title: Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics

abstract: Therapeutic mRNAs and vaccines are being developed for a broad range of human diseases, including COVID-19. However, their optimization is hindered by mRNA instability and inefficient protein expression. Here, we describe design principles that overcome these barriers. We develop a new RNA sequencing-based platform called PERSIST-seq to systematically delineate in-cell mRNA stability, ribosome load, as well as in-solution stability of a library of diverse mRNAs. We find that, surprisingly, in-cell stability is a greater driver of protein output than high ribosome load. We further introduce a method called In-line-seq, applied to thousands of diverse RNAs, that reveals sequence and structure-based rules for mitigating hydrolytic degradation. Our findings show that "superfolder" mRNAs can be designed to improve both stability and expression that are further enhanced through pseudouridine nucleoside modification. Together, our study demonstrates simultaneous improvement of mRNA stability and protein expression and provides a computational-experimental platform for the enhancement of mRNA medicines.

category: molecular biology


title: The Cdc42 GTPase activating protein Rga6 promotes the cortical localization of Septin

abstract: Septins are a family of filament-forming GTP-binding proteins that regulate fundamental cellular activities such as cytokinesis, cell polarity, and membrane remodelling. In general, Septin filaments function as barriers and scaffolds on the cell cortex. However, little is known about the mechanism that governs the recruitment and localization of the Septin complex to the cell cortex. Here, we identified the Cdc42 GTPase activating protein Rga6 as a key protein involved in promoting the localization of the Septin complex to the cell cortex in the fission yeast Schizosaccharomyces pombe. Rga6 interacts with the Septin complex and colocalizes with the Septin complex on the cell cortex. Live-cell microscopic analysis further showed Septin enrichment at the cortical regions adjacent to the growing cell tip. The Septin enrichment likely plays a crucial role in confining active Cdc42 to the growing cell tip. Hence, our findings support a model that Rga6 regulates polarized cell growth partly through promoting targeted localization of the Septin complex on the cell cortex.

category: cell biology


title: Caspase-2 regulates S-phase cell cycle events to protect from DNA damage accumulation independent of apoptosis

abstract: In addition to its classical role in apoptosis, accumulating evidence suggests that caspase-2 has non-apoptotic functions, including regulation of cell division. Loss of caspase-2 is known to increase proliferation rates but how caspase-2 is regulating this process is currently unclear. We show that caspase-2 is activated in dividing cells in G1- and early S-phase. In the absence of caspase-2, cells exhibit numerous S-phase defects including delayed exit from S-phase, S-phase-associated chromosomal aberrations, and increased DNA damage following S-phase arrest. In addition, caspase-2-deficient cells have a higher frequency of stalled replication forks, decreased DNA fiber length, and impeded progression of DNA replication tracts. This indicates that caspase-2 reduces replication stress and promotes replication fork protection to maintain genomic stability. These functions are independent of the pro-apoptotic function of caspase-2 because blocking caspase-2-induced cell death had no effect on cell division or DNA damage-induced cell cycle arrest. Thus, our data supports a model where caspase-2 regulates cell cycle events to protect from the accumulation of DNA damage independently of its pro-apoptotic function.


title: Shared and disease-specific host gene-microbiome interactions across human diseases

abstract: While the gut microbiome and host gene regulation separately contribute to gastrointestinal disorders, it is unclear how the two may interact to influence host pathophysiology. Here, we developed a machine learning-based framework to jointly analyze host transcriptomic and microbiome profiles from 416 colonic mucosal samples of patients with colorectal cancer, inflammatory bowel disease, and irritable bowel syndrome. We identified potential interactions between gut microbes and host genes that are disease-specific, as well as interactions that are shared across the three diseases, involving host genes and gut microbes previously implicated in gastrointestinal inflammation, gut barrier protection, energy metabolism, and tumorigenesis. In addition, we found that mucosal gut microbes that have been associated with all three diseases, such as Streptococcus, interact with different host pathways in each disease, suggesting that similar microbes can affect host pathophysiology in a disease-specific manner through regulation of different host genes.

category: genomics


title: Structural characterization of the antimicrobial peptides myxinidin and WMR in bacterial membrane mimetic micelles and bicelles

abstract: Antimicrobial peptides are a promising class of alternative antibiotics that interact selectively with negatively charged lipid bilayers. This paper presents the structural characterization of the antimicrobial peptides myxinidin and WMR associated with bacterial membrane mimetic micelles and bicelles by NMR, CD spectroscopy, and Molecular Dynamics simulations. Both peptides adopt a different conformation in the lipidic environment than in aqueous solution. The location of peptides in micelles and bicelles has been studied by paramagnetic relaxation enhancement experiments with paramagnetic tagged 5- and 16-doxyl stearic acid (5-/16-SASL). Multi-microsecond long molecular dynamics simulations of multiple copies of the peptides were used to gain an atomic level of detail on membrane-peptide and peptide-peptide interactions. Our results highlight an essential role of the negatively charged membrane mimetic in the structural stability of both myxinidin and WMR. The peptides localize predominantly in the membranes headgroup region and have a noticeable membrane thinning effect on the overall bilayer structure. Myxinidin and WMR show different tendency to selfaggregate, which is also influenced by the membrane composition (DOPE/DOPG versus DOPE/DOPG/CL) and can be related to the previously observed difference in the ability of the peptides to disrupt different types of model membranes.

category: biochemistry


title: Chaphamaparvovirus Antigen and Nucleic Acids are not Detected in Kidney Tissues from Cats with Chronic Renal Disease or Immunosuppressive Diseases

abstract: Mouse Kidney Parvovirus (MKPV) was recently recognized as the cause of murine inclusion body nephropathy, a disease reported for over 40 years in laboratory mice. Immunodeficient mice are persistently infected with MKPV, leading to chronic renal disease, morbidity and mortality whereas immunocompetent mice seroconvert with mild renal pathology. Given the high incidence of MKPV infection in wild mice in the New York City area, the first goal of this study was to evaluate the possibility of MKPV involvement in feline chronic kidney disease (CKD) from the same geographic region. As MKPV and related parvoviruses recently described in other animal species appear to have a tropism for kidney tissue, the second goal was to investigate the possible role of a virus of this group, other than MKPV, in the development of feline CKD, Presence of MKPV and related viruses was investigated in feline renal samples using PCR, RNA in situ hybridization (ISH) and immunohistochemistry (IHC). Cats were divided into three groups: normal (N=25), CKD (N=25) and immune suppressed (N=25). None of the kidney tissues from any of the 75 cats revealed the presence of MKPV DNA, RNA or antigen expression. Nor was "fechavirus" detected using PCR in renal tissue from cats with chronic kidney disease. We conclude that MKPV is an unlikely cause or contributor to feline CKD.

category: pathology


title: Acitretin mitigates uroporphyrin-induced bone defects in congenital erythropoietic porphyria models

abstract: Congenital erythropoietic porphyria (CEP) is a rare genetic disorder leading to accumulation of uro/coproporphyrin-I in tissues due to inhibition of uroporphyrinogen-III synthase. Clinical manifestations of CEP include bone fragility, severe photosensitivity and photomutilation. Currently there is no specific treatment for CEP, except bone marrow transplantation, and there is an unmet need for treating this orphan disease. Fluorescent porphyrins cause protein aggregation, which led us to hypothesize that uroporphyrin-I accumulation leads to protein aggregation and CEP-related bone phenotype. We developed a zebrafish model that phenocopies features of CEP. As in human patients, uroporphyrin-I accumulated in the bones of zebrafish, leading to impaired bone development. Furthermore, in an osteoblast-like cell line, uroporphyrin-I decreased mineralization, aggregated bone matrix proteins, activated endoplasmic reticulum stress and disrupted autophagy. Using high-throughput drug screening, we identified acitretin, a second-generation retinoid, and showed that it reduced uroporphyrin-I accumulation and its deleterious effects on bones. Our findings provide a new CEP experimental model and a potential repurposed therapeutic.


title: Understanding binary classifier model structure based on Shapley feature interaction patterns

abstract: There is increasing emphasis on the interpretability of machine learning models, including in understanding biological systems. The well-known Shapley value framework based on game theory works in principle with any models to attribute feature importance. While feature interactions are critical to understand and can be interpreted within this framework, much attention is paid in practice on global feature importance and general trends of interactions. The inter-relationships between underlying model structure and Shapley value and its decomposition is less clear. Here we use binary classifiers to systematically examine how logical and additive interactions affect marginal contributions. These decomposed main and interaction effects are reflected in resulting Shapley dependence plots. The directionality of inequalities or logical/additive operators influence independently the main and marginal/interaction effects. Lastly, we show that these principles are applicable for models with noise.

