lead to the development of localized clean energy technologies, where the benefits of plant growth for any purpose can be enhanced by obtaining significant amounts of clean energy.

category: bioengineering


title: Acetobase: database update and reanalysis offormyltetrahydrofolate synthetase amplicon sequencing data from the anaerobic digesters

abstract: AcetoBase is a public repository and database published in 2019, for the formyltetrahydrofolate synthetase (FTHFS) sequences. It is the first systematic collection of bacterial formyltetrahydrofolate nucleotide and protein sequences from the genomes and metagenome assembled genomes (MAGs), as well as sequences generated by clone library sequencing. In addition, AcetoBase was first to establish connection between FTHFS gene with the Wood-Ljungdahl pathway and 16S rRNA genes. Since the publication of AcetoBase, significant improvements were seen in the taxonomy of many bacterial lineages and accessibility/availability of public genomics and metagenomics data. Thus, an update to the AcetoBase database with new sequence data and taxonomy has been made along with improvements in web-functionality and user interface. The update in AcetoBase reference database version 2 was furthermore evaluated by reanalysis of publicly accessible FTHFS amplicon sequencing data previously analysed with AcetoBase version 1. The latest database update showed significant improvements in the taxonomic assignments of FTHFS sequences. AcetoBase with its enhancements in functionality and content is publicly accessible at https://acetobase.molbio.slu.se.

category: bioinformatics


title: A deep dilated convolutional residual network for predicting interchain contacts of protein homodimers

abstract:
MotivationDeep learning has revolutionized protein tertiary structure prediction recently. The cutting-edge deep learning methods such as AlphaFold can predict high-accuracy tertiary structures for most individual protein chains. However, the accuracy of predicting quaternary structures of protein complexes consisting of multiple chains is still relatively low due to lack of advanced deep learning methods in the field. Because interchain residue-residue contacts can be used as distance restraints to guide quaternary structure modeling, here we develop a deep dilated convolutional residual network method (DRCon) to predict interchain residue-residue contacts in homodimers from residue-residue co-evolutionary signals derived from multiple sequence alignments of monomers, intrachain residue-residue contacts of monomers extracted from true/predicted tertiary structures or predicted by deep learning, and other sequence and structural features.

ResultsTested on three homodimer test datasets (Homo_std dataset, DeepHomo dataset, and CASP14-CAPRI dataset), the precision of DRCon for top L/5 interchain contact predictions (L: length of monomer in a homodimer) is 43.46%, 47.15%, and 24.81% respectively, which is substantially better than two existing deep learning interchain contact prediction methods. Moreover, our experiments demonstrate that using predicted tertiary structure or intrachain contacts of monomers in the unbound state as input, DRCon still performs reasonably well, even though its accuracy is lower than when true tertiary structures in the bound state are used as input. Finally, our case study shows that good interchain contact predictions can be used to build high-accuracy quaternary structure models of homodimers.

AvailabilityThe source code of DRCon is available at https://github.com/jianlin-cheng/DRCon.

Contactchengji@missouri.edu


title: Named Entity Recognition and Linking: a Portuguese and Spanish Oncological Parallel Corpus

abstract: Biomedical literature is the main mean of communication for researchers to share their findings. Since biomedical literature is composed of a large collection of text expressed in natural language, the usage of text mining tools to extract information from those texts automatically is of utmost importance. The problem is that the majority of the state-of-the-art tools were not developed to deal with other languages besides English, which in biomedical literature is even more critical since a significant part of health-related texts is written in the authors native language. To address this issue, this work presents a deep learning NERL (Named Entity Recognition and Linking) system and a parallel corpus for the Spanish and Portuguese languages focused on the oncological domain. Both the system and the corpus are available at https://github.com/lasigeBioTM/ICERL_system-ICR_Corpus.