category: systems biology


title: The ascending arousal system shapes low-dimensional brain dynamics to mediate awareness of changes in intrinsic cognitive states

abstract: Models of cognitive function typically focus on the cerebral cortex and hence overlook functional links to subcortical structures. This view neglects the highly-conserved ascending arousal systems role and the computational capacities it provides the brain. In this study, we test the hypothesis that the ascending arousal system modulates cortical neural gain to alter the low-dimensional energy landscape of cortical dynamics. Our analyses of spontaneous functional magnetic resonance imaging data and phasic bursts in both locus coeruleus and basal forebrain demonstrate precise time-locked relationships between brainstem activity, low-dimensional energy landscapes, network topology, and spatiotemporal travelling waves. We extend our analysis to a cohort of experienced meditators and demonstrate locus coeruleus-mediated network dynamics were associated with internal shifts in conscious awareness. Together, these results present a novel view of brain organization that highlights the ascending arousal systems role in shaping both the dynamics of the cerebral cortex and conscious awareness.

category: neuroscience


title: Ultra-sensitive Protein-SIP to quantify activity and substrate uptake in microbiomes with stable isotopes

abstract: Stable isotope probing (SIP) approaches are a critical tool in microbiome research to determine associations between species and substrates. The application of these approaches ranges from studying microbial communities important for global biogeochemical cycling to host-microbiota interactions in the intestinal tract. Current SIP approaches, such as DNA-SIP or nanoSIMS, are limited in terms of sensitivity, resolution or throughput. Here we present an ultra-sensitive, high-throughput protein-based stable isotope probing approach (Protein-SIP). It allows for the determination of isotope incorporation into microbiome members with species level resolution using standard metaproteomics LC-MS/MS measurements. The analysis has been implemented as an open-source application (https://sourceforge.net/projects/calis-p/). We demonstrate sensitivity, precision and accuracy using bacterial cultures and mock communities with different labeling schemes. Finally, we measure translational activity using heavy water labeling in a 63-species community derived from human fecal samples grown on media simulating two different diets. Activity could be quantified on average for 27 species per sample, with 9 species showing significantly higher activity on a high protein diet, as compared to a high fiber diet. Surprisingly, among the species with increased activity on high protein were several Bacteroides species known as fiber consumers. Apparently, protein supply is a critical consideration when assessing growth of intestinal microbes on fiber, including fiber based prebiotics. In summary, we demonstrate that our Protein-SIP approach allows for the ultra-sensitive (0.01% to 10% label) detection of stable isotopes of elements found in proteins, using standard metaproteomics data.

category: microbiology


title: Hybrid histidine kinase BinK represses Vibrio fischeri biofilm signaling at multiple developmental stages

abstract:
The symbiosis between the Hawaiian bobtail squid, Euprymna scolopes, and its exclusive light-organ symbiont, Vibrio fischeri, provides a natural system in which to study host-microbe specificity and gene regulation during the establishment of a mutually-beneficial symbiosis. Colonization of the host relies on bacterial biofilm-like aggregation in the squid mucus field. Symbiotic biofilm formation is controlled by a two-component signaling (TCS) system consisting of regulators RscS-SypF-SypG, which together direct transcription of the Syp symbiotic polysaccharide. TCS systems are broadly important for bacteria to sense environmental cues and then direct changes in behavior. Previously, we identified hybrid histidine kinase BinK as a strong negative regulator of V. fischeri biofilm regulation, and here we further explore the function of BinK. To inhibit biofilm formation, BinK requires the predicted phosphorylation sites in both the histidine kinase (H362) and receiver (D794) domains. Furthermore, we show that RscS is not essential for host colonization when binK is deleted from strain ES114, and imaging of aggregate size revealed no benefit to the presence of RscS in a background lacking BinK. Strains lacking RscS still suffered in competition. Finally, we show that BinK functions to inhibit biofilm gene expression in the light organ crypts, providing evidence for biofilm gene regulation at later stages of host colonization. Overall, this study provides direct evidence for opposing activities of RscS and BinK and yields novel insights into biofilm regulation during the maturation of a beneficial symbiosis.

IMPORTANCEBacteria are often in a biofilm state, and transitions between planktonic and biofilm lifestyles are important for pathogenic, beneficial, and environmental microbes. The critical nature of biofilm formation during Vibrio fischeri colonization of the Hawaiian bobtail squid light organ provides an opportunity to study development of this process in vivo using a combination of genetic and imaging approaches. The current work refines the signaling circuitry of the biofilm pathway in V. fischeri, provides evidence that biofilm regulatory changes occur in the host, and identifies BinK as one of the regulators of that process. This study provides information about how bacteria regulate biofilm gene expression in an intact animal host.


title: To catch a hijacker: abundance, evolution and genetic diversity of P4-like bacteriophage satellites

abstract: Bacteriophages (phages) are bacterial parasites that can themselves be parasitized by phage satellites. The molecular mechanisms used by satellites to hijack phages are sometimes understood in great detail, but the origins, abundance, distribution, and composition of these elements are poorly known. Here, we show that P4-like elements are present in more than 10% of the genomes of Enterobacterales, and in almost half of those of Escherichia coli, sometimes in multiple distinct copies. We identified over 1000 P4-like elements with very conserved genetic organization of the core genome and a few hotspots with highly variable genes. These elements are never found in plasmids and have very little homology to known phages, suggesting an independent evolutionary origin. Instead, they are scattered across chromosomes, possibly because their integrases are often exchanged with other elements. The rooted phylogenies of hijacking functions are correlated and suggest longstanding co-evolution. They also reveal broad host ranges in P4-like elements, since almost identical elements can be found in distinct bacterial genuses. Our results show that P4-like phage satellites constitute a very distinct, widespread and ancient family of mobile genetic elements. They pave the way for studying the molecular evolution of antagonistic interactions between phages and their satellites.


title: Impact of blood storage conditions on the transcript profile of plasma cell-free RNA

abstract:
BACKGROUNDPlasma cell-free RNA (cfRNA) are potential biomarkers for disease prediction and diagnosis. However, pre-analysis factors, such as the delay in blood processing and storage may lead to unreliable results, though no study has systematically evaluated the effect of blood storage conditions on the whole transcriptome of plasma cfRNA yet.

METHODSWe collected peripheral blood samples from four healthy subjects and allowed them to stand at room temperature or 4{square} for different time periods (0h, 2h, 6h and 24h) prior to plasma separation. Then, plasma cfRNA stability was evaluated by measuring expression changes of cell-free mRNA, lncRNA and miRNA using high throughput sequencing-based profiling. Finally, their paired leukocyte RNA data were integrated to depict the effect of leukocytes on plasma cfRNA during storage.

RESULTSPlasma mRNA and lncRNA presented high correlations (Pearson R2 [≥] 0.8) and fewer variations when blood was stored at 4{square} for 6 hours or stored at RT for 2 hours. miRNA was more stable, with minimal R2 of 0.86 at 4{square} for at least 24 hours or at RT for 6 hours. Correlations of plasma RNA and leukocyte RNA increased with the incubation time, and the relative proportion of neutrophils in plasma grown from 14.3% to 61.2% at RT (P = 0.004), indicating leukocyte RNA contamination. Besides, the tissue enriched genes in plasma were down-regulated with the extension of storage time.

CONCLUSIONSOur results characterized the effects of short-term storage of blood samples on plasma cfRNA, which will facilitate further researches or clinical applications to avoid bias resulting from sample processing.


title: Targeted gene sequencing with Nanopore enables rapid and accurate confirmatory diagnostic of Tilapia lake virus

abstract: Infectious diseases represent one of the major challenges to sustainable aquaculture production. Rapid, accurate diagnosis and genotyping of emerging pathogens during early-suspected disease cases is critical to facilitate timely response to deploy adequate control measures and prevent or reduce spread. Currently, most laboratories use PCR to amplify partial pathogen genomic regions, occasionally combined with sequencing of PCR amplicon(s) using conventional Sanger sequencing services for confirmatory diagnosis. The main limitation of this approach is the lengthy turnaround time. Here, we report an innovative approach using a previously developed specific PCR assay for pathogen diagnosis combined with a new Oxford Nanopore Technologies (ONT)-based amplicon sequencing method for pathogen genotyping. Using fish clinical samples, we applied this approach for the rapid confirmation of PCR amplicon sequences identity and genotyping of tilapia lake virus (TiLV), a disease-causing virus affecting tilapia aquaculture globally. The consensus sequences obtained after polishing exhibit strikingly high identity to references derived by Illumina and Sanger methods (99.83-100%). This study suggests that ONT-based amplicon sequencing is a promising platform to deploy in regional aquatic animal health diagnostic laboratories in low and medium income countries, for fast identification and genotyping of emerging infectious pathogens from field samples within a single day.


title: Rapid genotyping of tilapia lake virus (TiLV) using Nanopore sequencing


title: Inhibition of selective autophagy by members of the herpesvirus ubiquitin-deconjugase family

abstract: Autophagy is an important component of the innate immune response that restricts infection by different types of pathogens. Viruses have developed multiple strategies to avoid autophagy to complete their replication cycle and promote spreading to new hosts. Here we report that the ubiquitin deconjugases encoded in the N-terminal domain of the large tegument proteins of Epstein-Barr virus (EBV), Kaposi Sarcoma herpesvirus (KSHV) and human cytomegalovirus (HCMV), but not herpes simplex virus-1 (HSV-1), regulate selective autophagy by inhibiting the activity of the autophagy receptor SQSTM1/p62. We found that all the homologs bind to and deubiquitinate SQSTM1/p62 but with variable efficiency, which correlates with their capacity to prevent the colocalization of LC3 with SQSTM1/p62 aggregates and promote the accumulation of a model autophagy substrate. The findings highlight important differences in the strategies by which herpesviruses interfere with selective autophagy.


title: Whole-genome-based Helicobacter pylori geographic surveillance: a visualized and expandable webtool

abstract: Helicobacter pylori exhibits specific geographic distributions that related to the clinical outcomes. Despite the high infection rate of H. pylori throughout the world, the genetic epidemiology surveillance of H. pylori still needs to be improved. Here, we used single nucleotide polymorphisms (SNPs) profiling approach based on whole genome sequencing (WGS) that facilitates genomic population analyses of H. pylori and encourages the dissemination of microbial genotyping strategies worldwide. A total number of 1,211 public H. pylori genomes were downloaded and used to construct the typing tool, named as HPTT (H. pylori Typing Tool). Combined with the metadata, we developed two levels of genomic typing, including a continent scale and a country scale that nested in the continent scale. Results showed that Asia was the largest isolates source in our dataset, while isolates from Europe and Oceania were comparatively more widespread. More specifically, Switzerland and Australia are the main source of widespread isolates in their corresponding continents. To integrate all the typing information and enable researchers to compare their own dataset against the existing global database in an easy and rapid way, a user-friendly website (https://db.cngb.org/HPTT/) was developed with both genomic typing tool and visualization tool. To further confirm the validity of the website, ten newly assembled genomes were downloaded and tested precisely located on the branch as we expected. In summary, H. pylori typing tool (HPTT) is a novel genomic epidemiological tool that can achieve high resolution analysis of genomic typing and visualizing simultaneously, providing insights into the genetic population structure analysis, evolution analysis and epidemiological surveillance of H. pylori.


title: A systematic structural comparison of all solved small proteins (4-6 kDa) reveals the weight of disulfide bonds in proteins' foldability

abstract: Defensins are small proteins, usually ranging from 4 to 6 kDa, amphipathic, disulfide-rich, and with a small or even absent hydrophobic core. Since a hydrophobic core is generally found in globular proteins that fold in an aqueous solvent, the peculiar fold of defensins can challenge tertiary protein structure predictors. We performed a PDB-wide survey of small proteins (4-6 kDa) to understand the similarities of defensins with other small disulfide-rich proteins. We found no differences when we compared defensins with non-defensins regarding the proportion and exposition to the solvent of apolar, polar, and charged residues. Then we divided all small proteins (4-6 kDa) deposited in PDB into two groups, one group with at least one disulfide bond (bonded, defensins included) and another group without any disulfide bond (unbonded). The group of bonded proteins presented apolar residues more exposed to the solvent than the unbonded group. The ab initio algorithm for tertiary protein structure prediction Robetta was more accurate to predict unbonded than bonded proteins. Our work highlights one more layer of complexity for the tertiary protein prediction structure: small disulfide-rich proteins ability to fold even with a poor hydrophobic core.


title: Spermiogenesis alterations in the absence of CTCF revealed by single cell RNA sequencing

abstract: CTCF is an architectonical protein that organizes the genome inside the cell nucleus in almost all eukaryotic cells. There is evidence that CTCF plays a critical role during spermatogenesis as its depletion produces abnormal sperm and infertility. However, the defects produced by the absence of CTCF throughout spermatogenesis have not been characterized. In this work, we performed single cell RNA sequencing in spermatogenic cells without CTCF. We uncovered defects in transcriptional programs that explain the severity of the damage in the produced sperm. At early stages of spermatogenesis, transcriptional alterations are mild. As germ cells go throughout the specialization stage or spermiogenesis, transcriptional profiles become more altered. We found spermatid defects that support the alterations in the transcriptional profiles, and thus we conclude that CTCF depletion alters several transcriptional profiles mostly during spermiogenesis. Our data highlights the importance of CTCF at the different stages of spermatogenesis.


title: Epigenetic clock and methylation studies in dogs

abstract: DNA methylation profiles have been used to develop biomarkers of aging known as epigenetic clocks, which predict chronological age with remarkable accuracy and show promise for inferring health status as an indicator of biological age. Epigenetic clocks were first built to monitor human aging but the principles underpinning them appear to be evolutionarily conserved, as they have now been successfully developed for over 120 mammalian species. Here we describe reliable and highly accurate epigenetic clocks shown to apply to 51 domestic dog breeds. The methylation profiles were generated using a custom array with DNA sequences that are conserved across all mammalian species (HorvathMammalMethylChip40). Canine epigenetic clocks were constructed to estimate age and also predict mortality risk by estimating average time-to-death. We also present two highly accurate human-dog dual species epigenetic clocks (R=0.97), which may facilitate the ready translation from canine to human use (or vice versa) of anti-aging treatments being developed for longevity and preventive medicine. Finally, epigenome-wide association studies herein reveal individual methylation sites that may underlie the inverse relationship between breed weight and lifespan. Overall, we describe robust biomarkers to measure aging and potentially health status in canines.

category: developmental biology


title: Osteoblast-Specific Wnt Secretion is Required for Skeletal Homeostasis and Loading-Induced Bone Formation in Adult Mice

abstract: Wnt signaling is critical to many aspects of skeletal regulation, but the importance of Wnt ligands in adult bone homeostasis and the anabolic response to mechanical loading is not well documented. We inhibited Wnt ligand secretion in adult (5-mo) mice using a systemic (drug) and a bone-targeted (genetic) approach, and subjected them to axial tibial loading to induce lamellar bone formation. Mice treated with the porcupine inhibitor WNT974 exhibited a decrease in bone formation in non-loaded limbs as well as a 54% decline in the periosteal bone formation response to tibial loading. Similarly, within 1-2 weeks of Wls deletion in osteoblasts (Osx-CreERT2;WlsF/F mice), skeletal homeostasis was altered with decreased bone formation and increased resorption, and the anabolic response to loading was reduced 65% compared to control (WlsF/F). These findings establish a requirement for Wnt ligand secretion by osteoblasts for adult bone homeostasis and the anabolic response to mechanical loading.

category: genetics


title: Deficiency of SYCP3-related XLR3 Disrupts the Initiation of Meiotic Sex Chromosome Inactivation in Mouse Spermatogenesis

abstract:
In mammals, the X and Y chromosomes share only small regions of homology called pseudo-autosomal regions (PAR) where pairing and recombination in spermatocytes can occur. Consequently, the sex chromosomes remain largely unsynapsed during meiosis I and are sequestered in a nuclear compartment known as the XY body where they are transcriptionally silenced in a process called meiotic sex chromosome inactivation (MSCI). MSCI mirrors meiotic silencing of unpaired chromatin (MSUC), the sequestration and transcriptional repression of unpaired DNA observed widely in eukaryotes. MSCI is initiated by the assembly of the axial elements of the synaptonemal complex (SC) comprising the structural proteins SYCP2 and SYCP3 followed by the ordered recruitment of DNA Damage Response (DDR) factors to effect gene silencing. However, the precise mechanism of how unsynapsed chromatin is detected in meiocytes is poorly understood. The sex chromosomes in eutherian mammals harbor multiple clusters of SYCP3-like amplicons comprising the Xlr gene family, only a handful of which have been functionally studied. We used a shRNA-transgenic mouse model to create a deficiency in the testis-expressed multicopy Xlr3 genes to investigate their role in spermatogenesis. Here we show that knockdown of Xlr3 in mice leads to spermatogenic defects and a skewed sex ratio that can be traced to MSCI breakdown. Spermatocytes deficient in XLR3 form the XY body and the SC axial elements therein, but are compromised in their ability to recruit DDR components to the XY body.

Author SummaryA key event in the production of sperm is the pairing and synapsis of homologous chromosome pairs to facilitate genetic recombination during the first meiotic division. Chromosomal abnormalities that undermine pairing and synapsis in spermatocytes trigger a checkpoint that leads to removal of the abnormal cells via programmed cell death. In mammals, the sex chromosomes, X and Y, lack homology through most of their length and remain largely unpaired. To avoid triggering the checkpoint the X and Y are sequestered in a specialized nuclear compartment called the XY body. DNA damage response (DDR) proteins are recruited to the XY body in a highly ordered progression leading to repression of all gene transcription from the sex chromosomes. We show that knocking down expression of the X-linked SYCR3-like gene, Xlr3, disrupts sex chromosome gene silencing by interfering with recruitment of DDR factors, leading to compromised sperm production.


title: The nuclear receptor DHR3/Hr46 is required in the blood brain barrier of mature males for courtship

abstract:
The blood brain barrier (BBB) forms a stringent barrier that protects the brain from components in the circulation that could interfere with neuronal function. At the same time, the BBB enables selective transport of critical nutrients and other chemicals to the brain. Many of these processes are still poorly understood. Beyond these functions, another recently recognized function is even less characterized, specifically the role of the BBB in modulating behavior by affecting neuronal function in a sex dependent manner. Notably, signaling in the adult Drosophila BBB is required for normal male courtship behavior. Courtship regulation also relies on male-specific molecules in the BBB. Our previous studies have demonstrated that adult feminization of these cells in males significantly lowered courtship. Here, we conducted microarray analysis of BBB cells isolated from males and females. Findings revealed that these cells contain male- and female-enriched transcripts, respectively. Among these transcripts, nuclear receptor DHR3/Hr46 was identified as a male-enriched BBB transcript. DHR3/Hr46 is best known for its essential roles in the ecdysone response during development and metamorphosis. In this study, we demonstrate that DR3/Hr46 is specifically required in the BBB cells of mature males for courtship behavior. The protein is localized in the nuclei of sub-perineurial glial cells (SPG), indicating that it might act as a transcriptional regulator. These data provide a catalogue of sexually dimorphic BBB transcripts and demonstrate a physiological adult role for the nuclear receptor DH3/Hr46 in the regulation of male courtship, a novel function that is independent of its developmental role.