title: A plastidial retrograde-signal potentiates biosynthesis of systemic stress response activators

abstract:
O_LIPlants employ an array of intricate and hierarchical signaling cascades to perceive and transduce informational cues to synchronize and tailor adaptive responses. Systemic stress response (SSR) is a recognized complex signaling and response network quintessential to plants local and distal responses to environmental triggers, however, the identity of the initiating signals has remained fragmented.
C_LIO_LIHere, we show that both biotic (aphids and viral pathogens) and abiotic (high-light and wounding) stresses induce accumulation of the plastidial-retrograde-signaling metabolite, methylerythritol cyclodiphosphate (MEcPP), leading to reduction of the phytohormone, auxin, and the subsequent decreased expression of the phosphatase, PP2C.D1.
C_LIO_LIThis enables phosphorylation of mitogen-activated protein kinases (MAPK3/6), and the consequential induction of the downstream events ultimately resulting in biosynthesis of the two SSR priming metabolites, pipecolic- and N-hydroxy-pipecolic acid.
C_LIO_LIThis work identifies plastids as the initiation site, and the plastidial retrograde-signal, MEcPP as the initiator of a multi-component signaling cascade potentiating the biosynthesis of SSR activators, in response to biotic and abiotic triggers.
C_LI


title: Impact of zoonotic disease outbreaks on international wildlife trade dynamics in Ghana

abstract: Wildlife is under intense pressure from trade, which most likely contributes to zoonotic diseases. The study explores the impact of zoonotic disease outbreaks on the wildlife trade of Ghana. This study provides an in-depth analysis of the trends of taxa trade and factors that influence trade from 1975-2018 by combining zoonotic disease data with data from the CITES database. Trade flow showed that reptiles were the most traded group, followed by birds, mammals, and amphibians. Species of the families Pythonidae, Dendrobatidae, Cercopithecidae, and Psittacidae were the most traded. The decade mean number of trade for 1997-2007 was the highest (n = 62) followed by 2008-2018 (n = 54.4). Most exporter countries that traded with Ghana are from Africa and importers from the United States of America, Europe and Asia. Continuous trade in reptiles and birds, especially the endangered pythons and psittacus species, could lead to their extinction in the wild. The outbreak of zoonotic diseases influenced the dynamics of the wildlife trade in Ghana as traders shifted their activities among taxa over a period of time. Because those taxa were observed to harbour zoonotic diseases and constitute high health risks when traded. Mammals trade flow decreased with disease outbreaks over time, while reptiles increased. Early detection of zoonotic diseases and the adoption of an expanded education module on avoiding species capable of harbouring pathogens will most likely help reduce trade in wildlife.

category: zoology


title: Educational Attainment Polygenic Scores, Socioeconomic Factors, and Cortical Structure in Children and Adolescents

abstract:
BackgroundGenome-wide polygenic scores for educational attainment (PGS-EA) and socioeconomic factors, which are correlated with each other, have been consistently associated with academic achievement and general cognitive ability in children and adolescents. Yet, the independent associations of PGS-EA and socioeconomic factors with specific underlying factors at the neural and neurocognitive levels are not well understood. The goal of this study was to examine the unique contributions of PGS-EA and parental education to cortical thickness (CT), cortical surface area (SA), and neurocognitive skills in children and adolescents.

MethodsParticipants were typically developing children and adolescents (3-21 years of age; 53% male; N = 391). High-resolution, T1-weighted magnetic resonance imaging data were acquired. PGS-EA were computed based on the most recent genome-wide association study of educational attainment. Sustained attention, inhibitory control, working memory, vocabulary, and episodic memory were measured.

ResultsPGS-EA and parental education were independently and significantly associated with SA, vocabulary, and attention outcomes but were not associated with CT. Vertex-wise analyses indicated that higher PGS-EA was significantly associated with greater SA in the left medial orbitofrontal gyrus and inferior frontal gyrus after accounting for parental education. Higher parental education was significantly associated with greater SA in the left parahippocampal gyrus after accounting for PGS-EA.

ConclusionsThese findings suggest that education-linked genetics may influence SA, particularly in certain frontal regions, leading to variability in academic achievement. Results suggested genetic confounding in associations between parental education and SA in children and adolescents, with these associations remaining significant after controlling for PGS-EA.