Author summaryThe blood brain barrier very tightly regulates which molecules can enter the brain. This is an important protection for the brain, however, it also complicates communication between molecules in the circulating fluid and the brain. In fly courtship, for example, circulating male-specific products are crucially required for normal courtship. But the neuronal circuits that ultimately control the behavior are inside the brain, separated from these molecules by the blood brain barrier. The mechanisms of this communication are not known. Here we show that the blood brain barrier itself contains sex-specific RNAs and we show that one of them, a nuclear receptor called DHR3, is required in adult males for normal courtship. These findings promise new insight into the communication between blood brain barrier and the brain.

category: animal behavior and cognition


title: Epigenome erosion drives neural crest-like phenotypic mimicry in triple-negative breast cancer and other SOX10+ malignancies

abstract:
AO_SCPLOWBSTRACTC_SCPLOWO_ST_ABSBackgroundC_ST_ABSIntratumoural heterogeneity is a poor prognostic feature in triple-negative breast cancer (TNBC) and other high-grade malignancies. It is caused by genomic instability and phenotypic plasticity, but how these features co-evolve during tumour development remains unclear. SOX10 is a transcription factor, neural crest stem cell (NCSC) specifier and candidate mediator of cancer-associated phenotypic plasticity.

MethodsUsing immunophenotyping, we investigated the expression of SOX10 in normal human breast tissue and breast cancer (n=21 cosmetic breast reduction and 1,860 tumour samples with clinical annotation). We then defined the context and evolution of its expression in TNBC compared to 21 other malignancies using systems-level transcriptomics.

ResultsSOX10 was detected in nuclei of normal mammary luminal progenitor cells, the histogenic origin of most TNBCs. In breast cancer, nuclear SOX10 predicted poor outcome amongst cross-sectional (log-rank p=0.0015, hazard ratio 2.02, n=224) and metaplastic (log-rank p=0.04, n=66) TNBCs. Systems-level transcriptional network analysis identified a core module in SOX10s normal mammary epithelial transcription program that is rewired to NCSC genes in TNBC. Reprogramming was proportional to DNA damage and genome-wide promoter hypomethylation, particularly at CpG island shores. Using a novel network analysis pipeline, we found that NCSC-like transcriptional reprogramming is also strongly associated with promoter hypomethylation in other SOX10+ malignancies: glioma and melanoma.

ConclusionsWe propose that cancer-associated genome hypomethylation simulates the open chromatin landscape of more primitive cell states, and that on this relatively unrestricted background, SOX10 recreates its ancestral gene regulatory circuits by default. These findings provide new insights about the basis of intratumoural heterogeneity and resurrection of developmental phenotypes in cancer; and highlight the potential for therapeutics that limit chromatin remodelling.

category: cancer biology


title: Novel insights in the pathophysiology of α-synuclein dysregulation on D2 receptor activity contributing to the vulnerability of dopamine neurons

abstract: Pathophysiological damages and loss of function of dopamine neurons precedes their demise and contributes to the early phases of Parkinsons disease. The presence of aberrant intercellular pathological inclusions of the protein -synuclein within ventral midbrain dopaminergic neurons is one of the cardinal features of Parkinsons disease. We employed multiple complementary approaches in molecular biology, electrophysiology, and live-cell imaging to investigate how excessive -synuclein levels alters multiple characteristics of dopaminergic neuronal dynamics and dopamine transmission prior to neuronal demise. These studies demonstrate that -synuclein dysregulation of D2 receptor autoinhibition contributes to the vulnerability of dopaminergic neurons, and that modulation thereof can ameliorate the resulting pathophysiology. These novel findings provide mechanistic insights in the insidious loss of dopaminergic function and neurons that characterize Parkinsons disease progression with significant therapeutic implications.


title: Unaffected functional recovery after spinal cord contusions at different circadian times

abstract: The circadian rhythms of gene expression drive diurnal oscillations of physiological processes that determine the acute injury response including immunity, inflammation and hemostasis. While outcomes of various acute injuries are affected by the time of day at which the original insult occurred, such diurnal influences on recovery after spinal cord injury (SCI) are unknown. We report that several key regulators of circadian gene expression are differentially expressed in uninjured spinal cord tissue of naive mice at Zeitgeber time 1 (ZT1) or ZT12, where ZT0 or ZT12 are times when lights are turned on or off, respectively. However, mice that received moderate, T9 contusive SCI at ZT0 or ZT12 showed similar recovery of locomotion as determined using the ladder walking test and the Basso mouse scale (BMS) over a 6 week post-injury period. Consistent with those findings, terminal histological analysis revealed no significant differences in white matter sparing at the injury epicenter. Therefore, locomotor recovery after thoracic contusive SCI is not affected by the time of day at which the neurotrauma occurred at least when comparing the beginning to the end of the mouse active period.


title: Taming Strong Selection with Large Sample Sizes

abstract:
1The fate of mutations and the genetic load of populations depend on the relative importance of genetic drift and natural selection. In addition, the accuracy of numerical models of evolution depends on the strength of both selection and drift: strong selection breaks the assumptions of the nearly neutral model, and drift coupled with large sample sizes breaks Kingmans coalescent model.

Thus, the regime with strong selection and large sample sizes, relevant to the study of pathogenic variation, appears particularly daunting. Surprisingly, we find that the interplay of drift and selection in that regime can be used to define asymptotically closed recursions for the distribution of allele frequencies that are accurate well beyond the strong selection limit.

Selection becomes more analytically tractable when the sample size n is larger than twice the population-scaled selection coefficient: n [≥] 2Ns (4Ns in diploids). That is, when the expected number of coalescent events in the sample is larger than the number of selective events. We construct the relevant transition matrices, show how they can be used to accurately compute distributions of allele frequencies, and show that the distribution of deleterious allele frequencies is sensitive to details of the evolutionary model.


title: Fine-tuning of SUMOylation modulates drought tolerance

abstract: SUMOylation is involved in various aspects of plant biology, including drought stress. However, the relationship between SUMOylation and drought stress tolerance is complex; whether SUMOylation has a crosstalk with ubiquitination in response to drought stress remains largely unclear. In this study, we found that both increased and decreased SUMOylation led to increased survival of apple (Malus x domestica) under drought stress: both transgenic MdSUMO2A overexpressing (OE) plants and MdSUMO2 RNAi plants exhibited enhanced drought tolerance. We further confirmed that MdDREB2A is one of the MdSUMO2 targets. Both transgenic MdDREB2A OE and MdDREB2AK192R OE plants (which lacked the key site of SUMOylation by MdSUMO2A) were more drought tolerant than wild-type plants. However, MdDREB2AK192R OE plants had a much higher survival rate than MdDREB2A OE plants. We further showed SUMOylated MdDREB2A was conjugated with ubiquitin by MdRNF4 under drought stress, thereby triggering its protein degradation. In addition, MdRNF4 RNAi plants were more tolerant to drought stress. These results revealed the molecular mechanisms that underlie the relationship of SUMOylation with drought tolerance and provided evidence for the tight control of MdDREB2A accumulation under drought stress mediated by SUMOylation and ubiquitination.

category: plant biology


title: Organelle calcium-derived voltage oscillations in pacemaker neurons drive food-seeking behavior in Aplysia

abstract: The expression of motivated behaviors depends on both external and internally-arising neural stimuli, yet the intrinsic releasing mechanisms for such variably occurring behaviors remain elusive. In isolated nervous system preparations of Aplysia, we have found that irregularly expressed cycles of motor output underlying food-seeking behavior arise from regular membrane potential oscillations of varying magnitude in an identified pair of interneurons (B63) in the bilateral buccal ganglia. This rhythmic signal, which is endogenous and specific to the B63 cells, is generated by organelle-derived intracellular calcium fluxes that activate voltage-independent plasma membrane channels. The resulting voltage oscillation spreads throughout a subset of gap junction-coupled buccal network neurons and by triggering plateau potential-mediated bursts in B63, can initiate motor output driving food-seeking action. Thus, an atypical neuronal pacemaker mechanism, based on rhythmic intracellular calcium store release and intercellular propagation, can act as an autonomous intrinsic releaser for the occurrence of a motivated behavior.


title: Population Codes Enable Learning from Few Examples By Shaping Inductive Bias

abstract: Learning from a limited number of experiences requires suitable inductive biases. To identify how inductive biases are implemented in and shaped by neural codes, we analyze sample-efficient learning of arbitrary stimulus-response maps from arbitrary neural codes with biologically-plausible readouts. We develop an analytical theory that predicts the generalization error of the readout as a function of the number of observed examples. Our theory illustrates in a mathematically precise way how the structure of population codes shapes inductive bias, and how a match between the code and the task is crucial for sample-efficient learning. We observe that many different codes can support the same inductive bias. By analyzing recordings from the mouse primary visual cortex, we demonstrate that biological codes have lower total activity than other codes with identical bias. Using these mouse primary visual cortex responses, we demonstrate the existence of an efficiency bias towards low frequency orientation discrimination tasks for grating stimuli and low spatial frequency reconstruction tasks for natural images. We reproduce the discrimination bias in a simple model of primary visual cortex, and further show how invariances in the code to certain stimulus variations alter learning performance. We extend our methods to time-dependent neural codes and predict the sample efficiency of readouts from recurrent networks. Finally, we discuss implications of our theory in the context of recent developments in neuroscience and artificial intelligence. Overall, our study provides a concrete method for elucidating inductive biases of the brain and promotes sample-efficient learning as a general normative coding principle.


title: Genome sequencing and functional genes comparison between Sphingopyxis USTB-05 and Sphingomonas morindae NBD5

abstract:
Sphingomonadaceae has a large number of strains that can biodegrade hepatotoxins or environmental pollutants. The latest research reported that certain strains can also produce lutein. Based on the third-generation sequencing technology, we analyzed the whole genome sequence and compared related functional genes of two strains of Sphingomonadaceae isolated from different habitats. The genome of Sphingopyxis USTB-05 was 4,679,489 bp and contained 4312 protein coding genes. The 4,239,716 bp nuclear genome of Sphingomonas morindae NBD5, harboring 3882 protein coding genes, has two sets of chromosomes. Both strains had lutein synthesis metabolism pathway sharing identical synthetic genes of crtB, crtE, crtI, crtQ, crtL, crtR, atoB, dxs, dxr, ispD, ispE, ispDF, gcpE, ispG, ispH, ispA, ispB and ispU. Sphingopyxis USTB-05 had hepatotoxins microcystins and nodularin metabolic pathways related to 16 genes (ald, ansA, gdhA, crnA, phy, ocd, hypdh, spuC, nspC, speE, murI, murD, murC, hmgL, bioA and glsA), while these genes were not found in Sphingomonas morindae NBD5. The unique protein sequences of strain NBD5 and strain USTB-05 were 155 and 199, respectively. The analysis of whole genome of the two Sphingomonadaceae strains provides insights into prokaryote evolution, the new pathway for lutein production and the new genes for environmental pollutant biodegradation.

IMPORTANCEUnderstanding the functional genes related to the special functions of strains is essential for humans to utilize microbial resources. The ability of Sphingopyxis USTB-05 to degrade hepatotoxins microcystins and nodularin has been studied in depth, however the complete metabolic process still needs further elucidation. Sphingomonas morindae NBD5 can produce lutein, and it is necessary to determine whether there is a new pathway of lutein. In this study, the whole genome sequencing of Sphingopyxis USTB-05 and Sphingomonas morindae NBD5 were performed for the first time. Lutein synthesis metabolic pathways and synthetic genes were discovered in Sphingomonadaceae. We predicted the existence of new lutein synthesis pathways and revealed most of the genes of the new synthesis pathways. A comparative analysis of the functional genes of the two strains revealed that Sphingopyxis USTB-05 contains a large number of functional genes related to the biodegradation of hepatotoxins or hexachlorocyclohexane. Among them, the functional genes related to the biodegradation and metabolism of hexachlorocyclohexane had not been previously reported. These findings lay the foundation for the biosynthesis of lutein using Sphingomonas morindae NBD5 or Sphingopyxis USTB-05 and the application of Sphingopyxis USTB-05 for the biodegradation of hepatotoxins microcystins and nodularin or environmental pollutants.


title: Extracellular vesicles synchronize cellular phenotypes of differentiating cells

abstract: During embryonic development, cells differentiate in a coordinated manner, aligning their fate decisions and differentiation stages with those of surrounding cells. However, little is known about the mechanisms that regulate this synchrony. Here we show that cells in close proximity synchronize their differentiation stages and cellular phenotypes with each other via extracellular vesicle (EV)-mediated cellular communication. We previously established a mouse embryonic stem cell (ESC) line harboring an inducible constitutively active protein kinase A (CA-PKA) gene and found that the ESCs rapidly differentiated into mesoderm after PKA activation. In the present study, we performed a co-culture of control ESCs and PKA-ESCs, finding that both ESCs rapidly differentiated in synchrony even when PKA was activated only in PKA-ESCs, a phenomenon we named "Phenotypic Synchrony of Cells (PSyC)". We further demonstrated PSyC was mediated by EVs containing miR-132. PKA-ESC-derived EVs and miR-132-containing artificial nano-vesicles similarly enhanced mesoderm and cardiomyocyte differentiation in ESCs and ex vivo embryos, respectively. PSyC is a new form of cell-cell communication mediated by EV regulation of neighboring cells and could be broadly involved in tissue development and homeostasis.


title: Arf GTPase-Activating proteins SMAP1 and AGFG2 regulate the size of Weibel-Palade bodies and exocytosis of von Willebrand factor

abstract:
Arf GTPase-Activating proteins (ArfGAPs) mediate the hydrolysis of GTP bound to ADP-ribosylation factors, which are important for intracellular transport. ArfGAPs have been shown to be critical for cargo sorting in the Golgi-to-ER and post-Golgi traffic. However, their roles in the sorting of secretory proteins remains unclear. Weibel-Palade bodies (WPBs) are cigar-shaped secretory granules in endothelial cells that contain von Willebrand factor (vWF) as their main cargo. WPBs are formed at the trans-Golgi Network, and this process is thought to be coupled with the sorting of vWF. WPB biogenesis was reported to be regulated by ADP-ribosylation factors and their regulators, but the role of ArfGAPs has been unknown. In this study, we performed siRNA screening of ArfGAPs to investigate the biogenesis of WPBs. We found two ArfGAPs, SMAP1 and AGFG2, to be involved in WPB size and vWF exocytosis, respectively. SMAP1 depletion resulted in small-sized WPBs, and the lysosomal inhibitor leupeptin recovered the size of WPBs. These results indicate that SMAP1 functions in preventing the degradation of cigar-shaped WPBs. However, AGFG2 downregulation resulted in the inhibition of vWF secretion upon Phorbol 12-myristate 13-acetate (PMA)-stimulation, suggesting that AGFG2 plays a role in vWF exocytosis. Our study revealed unexpected processes regulated by ArfGAPs for vWF transport.

Summary StatementThe ArfGAP proteins SMAP1 and AGFG2 were identified as regulating WPB size and vWF exocytosis.


title: Expression of Sonic Hedgehog and Pathway Components in the Embryonic Mouse Head: Anatomical Relationships Between Regulators of Positive and Negative Feedback

abstract:
ObjectiveThe Hedgehog pathway is a fundamental signaling pathway in organogenesis. The expression patterns of the ligand Sonic Hedgehog (Shh) and key pathway components have been studied in many tissues but direct spatial comparisons across tissues with different cell compositions and structural organization are not common and could reveal tissue-specific differences in pathway dynamics.