title: Unsupervised mining of HLA-I peptidomes reveals new binding motifs and substantial false positives in community database

abstract: Modern vaccine designs and studies of human leukocyte antigen (HLA)-mediated immune responses rely heavily on the knowledge of HLA allele-specific binding motifs and computational prediction of HLA-peptide binding affinity. Breakthroughs in HLA peptidomics have considerably expanded the databases of natural HLA ligands and enabled detailed characterizations of HLA-peptide binding specificity. However, cautions must be made when analyzing HLA peptidomics data because identified peptides may be contaminants in mass spectrometry or may weakly bind to the HLA molecules. Here, a hybrid de novo peptide sequencing approach was applied to large-scale mono-allelic HLA peptidomics datasets to uncover new ligands and refine current knowledge of HLA binding motifs. Up to 12-40% of the peptidomics data were low-binding affinity peptides with an arginine or a lysine at the C-terminus and likely to be tryptic peptide contaminants. Thousands of these peptides have been reported in a community database as legitimate ligands and might be erroneously used for training prediction models. Furthermore, unsupervised clustering of identified ligands revealed additional binding motifs for several HLA class I alleles and effectively isolated outliers that were experimentally confirmed to be false positives. Overall, our findings expanded the knowledge of HLA binding specificity and advocated for more rigorous interpretation of HLA peptidomics data that will ensure the high validity of community HLA ligandome databases.

category: immunology


title: Global fitting for high-accuracy multi-channel single-molecule localization

abstract: Multi-channel detection in single-molecule localization microscopy (SMLM) greatly increases information content for various biological applications. Here, we present globLoc, a graphics processing unit (GPU) based global fitting algorithm with flexible PSF modeling and parameter sharing, to extract maximum information from multi-channel single molecule data. We show, both in simulations and experiments, that global fitting can substantially improve the 3D localization precision for biplane and 4Pi SMLM and color assignment for ratiometric multicolor imaging.


title: A mechanism for migrating bacterial populations to non-genetically adapt to new environments

abstract: Populations of chemotactic bacteria can rapidly expand into new territory by consuming and chasing an attractant cue in the environment, increasing the populations overall growth in nutrient-rich environments. Although the migrating fronts driving this expansion contain cells of multiple swimming phenotypes, the consequences of non-genetic diversity for population expansion are unknown. Here, through theory and simulations, we predict that expanding populations non-genetically adapt their phenotype composition to migrate effectively through multiple physical environments. Swimming phenotypes in the migrating front are spatially sorted by chemotactic performance, but the mapping from phenotype to performance depends on the environment. Therefore, phenotypes that perform poorly localize to the back of the group, causing them to selectively fall behind. Over cell divisions, the group composition dynamically enriches for high-performers, enhancing migration speed and overall growth. Furthermore, non-genetic inheritance controls a trade-off between large composition shifts and slow responsiveness to new environments, enabling a diverse population to out-perform a non-diverse one in varying environments. These results demonstrate that phenotypic diversity and collective behavior can synergize to produce emergent functionalities. Non-genetic inheritance may generically enable bacterial populations to transiently adapt to new situations without mutations, emphasizing that genotype-to-phenotype mappings are dynamic and context-dependent.


title: Selective Microautophagy of Proteasomes is Initiated by ESCRT-0 and Is Promoted by Proteasome Ubiquitylation

abstract:
The proteasome is central to proteolysis by the ubiquitin-proteasome system under normal growth conditions but is itself degraded through macroautophagy under nutrient stress. A recently described AMPK (AMP-activated protein kinase)-regulated ESCRT (endosomal sorting complex required for transport)-dependent microautophagy pathway also regulates proteasome trafficking and degradation in low glucose conditions in yeast. Aberrant proteasomes are more prone to microautophagy, suggesting the ESCRT system fine-tunes proteasome quality control under low glucose stress. Here we uncover additional features of the selective microautophagy of proteasomes. Genetic or pharmacological induction of aberrant proteasomes is associated with increased mono- or oligo-ubiquitylation of proteasome components, which appear to be recognized by ESCRT-0. AMPK controls this pathway in part by regulating the trafficking of ESCRT-0 to the vacuole surface, which also leads to degradation of the Vps27 subunit of ESCRT-0. The Rsp5 ubiquitin ligase contributes to proteasome subunit ubiquitylation, and multiple ubiquitin-binding elements in Vps27 are involved in their recognition. We propose that ESCRT-0 at the vacuole surface recognizes ubiquitylated proteasomes and initiates their microautophagic elimination during glucose depletion.

Saummary statementESCRT-0 selectively targets aberrant proteasomes for microautophagy by recognition of proteasome ubiquitylation status to fine-tune proteasome quality control under low glucose conditions.


title: Brain I3 Binding Protein regulates K-Ras4B membrane localization and signaling

abstract: Membrane localization of Ras proteins is necessary for their biological functions and oncogenic activity. We report here on the identification of Brain I3 Binding Protein (BRI3BP) as a novel binding partner for Ras. We show that K-Ras4B plasma membrane localization and biological function are reduced in the absence of BRI3BP. BRI3BP interacts with K-Ras4B and K-Ras4A and our data suggest that BRI3BP operates within the recycling endosomal compartment to regulate K-Ras localization to the plasma membrane. This study uncovers a new regulatory protein for Ras membrane localization.


title: SpinX: Time-resolved 3D Analysis of Mitotic Spindle Dynamics using Deep Learning Techniques and Mathematical Modelling

abstract: Time-lapse microscopy movies have transformed the study of subcellular dynamics. However, manual analysis of movies can introduce bias and variability, obscuring important insights. While automation can overcome such limitations, spatial and temporal discontinuities in time-lapse movies render methods such as object segmentation and tracking difficult. Here we present SpinX, a framework for reconstructing gaps between successive frames by combining Deep Learning and mathematical object modelling. By incorporating expert feedback through selective annotations, SpinX identifies subcellular structures, despite confounding neighbour-cell information, non-uniform illumination and variable marker intensities. The automation and continuity introduced allows precise 3-Dimensional tracking and analysis of spindle movements with respect to the cell cortex for the first time. We demonstrate the utility of SpinX using distinct spindle markers and drug treatments. In summary, SpinX provides an exciting opportunity to study spindle dynamics in a sophisticated way, creating a framework for step changes in studies using time-lapse microscopy.


title: Membrane-mediated dimerization potentiates PIP5K lipid kinase activity

abstract: The phosphatidylinositol 4-phosphate 5-kinase (PIP5K) family of lipid modifying enzymes generate the majority of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) lipids found at the plasma membrane in eukaryotic cells. PI(4,5)P2 lipids serve a critical role in regulating receptor activation, ion channel gating, endocytosis, and actin nucleation. Here we describe how PIP5K activity is regulated by cooperative binding to PI(4,5)P2 lipids and membrane-mediated dimerization of the kinase domain. In contrast to constitutively dimeric phosphatidylinositol 5-phosphate 4-kinase (PIP4K, type II PIPK), solution PIP5K exists in a weak monomer-dimer equilibrium. PIP5K monomers can associate with PI(4,5)P2 containing membranes and dimerize in a protein density dependent manner. Although dispensable for PI(4,5)P2 binding and lipid kinase activity, dimerization enhances the catalytic efficiency of PIP5K through a mechanism consistent with allosteric regulation. Additionally, dimerization amplifies stochastic variation in the kinase reaction velocity and strengthens effects such as the recently described stochastic geometry sensing. Overall, the mechanism of PIP5K membrane binding creates a broad dynamic range of lipid kinase activities that are coupled to the density of PI(4,5)P2 and membrane bound kinase.


title: Cell-surface tethered promiscuous biotinylators enable small-scale surface proteomics of human exosomes

abstract: Characterization of cell surface proteome differences between cancer and healthy cells is a valuable approach for the identification of novel diagnostic and therapeutic targets. However, selective sampling of surface proteins for proteomics requires large samples (>10e7 cells) and long labeling times. These limitations preclude analysis of material-limited biological samples or the capture of rapid surface proteomic changes. Here, we present two labeling approaches to tether exogenous peroxidases (APEX2 and HRP) directly to cells, enabling rapid, small-scale cell surface biotinylation without the need to engineer cells. We used a novel lipidated DNA-tethered APEX2 (DNA-APEX2), which upon addition to cells promoted cell agnostic membrane-proximal labeling. Alternatively, we employed horseradish peroxidase (HRP) fused to the glycan binding domain of wheat germ agglutinin (WGA-HRP). This approach yielded a rapid and commercially inexpensive means to directly label cells containing common N-Acetylglucosamine (GlcNAc) and sialic acid glycans on their surface. The facile WGA-HRP method permitted high surface coverage of cellular samples and enabled the first comparative surface proteome characterization of cells and cell-derived exosomes, leading to the robust quantification of 1,020 cell and exosome surface proteins. We identified a newly-recognized subset of exosome-enriched markers, as well as proteins that are uniquely upregulated on Myc oncogene-transformed prostate cancer exosomes. These two cell-tethered enzyme surface biotinylation approaches are highly advantageous for rapidly and directly labeling surface proteins across a range of material-limited sample types.