ResultsWe directly compared the expression characteristics of Shh, and four genes with functional roles in signaling and whose expression levels serve as readouts of pathway activity in multiple tissues of the embryonic mouse head at embryonic day 15.5 by serial in situ hybridization. The four readout genes were the positive feedback regulator Gli1, and three negative feedback regulators, Patched1, Patched2, and Hedgehog Interacting Protein. While the relative abundance of Gli1 was similar across tissues, the relative expression levels and spatial distribution of Shh and the negative feedback regulators differed, suggesting that feedback regulation of hedgehog signaling is context dependent. This comparative analysis offers insight into how consistent pathway activity could be achieved in tissues with different morphologies and characteristics of ligand expression.


title: Contact-tracing in cultural evolution: a Bayesian mixture model to detect geographic areas of language contact

abstract: When speakers of two or more languages interact, they are likely to influence each other: contact leaves traces in the linguistic record, which in turn can reveal geographic areas of past human interaction and migration. However the complex, multi-dimensional nature of contact has hindered the development of a rigorous methodology for detecting its traces. Specifically, other factors may contribute to similarities between languages. Inheritance (a property is passed from an ancestor to several descendant languages), and universal preference (a property is universally preferred), may both overshadow contact signals. How can we find geographic contact areas in language data, while accounting for the confounding effects of inheritance and universal preference? We present sBayes, an algorithm for Bayesian clustering in the presence of confounding effects. The algorithm learns which similarities in a set of features are better accounted for by confounders, and which are due to contact effects. Contact areas are free to take any shape or size, but an explicit geographic prior ensures their spatial coherence. We test the clustering method on simulated data and apply it in two case studies to reveal language contact in South America and the Balkans. Our results are supported by --mostly qualitative-- findings from previous studies. While we focus on the specific problem of language contact, the method can also be used to uncover other traces of shared history in cultural evolution, and more generally, to reveal latent spatial clusters in the presence of confounders.

category: evolutionary biology


title: Dopamine facilitates fear memory formation in the human amygdala

abstract: Learning which environmental cues that predict danger is crucial for survival and accomplished through Pavlovian fear conditioning. In humans and rodents alike, fear conditioning is amygdala-dependent and rests on similar neurocircuitry. Rodent studies have implicated a causative role for dopamine in the amygdala during fear memory formation, but the role of dopamine in aversive learning in humans is unclear. Here, we show dopamine release in the amygdala and striatum during fear learning in humans. Using simultaneous positron emission tomography and functional magnetic resonance imaging, we demonstrate that the amount of dopamine release is linked to strength of conditioned fear responses and linearly coupled to learning-induced memory trace activity in the amygdala. Thus, like in rodents, formation of amygdala-dependent fear memories in humans seems to be facilitated by endogenous dopamine release, supporting an evolutionary conserved neurochemical mechanism for aversive memory formation.


title: Micrococcal nuclease sequencing of porcine sperm suggests a nucleosomal involvement on semen quality and early embryo development

abstract:
BackgroundThe mammalian mature spermatozoon has a unique chromatin structure in which the vast majority of histones are replaced by protamines during spermatogenesis and a small fraction of nucleosomes are retained at specific locations of the genome. The chromatin structure of sperm remains unresolved in most livestock species, including the pig. However, its resolution could provide further light into the identification of the genomic regions related to sperm biology and embryo development and it could also help identifying molecular markers for sperm quality and fertility traits. Here, for the first time in swine, we performed Micrococcal Nuclease coupled with high throughput sequencing on pig sperm and characterized the mono-nucleosomal (MN) and sub-nucleosomal (SN) chromatin fractions.

ResultsWe identified 25,293 and 4,239 peaks in the mono-nucleosomal and sub-nucleosomal fractions, covering 0.3% and 0.02% of the porcine genome, respectively. A cross-species comparison of nucleosome-associated DNAs in sperm revealed positional conservation of the nucleosome retention between human and pig. Gene ontology analysis of the genes mapping nearby the mono-nucleosomal peaks and identification of putative transcription factor binding motifs within the mono-nucleosomal peaks showed enrichment for sperm function and embryo development related processes. We found motif enrichment for the transcription factor Znf263, which in humans was suggested to be a key regulator of the genes with paternal preferential expression during early embryo development. Moreover, we found enriched co-occupancy between the RNAs present in pig sperm and the RNA related to sperm quality, and the mono-nucleosomal peaks. We also found preferential co-location between GWAS hits for semen quality in swine and the mono-nucleosomal sites identified in this study.

ConclusionsThese results suggest a clear relationship between nucleosome positioning in sperm and sperm and embryo development.


title: The truncate soft-shell clam, Mya truncata, as a biomonitor of municipal wastewater exposure and historical anthropogenic impacts in the Canadian Arctic.

abstract: Municipal wastewater is a large source of pollution to Canadian waters, yet its effects on Arctic marine ecosystems remains relatively unknown. We characterized the impacts of municipal wastewater from a growing northern community, Iqaluit, Nunavut on the Arctic truncate soft-shell clam, Mya truncata. Clams were sampled from six locations that varied in proximity to the wastewater treatment plant and shell biogeochemical analysis revealed that clams nearest the wastewater treatment plant had slower growth rates, lower carbon and oxygen stable isotope ratios, and elevated concentrations of copper and lead. A parallel analysis on mRNA expression profiles characterized M. truncatas physiological response to wastewater effluent. Clams nearest the wastewater treatment plant had significantly lower mRNA expression of genes associated with metabolism, antioxidants, molecular chaperones, and phase I and II detoxification, but had heightened mRNA expression in genes coding for enzymes that bind and remove contaminants. These results demonstrated a biological response to Iqaluits wastewater effluent and highlight M. truncatas potential to act as a biomonitor of municipal wastewater along Canadian Arctic coastlines.

category: pharmacology and toxicology


title: Robust phylogenetic position of the enigmatic hydrozoan, Margelopsis haeckelii revealed within the family Corymorphidae

abstract: The life-cycle and polyp morphology of Margelopsidae representatives are very different from all other Aplanulata cnidarians. Until recently, their evolutionary origin and phylogenetic position has been a subject of significant speculation. A recent molecular study based only on COI data unexpectedly placed Margelopsidae as a sister group to all Aplanulata, despite the Margelopsid morphology suggests affiliation with Tubulariidae or Corymorphidae. Here we used multigene analyses, including nuclear (18S rRNA and 28S rRNA) and mitochondrial (16S rRNA and COI) markers of the Margelopsidae hydroid Margelopsis haeckelii Hartlaub, 1897, to resolve its phylogenetic position with respect to other hydrozoans. Our data provides strong evidence that M. haeckelii is a member of the family Corymorphydae, making the family Margelopsidae invalid. Furthermore, we show that medusa previously known as M. hartlaubii Browne, 1903 is sister to Plotocnide borealis, Wagner, 1885 and might be a member of Boreohydridae. The phylogenetic signal of polyp and medusa stages is discussed in light of concept of inconsistent evolution and molecular phylogenetic analysis.

category: zoology


title: TGIF1 is required for chicken ovarian cortical development and generation of the juxtacortical medulla

abstract:
During early embryogenesis in amniotic vertebrates, the gonads differentiate into either ovaries or testes. The first cell lineage to differentiate gives rise to the supporting cells; Sertoli cells in males and pre-granulosa cells in females. These key cell types direct the differentiation of the other cell types in the gonad, including steroidogenic cells. The gonadal surface epithelium and the interstitial cell populations are less well studied, and little is known about their sexual differentiation programs. Here, we show the requirement of the transcription factor gene TGIF1 for ovarian development in the chicken embryo. TGIF1 is expressed in the two principal ovarian somatic cell populations, the cortex and the pre-granulosa cells of the medulla. TGIF1 expression is associated with an ovarian phenotype in sex reversal experiments. In addition, targeted over-expression and gene knockdown experiments indicate that TGIF1 is required for proper ovarian cortical formation. TGIF1 is identified as the first known regulator of juxtacortical medulla formation. These findings provide new insights into chicken ovarian differentiation and development, specifically in the process of cortical and juxtacortical medulla formation, a poorly understood area.

SUMMARY STATEMENTThe transcription factor TGIF1 is required for proper ovarian sex differentiation in chicken embryos, regulating development of the cortical and juxtacortical medulla, independently of the supporting cell sex lineage.


title: Taxonomic scale dependency of Bergmann's patterns: A cross-scale comparison of hawkmoths and birds along a tropical elevational gradient

abstract: Bergmanns rule predicts a larger body size for endothermic organisms in colder environments. The multiplicity of patterns and processes is expected because body size and temperature are two most fundamental factors on which many physiological, ecological and evolutionary processes depend, affecting all levels of biological organization, from individuals to communities. The confounding results from previous studies may be due to the differences in taxonomic (intraspecific, interspecific and community) and spatial (latitudinal vs elevational) scales. We compared Bergmanns patterns for endotherms (Aves) and ectotherms (Lepidoptera: Sphingidae) along a same 2.6 km elevational transect in the eastern Himalayas. Using a large data spanning 3,302 hawkmoths (76 morpho-species) and 15,746 birds (245 species), we compared the patterns at the intraspecific (hawkmoths only), interspecific and community scales. At the interspecific scale, we account for phylogenetic non-independence in body mass by using a heirarchical linear mixed effects model for hawkmoths, and a phylogenetic generalised least squares model for birds. We assess the importance of using abundance-weighted metrics at the community scales, after accounting for spatial auto-correlation in communities. Hawkmoths exhibited positive Bergmanns pattern at the intraspecific and abundance-weighted community scale. Intraspecific variation accounted for a substantial 33% variation at the community level. Contrary to this, birds exhibited a strong converse-Bergmanns pattern at interspecific and community scales, both with- and without-abundance. Overall, all metrics which incorporate local traits and/or species abundances show stronger correlations than when this information is lacking. The multiplicity of patterns at a single location provides the opportunity to disentangle the relative contribution of individual- and species-level processes by integrating data across multiple nested taxonomic scales for the same taxa. We suggest that future studies of Bergmanns patterns should explicitly address taxonomic- and spatial-scale dependency, with species relative abundance and intraspecific trait variation as essential ingredients especially at short elevational scales.