title: The Incubascope : a simple, compact and large field of view microscope for long-term imaging inside an incubator

abstract:
Optical imaging has rapidly evolved in the last decades. Sophisticated microscopes allowing optical sectioning for 3D imaging or sub-diffraction resolution are available. Due to price and maintenance issues, these microscopes are often shared between users in facilities. Consequently, long term access is often prohibited and does not allow to monitor slowly evolving biological systems or to validate new models like organoids. Preliminary coarse long-term data that do not require acquisition of terabytes of high-resolution images are important as a first step. By contrast with expansive all-in-one commercialized stations, standard microscopes equipped with incubator stages offer a more cost-effective solution despite imperfect long run atmosphere and temperature control.

Here, we present the Incubascope, a custom-made compact microscope that fits into a table-top incubator. It is cheap and simple to implement, user-friendly and yet provides high imaging performances. The system has a field of view of 5.5 x 8 mm2, a 3 m resolution, a 10 frames/second acquisition rate, and is controlled with a Python-based graphical interface. We exemplify the capabilities of the Incubascope on biological applications such as the hatching of Artemia salina eggs, the growth of the slime mold Physarum polycephalum and of encapsulated spheroids of mammalian cells.

category: biophysics


title: High-resolution imaging and manipulation of endogenous AMPA receptor surface mobility during synaptic plasticity and learning

abstract: Regulation of synaptic neurotransmitter receptor content is a fundamental mechanism for tuning synaptic efficacy during experience-dependent plasticity and behavioral adaptation. However, experimental approaches to track and modify receptor movements in integrated experimental systems are limited. Exploiting AMPA-type glutamate receptors (AMPAR) as a model, we generated a knock-in mouse expressing the biotin acceptor peptide (AP) tag on the GluA2 extracellular N-terminus. Cell-specific introduction of biotin ligase allows the use of monovalent or tetravalent avidin variants to respectively monitor or manipulate the surface mobility of endogenous AMPAR containing biotinylated AP-GluA2 in neuronal subsets. AMPAR immobilization precluded the expression of long-term potentiation and formation of contextual fear memory, allowing for target-specific control of the expression of synaptic plasticity and animal behavior. The AP tag knock-in model offers unprecedented access to resolve and control the spatiotemporal dynamics of endogenous receptors, and opens new avenues to study the molecular mechanisms of synaptic plasticity and learning.


title: Design of Peptide Vaccine for COVID19: CD8+ and CD4+ T cell epitopes from SARS-CoV-2 open-reading-frame protein variants

abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, has challenged public health at an unprecedented scale which has led to a dramatic loss of human life worldwide. To design a protective vaccine against SARS-CoV-2, it is necessary to understand which SARS-CoV-2 specific epitopes can elicit a T cell response and provide protection across a broad population. In this study, PLpro and RdRp, two immunogenic non-structural proteins from an immunodominant gene region ORF1ab, as well as ORF3a and ORF9b are identified as potential vaccine targets against SARS-CoV-2. To select top epitopes for vaccine design, we used various clinical properties, such as antigenicity, allergenicity, toxicity and IFN-y secretion. The analysis of CD8 and CD4 T cell epitopes revealed multiple potential vaccine constructs that cover a high percentage of the world population. We identified 8 immunogenic, antigenic, non-allergenic, non-toxic, stable and IFN-y inducing CD8 proteins for nsp3, 4 for nsp12, 11 for ORF3a and 3 for ORF9b that are common across four lineages of variants of concern - B.1.1.7, P.1, B.1.351 and B.1.617.2, which protect 98.12%, 87.08%, 96.07% and 63.8% of the world population, respectively. We also identified variant specific T cell epitopes that could be useful in targeting each variant strain separately. Including the prediction of mouse MHC affinity towards our top CD8 epitopes, our study revealed a total of 3 immunogenic, antigenic, non-allergenic, non-toxic, stable and IFN-y inducing CD8 epitopes overlapping with 6 antigenic, non-allergenic, non-toxic, stable and IFN-y inducing CD4 epitopes across all four variants of concern which can effectively be utilized in pre-clinical studies. The landscape of SARS-CoV-2 T cell epitopes that we identified can help lead SARS-CoV-2 vaccine development as well as epitope-based peptide vaccine research in the future.