category: ecology


title: Influence of short-term temperature drops on sex-determination in sea turtles

abstract: All sea turtles exhibit temperature-dependent sex-determination, where warmer temperatures produce mostly females and cooler temperatures produce mostly males. As global temperatures continue to rise, sea turtle sex-ratios have become increasingly female-biased, threatening the long-term viability of many populations. Nest temperatures are dependent on sand temperature, and heavy rainfall events reduce sand temperatures for a brief period. However, it is unknown whether these short-term temperature drops are large and long enough to produce male hatchlings. To discover if short-term temperature drops within the sex-determining period can lead to male hatchling production, we exposed green and loggerhead turtle eggs to short-term temperature drops conducted in constant temperature rooms. We dropped incubation temperature at four different times during the sex-determining period for a duration of either 3 or 7 days to mimic short-term drops in temperature caused by heavy rainfall in nature. Some male hatchlings were produced when exposed to temperature drops for as little as 3 days, but the majority of male production occurred when eggs were exposed to 7 days of lowered temperature. More male hatchlings were produced when the temperature drop occurred during the middle of the sex-determining period in green turtles, and the beginning and end of the sex-determining period in loggerhead turtles. Inter-clutch variation was evident in the proportion of male hatchlings produced, indicating that maternal and or genetic factors influence male hatchling production. Our findings have management implications for the long-term preservation of sea turtles on beaches that exhibit strongly female-biased hatchling sex-ratios.


title: Extensive aquatic subsidies lead to territorial breakdown and high density of an apex predator

abstract: Energetic subsidies between terrestrial and aquatic ecosystems can strongly influence food webs and population dynamics. Our objective was to study how aquatic subsidies affected jaguar (Panthera onca) diet, sociality, and population density in a seasonally flooded protected area in the Brazilian Pantanal. The diet (n = 138 scats) was dominated by fish (46%) and aquatic reptiles (55%), representing the first jaguar population known to feed extensively on fish and to minimally consume mammals (11%). These aquatic subsidies supported the highest jaguar population density estimate to date (12.4 per 100 km2) derived from camera traps (8,065 trap nights) and GPS collars (n = 13). Contrary to their mostly solitary behavior elsewhere, we documented social interactions previously unobserved between same-sex adults including cooperative fishing, co-traveling, and play. Our research demonstrates that aquatic subsidies seen in omnivores can be highly influential to obligate carnivores leading to high population density and altered social structure.


title: Dynamics of data availability in disease modeling: An example evaluating the trade-offs of ultra-fine-scale factors applied to human West Nile virus disease models in the Chicago area, USA

abstract:
BackgroundSince 1999, West Nile virus (WNV) has moved rapidly across the United States, resulting in tens of thousands of human cases. Both the number of human cases and the minimum infection rate (MIR) in vector mosquitoes vary across time and space and are driven by numerous abiotic and biotic forces, ranging from differences in microclimates to socio-demographic factors. Because the interactions among these multiple factors affect the locally variable risk of WNV illness, it has been especially difficult to model human disease risk across varying spatial and temporal scales. Cook and DuPage Counties, comprising the city of Chicago and surrounding suburbs, experience some of the highest numbers of human neuroinvasive cases of WNV in the United States. Despite active mosquito control efforts, there is consistent annual WNV presence, resulting in more than 285 confirmed WNV human cases and 20 deaths from the years 2014-2018 in Cook County alone.

MethodsA previous Chicago-area WNV model identified the fifty-five most high and low risk locations in the Northwest Mosquito Abatement District (NWMAD), an enclave [1/4] the size of the combined Cook and DuPage county area. In these locations, human WNV risk was stratified by model performance, as indicated by differences in studentized residuals. Within these areas, an additional two-years of field collections and data processing was added to a 12-year WNV dataset that includes human cases, MIR, vector abundance, and land-use, historical climate, and socio-economic and demographic variables, and was assessed by an ultra-fine-scale (1 km spatial x 1 week temporal resolution) multivariate logistic regression model.

ResultsMultivariate statistical methods applied to the ultra-fine-scale model identified fewer explanatory variables while improving upon the fit of the previous model. Beyond MIR and climatic factors, efforts to acquire additional covariates only slightly improved model predictive performance.

ConclusionsThese results suggest human WNV illness in the Chicago area may be associated with fewer, but increasingly critical, key variables at finer scales. Given limited resources, these findings suggest large variations in model performance occur, depending on covariate availability, and provide guidance in variable selection for optimal WNV human illness modeling.


title: H3K9 methyltransferase EHMT2/G9a controls ERVK-driven non-canonical imprinted genes

abstract: Unlike regular imprinted genes, non-canonical imprinted genes are known to not depend on gamete-specific DNA methylation difference. Instead, the paternal allele-specific expression of these genes in the extra-embryonic lineages depends on an H3K27me3-based imprint in the oocyte, but this marking is not maintained beyond pre-implantation development. The maintenance of non-canonical imprinting corresponds to maternal allele-specific DNA methylation and paternal allele-specific H3K4me3 at their somatic DMRs, which occur at ERVK repeats. We hypothesized that EHMT2, the main euchromatic H3K9 methyltransferase, also has a role in this process. Using reciprocal mouse crosses and allele-specific RNA-seq analysis, we found that the maternal allele of each known non-canonical imprinted gene was derepressed from its ERVK promoter in the Ehmt2-/- ectoplacental cone of somite-matched 8.5 dpc embryos. In the Ehmt2-/- embryos, loss of DNA methylation accompanied the derepression of both parental alleles of those ERVK promoters. Our study identifies EHMT2 as an essential player that maintains the repressed chromosomal state in non-canonical imprinting.


title: Genomic analysis of patient-derived xenograft models reveals intra-tumor heterogeneity in endometrial cancer and can predict tumor growth inhibition with talazoparib

abstract:
BackgroundEndometrial cancer (EC) is a major gynecological cancer with increasing incidence. It comprised of four molecular subtypes with differing etiology, prognoses, and response to chemotherapy. In the future, clinical trials testing new single agents or combination therapies will be targeted to the molecular subtype most likely to respond. Pre-clinical models that faithfully represent the molecular subtypes of EC are urgently needed, we sought to develop and characterize a panel of novel EC patient-derived xenograft (PDX) models.

MethodsHere, we report whole exome or whole genome sequencing of 11 PDX models and the matched primary tumor. Analysis of multiple PDX lineages and passages was performed to study tumor heterogeneity across lineages and/or passages. Based on recent reports of frequent defects in the homologous recombination (HR) pathway in EC, we assessed mutational signatures and HR deficiency scores and correlated these with in vivo responses to the PARP inhibitor (PARPi) talazoparib in six PDXs representing the different molecular subtypes of EC.

ResultsPDX models were successfully generated from all four molecular subtypes of EC and uterine carcinosarcomas, and they recapitulated morphology and the molecular landscape of primary tumors without major genomic drift. We also observed a wide range of inter-tumor and intra-tumor heterogeneity, well captured by different PDX lineages, which could lead to different treatment responses. An in vivo response to talazoparib was detected in two p53mut models consistent with stable disease, however both lacked the HR deficiency genomic signature.

ConclusionsEC PDX models represent the four molecular subtypes of disease and can capture intra-tumoral heterogeneity of the original primary tumor. PDXs of the p53mut molecular subtype showed sensitivity to PARPi, however, deeper and more durable responses will likely require combination of PARPi with other agents.


title: scAMACE: Model-based approach to the joint analysis of single-cell data on chromatin accessibility, gene expression and methylation

abstract: The advancement in technologies and the growth of available single-cell datasets motivate integrative analysis of multiple single-cell genomic datasets. Integrative analysis of multimodal single-cell datasets combines complementary information offered by single-omic datasets and can offer deeper insights on complex biological process. Clustering methods that identify the unknown cell types are among the first few steps in the analysis of single-cell datasets, and they are important for downstream analysis built upon the identified cell types. We propose scAMACE for the integrative analysis and clustering of single-cell data on chromatin accessibility, gene expression and methylation. We demonstrate that cell types are better identified and characterized through analyzing the three data types jointly. We develop an efficient expectation-maximization (EM) algorithm to perform statistical inference, and evaluate our methods on both simulation study and real data applications. We also provide the GPU implementation of scAMACE, making it scalable to large datasets. The software and datasets are available at https://github.com/cuhklinlab/scAMACE_py (python implementation) and https://github.com/cuhklinlab/scAMACE (R implementation).