title: Identification of HLA-A*24:02-restricted CTL candidate epitopes derived from the non-structural polyprotein 1a of SARS-CoV-2 and analysis of their conservation using the mutation database of SARS-CoV-2 variants

abstract:
COVID-19 vaccines are currently being administrated worldwide and playing a critical role in controlling the pandemic. They have been designed to elicit neutralizing antibodies against Spike protein of the original SARS-CoV-2, and hence they are less effective against SARS-CoV-2 variants with mutated Spike than the original virus. It is possible that novel variants with abilities of enhanced transmissibility and/or immunoevasion will appear in the near future and perfectly escape from vaccine-elicited immunity. Therefore, the current vaccines may need to be improved to compensate for the viral evolution. For this purpose, it may be beneficial to take advantage of CD8+ cytotoxic T lymphocytes (CTLs). Several lines of evidence suggest the contribution of CTLs on the viral control in COVID-19, and CTLs target a wide range of proteins involving comparatively conserved non-structural proteins. Here, we identified twenty-two HLA-A*24:02-restricted CTL candidate epitopes derived from the non-structural polyprotein 1a (pp1a) of SARS-CoV-2 using computational algorithms, HLA-A*24:02 transgenic mice and the peptide-encapsulated liposomes. We focused on pp1a and HLA-A*24:02 because pp1a is relatively conserved and HLA-A*24:02 is predominant in East Asians such as Japanese. The conservation analysis revealed that the amino acid sequences of 7 out of the 22 epitopes were hardly affected by a number of mutations in the Sequence Read Archive database of SARS-CoV-2 variants. The information of such conserved epitopes might be useful for designing the next-generation COVID-19 vaccine that is universally effective against any SARS-CoV-2 variants by the induction of both anti-Spike neutralizing antibodies and CTLs specific for conserved epitopes.

ImportanceCOVID-19 vaccines have been designed to elicit neutralizing antibodies against the Spike protein of the original SARS-CoV-2, and hence they are less effective against variants. It is possible that novel variants will appear and escape from vaccine-elicited immunity. Therefore, the current vaccines may need to be improved to compensate for the viral evolution. For this purpose, it may be beneficial to take advantage of CD8+ cytotoxic T lymphocytes (CTLs). Here, we identified twenty-two HLA-A*24:02-restricted CTL candidate epitopes derived from the non-structural polyprotein 1a (pp1a) of SARS-CoV-2. We focused on pp1a and HLA-A*24:02 because pp1a is conserved and HLA-A*24:02 is predominant in East Asians. The conservation analysis revealed that the amino acid sequences of 7 out of the 22 epitopes were hardly affected by mutations in the database of SARS-CoV-2 variants. The information might be useful for designing the next-generation COVID-19 vaccine that is universally effective against any variants.


title: Mechanical stimulation promotes enthesis injury repair by mobilizing Prx1+ cells via ciliary TGF-β signaling

abstract: Proper mechanical stimulation can improve rotator cuff enthsis injury repair. However, the underlying mechanism of mechanical stimulation promoting injury repair is still unknown. In this study, we found that Prx1+ cell was essential for murine rotator cuff enthesis development identified by single-cell RNA sequence and involved in the injury repair. Proper mechanical stimulation could promote the migration of Prx1+ cells to enhance enthesis injury repair. Meantime, TGF-{beta} signaling and primary cilia played an essential role in mediating mechanical stimulation signaling transmission. Proper mechanical stimulation enhanced the release of active TGF-{beta}1 to promote migration of Prx1+ cells. Inhibition of TGF-{beta} signaling eliminated the stimulatory effect of mechanical stimulation on Prx1+ cell migration and enthesis injury repair. In addition, knockdown of Pallidin to inhibit TGF-{beta}R2 translocation to the primary cilia or deletion of IFT88 in Prx1+ cells also restrained the mechanics-induced Prx1+ cells migration. These findings suggested that mechanical stimulation could increase the release of active TGF-{beta}1 and enhance the mobilization of Prx1+ cells to promote enthesis injury repair via ciliary TGF-{beta} signaling.


title: Crowding within synaptic junctions influence the degradation of adenoside nucleotides by CD39 and CD73 ectonucleotidases

abstract: 1Synapsed cells can communicate using exocytosed nucleotides like adenosine triphosphate (ATP). Ectonucleotidases localized to a synaptic junction degrade such nucleotides into metabolites like adenosine monophosphate (AMP) or adenosine, oftentimes in a sequential manner. CD39 and CD73 are a representative set of coupled ectonucleotidases, where CD39 first converts ATP and adenosine diphosphate (ADP) into AMP, after which the AMP product is dephosphorylated into adenosine by CD73. Hence, CD39/CD73 help shape cellular responses to extracellular ATP. In a previous study [1] we demonstrated that the rates of coupled CD39/CD73 activity within synapse-like junctions are strongly controlled by the enzymes co-localization, their surface charge densities, and the electrostatic potential of the surrounding cell membranes. In this study, we demonstrate that crowders within a synaptic junction, which can include globular proteins like cytokines and membrane-bound proteins, impact coupled CD39/CD73 electronucleotidase activity and in turn, the availability of intrasynapse ATP. Specifically, we simulated a spatially-explicit, reaction-diffusion model for the coupled conversion of ATP[->]AMP and AMP[->]adenosine in a model synaptic junction with crowders via the finite element method. Our modeling results suggest that the association rate for ATP to CD39 is strongly influenced by the density of intrasynaptic protein crowders, as increasing crowder density suppressed ATP association kinetics. Much of this suppression can be rationalized based on a loss of configurational entropy. The surface charges of crowders can further influence the association rate, with the surprising result that favorable crowder/nucleotide electrostatic interactions can yield CD39 association rates that are faster than crowder-free configurations. However, attractive crowder/nucleotide interactions decrease the rate and efficiency of adenosine production, which in turn increases the availability of ATP and AMP within the synapse relative to crowder-free configurations. These findings highlight how CD39/CD73 ectonucleotidase activity, electrostatics and crowding within synapses influence the availability of nucleotides for intercellular communication.


title: Epithelial Plasticity and Innate Immune Activation Promote Lung Tissue Remodeling following Respiratory Viral Infection.

abstract: Epithelial plasticity has been suggested in lungs of mice following genetic depletion of stem cells but is of unknown physiological relevance. Viral infection and chronic lung disease share similar pathological features of stem cell loss in alveoli, basal cell (BC) hyperplasia in small airways, and innate immune activation, that contribute to epithelial remodeling and loss of lung function. We show that a novel lineage of distal airway secretory cells, intralobar serous (IS) cells, are activated to assume BC fates following influenza virus infection. Nascent BC differ from pre-existing BC by high expression of IL-22ra1 and undergo IL-22-dependent expansion for colonization of injured alveoli. Resolution of virus-elicited inflammation resulted in BC>IS re-differentiation in repopulated alveoli, and increased local expression of antimicrobial factors, but failed to replace normal alveolar epithelium. Epithelial plasticity therefore protects against mortality from acute respiratory viral infection but results in distal lung remodeling and loss of lung function.


title: Dynamic regulation and requirement for ribosomal RNA transcription duringmammalian development

abstract:
Ribosomal RNA (rRNA) transcription by RNA Polymerase I (Pol I) is a critical rate-limiting step in ribosome biogenesis, which is essential for cell survival. Despite its global function, disruptions in ribosome biogenesis cause tissue-specific birth defects called ribosomopathies, which frequently affect craniofacial development. Here, we describe a cellular and molecular mechanism underlying the susceptibility of craniofacial development to disruptions in Pol I transcription. We show that Pol I subunits are highly expressed in the neuroepithelium and neural crest cells (NCC), which generate most of the craniofacial skeleton. High expression of Pol I subunits sustains elevated rRNA transcription in NCC progenitors, which supports their high tissue-specific levels of protein translation, but also makes NCC particularly sensitive to rRNA synthesis defects. Consistent with this model, NCC-specific deletion of Pol I subunits Polr1a, Polr1c, and associated factor Tcof1 in mice cell-autonomously diminishes rRNA synthesis, which causes an imbalance between rRNA and ribosomal proteins. This leads to increased binding of ribosomal proteins Rpl5 and Rpl11 to Mdm2 and concomitantly diminished binding between Mdm2 and p53. Consequently, p53 protein accumulates, resulting in NCC apoptosis and craniofacial anomalies. Furthermore, compound mutations in Pol I subunits and associated factors specifically exacerbates the craniofacial anomalies characteristic of the ribosomopathies Treacher Collins Syndrome and Acrofacial Dysostosis-Cincinnati Type. Altogether, our novel results demonstrate a dynamic spatiotemporal requirement for rRNA transcription during mammalian cranial NCC development and corresponding tissue-specific threshold sensitivities to disruptions in rRNA transcription in the pathogenesis of congenital craniofacial disorders.

Significance statementRNA Polymerase I (Pol I) mediated rRNA transcription is required for protein synthesis in all tissues for normal growth and survival as well as for proper embryonic development. Interestingly, disruptions in Pol I mediated transcription perturb ribosome biogenesis and lead to tissue-specific birth defects, which commonly affect the head and face. Our novel results show that during mouse development, Pol I mediated rRNA transcription and protein translation is tissue-specifically elevated in neural crest cells, which give rise to bone, cartilage, and ganglia of the head and face. Using new mouse models, we further show that neural crest cells are highly sensitive to disruptions in Pol I and that when rRNA synthesis is genetically downregulated, it specifically results in craniofacial anomalies.

category: developmental biology


title: Calculating the force-dependent unbinding rate of biological macromolecular bonds from the force-ramp optical trapping assays

abstract: The non-covalent biological bonds that constitute protein-protein or protein-ligand interactions play crucial roles in many cellular functions, including mitosis, motility, and cell-cell adhesion. The effect of external force (F) on the unbinding rate (koff(F)) of macromolecular interactions is a crucial parameter to understanding the mechanisms behind these functions. Optical tweezer-based single-molecule force spectroscopy is frequently used to obtain quantitative force-dependent dissociation data on slip, catch, and ideal bonds. However, analyses of this data using dissociation time or dissociation force histograms often quantitatively compare bonds without fully characterizing their underlying biophysical properties. Additionally, the results of histogram-based analyses can depend on the rate at which force was applied during the experiment and the experiments sensitivity. Here, we present an analytically derived cumulative distribution function-like approach to analyzing force-dependent dissociation force spectroscopy data. We demonstrate the benefits and limitations of the technique using stochastic simulations of various bond types. We show that it can be used to obtain the detachment rate and force sensitivity of biological macromolecular bonds from force spectroscopy experiments by explicitly accounting for loading rate and noisy data. We also discuss the implications of our results on using optical tweezers to collect force-dependent dissociation data.


title: Biomass accumulation and physiological responses of tomato plants to magnetically-treated water in hydroponic conditions

abstract: Magnetically-treated water (MTW) has been reported to enhance biomass accumulation in plants. However, the underlying mechanisms are not fully understood, and the existing reports only deal with soil-grown plants. Thus, the purpose of this experiment was to assess whether or not MTW affects main physiological processes (gas exchange, biomass accumulation and water potential) in tomato plants whose water supply was only MTW. Two experiments were done in hydroponic semi-controlled conditions, consisting of a loop system with permanent recirculation of water through a non-uniform magnet. The plants grown under MTW showed a significant increase in chlorophyll content, photosynthesis and transpiration at high light irradiances, although the increase in stomatal conductance was less significant. MTW also increased fruit fresh biomass, number of fruits and root dry biomass in 61.7 %, 85.3 % and 30.3 % respectively, but this was only achieved at natural sunlight conditions. Moreover, treated plants showed higher root hydraulic conductance and leaf water potential, which is thought to be related with a lower surface tension of MTW, an effect that is consistent with previous studies. The higher biomass accumulation in tomato plants under MTW is likely explained because of a faster water transport from the roots to the leaves via xylem, which in turn increases H2O efflux and CO2 assimilation in the leaves, thanks to a higher stomatal conductance.