category: bioinformatics


title: ALOHA: Aggregated local extrema splines for high-throughput dose-response analysis

abstract: Computational methods for genomic dose-response integrate dose-response modeling with bioinformatics tools to evaluate changes in molecular and cellular functions related to pathogenic processes. These methods use parametric models to describe each genes dose-response, but such models may not adequately capture expression changes. Additionally, current approaches do not consider gene co-expression networks. When assessing co-expression networks, one typically does not consider the dose-response relationship, resulting in  co-regulated gene sets containing genes having different dose-response patterns. To avoid these limitations, we develop an analysis pipeline called Aggregated Local Extrema Splines for High-throughput Analysis (ALOHA), which computes individual genomic dose-response functions using a flexible class Bayesian shape constrained splines and clusters gene co-regulation based upon these fits. Using splines, we reduce information loss due to parametric lack-of-fit issues, and because we cluster on dose-response relationships, we better identify co-regulation clusters for genes that have co-expressed dose-response patterns from chemical exposure. The clustered pathways can then be used to estimate a dose associated with a pre-specified biological response, i.e., the benchmark dose (BMD), and approximate a point of departure dose corresponding to minimal adverse response in the whole tissue/organism. We compare our approach to current parametric methods and our biologically enriched gene sets to cluster on normalized expression data. Using this methodology, we can more effectively extract the underlying structure leading to more cohesive estimates of gene set potency.


title: Modeling High-grade serous ovarian carcinoma using a combination of in vivo electroporation and CRISPR/Cas9 mediated genome editing

abstract:
Ovarian cancer remains the most lethal gynecological cancer today. High-grade serous ovarian carcinoma (HGSC) is the most common and lethal type of ovarian cancer and is most frequently diagnosed at advanced stages. Here, we developed a novel strategy to generate somatic ovarian cancer mouse models using a combination of in vivo electroporation and CRISPR/Cas9 mediated genome editing. We mutated tumor suppressor genes associated with HGSC in two different combinations; Brca1, Tp53, Pten with/without Lkb1 and successfully generated HGSC, however, with different latencies and pathophysiology. By utilizing Cre lineage tracing in our system, we visualized peritoneal micrometastases in an immune-competent environment. Because our strategy is flexible in selecting mutation combinations and targeting areas, it would be useful for generating ovarian cancer mouse models.

Significance StatementHigh grade serous ovarian cancer (HGSC) is the most common ovarian malignancy but our knowledge of early tumorigenesis is still quite limited due to late diagnosis of patients. We developed a new strategy of generating mouse ovarian cancer models using a combination of in vivo electroporation and CRISPR-mediated genome editing. We demonstrated that a combination of three tumor suppressor gene mutations (Brca1, Tp53 and Pten) is sufficient to develop HGSCs. Interesting, an additional mutation in Lkb1 drastically changed tumor latency, penetrance and pathophysiology through changing its cell-of-origin. Our strategy is highly flexible in selection of mutation combinations and targeting areas in immune competent mice and useful to study early tumorigenesis of ovarian cancer.


title: Neocortical rhythm entrainment by parvalbumin-positive interneurons across cortical layers

abstract: Neocortical interneurons provide local inhibition responsible for organizing neuronal activity into brain oscillations that subserve several functions such as memory, attention and neuronal communication. However, little is known about the contribution of interneurons to the entrainment of neocortical oscillations across cortical layers. Here, using layer-specific optogenetic stimulations with micro-Light-Emitting-Diode (LED) arrays, directed toward parvalbumin-expressing (PV) interneurons in non-anesthetized awake mice, we find that supragranular layer stimulations of PV neurons were most efficient at entraining supragranular layer neurons and local field potential (LFP) oscillations at gamma frequencies ({gamma}: 25 - 80 Hz), whereas infragranular layer stimulation of PV neurons better entrained delta ({delta}: 2 - 5 Hz) and theta ({theta}: 6 - 10 Hz) frequency LFP oscillations. We found that PV neurons tightly control the transmission of multiple rhythms to the network across cortical layers in an orientation-selective manner. Intrinsic resonant properties of neurons could underlie such layer-specific properties of rhythm entrainment.


title: Building reusable phage and antibiotic treatments via exploitation of bacteria-phage coevolutionary dynamics.

abstract: People with chronic (long-lasting) infections face the problem that treatment options diminish in time as the pathogen evolves increasing resistance. To address this challenge, we exploit phage and bacterial co-evolution, producing dynamic selection pressures that can return the pathogen to a state of susceptibility to the initial (regulator-approved) therapy. We show that phage OMKO1 alone triggers Arms Race Dynamic (ARD) co-evolution with the pathogen Pseudomonas aeruginosa, leading to generalized phage resistance and crucially - failure at reuse. In contrast, co-administration of the phage with antibiotics triggers Fluctuating Selection Dynamics (FSD) co-evolution, allowing for effective reuse after 20 days of treatment. We pursue medical relevance in our experiments with the use of clinically important pathogens, antibiotics, phage, and a benchmarked synthetic sputum medium. Phenotypic and genomic characterization of evolved isolates demonstrates that efflux-targeting phage OMKO1 exerts continued selection for antibiotic susceptibility regardless of co-evolutionary dynamic or antibiotic co-treatment, opening the door for evolutionary robust phage therapy.


title: Are narrow-ranging species doomed to extinction? Dramatic and generalised decline in future climate suitability of highly threatened species

abstract: Narrow-ranging species are usually omitted from Species distribution models (SDMs) due to statistical constraints, while they are predicted to be particularly vulnerable to climate change. The recently available high-resolution environmental predictors, along with recently developed methods enable to increase the eligibility of narrow-ranging species for SDMs, provided their distribution is well known. We fill a gap of knowledge on the effect of predicted climate change on narrow-ranging species. We modelled the distribution of the golden mantella frog Mantella aurantiaca and the Manapany day gecko Phelsuma inexpectata, for which the distribution of their occurrence records is well documented. Our modelling scheme included a range of processes susceptible to address statistical issues related to narrow-ranging species. We predict an alarming decline in climate suitability in the whole current distribution area of both species by 2070, potentially leading to a complete extinction in most scenarios. We identified the areas with the best climate suitability in the future, but these remain largely suboptimal regarding species climatic niche. The high level of habitat fragmentation suggests that both species likely need to be at least partly translocated. Climate change may not only drive range contractions or distribution shifts in narrow-ranging species, but may lead to the complete extirpation of suitable environments across their entire region. This study suggests that the level of threats of narrow-ranging species already identified as threatened may be underestimated, especially in heterogeneous tropical areas. We stress the need to develop sampling campaigns and implement proactive actions for narrow-ranging species in the tropics.


title: Are narrow-ranging species doomed to extinction? Probable dramatic and generalised decline in future climate suitability of highly threatened species


title: Are narrow-ranging species doomed to extinction? Projected dramatic decline in future climate suitability of two highly threatened species


title: DiaThor: R package for computing diatom metrics and biotic indices

abstract:
O_LIDiatoms are widely used to detect changes in water quality due to their specific sensibility to a variety of environmental conditions. Among the different diatom-based tools to measure water quality, the biotic indices, ecological and morphological traits are the most commonly used.
C_LIO_LIDiaThor, contains 18 functions that provide morphological data of the samples (number and shape of chloroplasts, total biovolume), ecological data (species richness, evenness, diversity, size classes, ecological guilds, ecological preferences) and biotic indices (Descy Index, EPID Index, Indice Diatomique Artois-Picardie, Swiss Diatom Index, Pampean Diatom Index, ILM index, Specific Pollution sensitivity Index, Lobo Index, Sladecek index, SPEARherbicides index and the Trophic index). A web application (in Shiny) was also developed to provide access to the package for those users not familiar with the R environment.
C_LIO_LIThe structure of the package, how it functions and the structure of the input and output data are explained. To demonstrate the most common use of DiaThor, an example of the package performance is also provided.
C_LIO_LIThe DiaThor package aims to contribute to the water quality assessment based on diatom assemblages, while also providing researchers with an open platform to suggest new statistics and functionalities to be integrated into future builds.
C_LI


title: Memory specific to temporal features of sound is formed by cue-selective enhancements in temporal coding enabled by inhibition of an epigenetic regulator

abstract:
Recent investigation of memory-related functions in the auditory system have capitalized on the use of memory-modulating molecules to probe the relationship between memory and its substrates in auditory system coding. For example, epigenetic mechanisms, which regulate gene expression necessary for memory consolidation, are powerful modulators of learning-induced neuroplasticity and long-term memory formation (LTM). Inhibition of the epigenetic regulator histone deacetylase 3 (HDAC3) promotes LTM that is highly specific for spectral features of sound. The present work demonstrates for the first time that HDAC3 inhibition also enables memory for temporal features of sound. Rats trained in an amplitude modulation (AM) rate discrimination task and treated with a selective inhibitor of HDAC3 formed memory that was unusually specific to the AM rate paired with reward. Unusually sound-specific memory revealed behaviorally was associated with a signal-specific enhancement in temporal coding in the auditory system: stronger phase-locking that was specific to the rewarded AM rate was revealed in both the surface-recorded frequency following response (FFR) and auditory cortical multiunit activity in rats treated with the HDAC3-inhibitor. Furthermore, HDAC3inhibition increased trial-to-trial cortical response consistency (relative to naive and trained vehicle-treated rats) that generalized across different AM rates. Stronger signal-specific phase-locking correlated with individual behavioral differences in memory specificity for the AM signal. Together, these findings support that epigenetic mechanisms regulate activity-dependent processes that enhance discriminability of sensory cues encoded into LTM in both spectral and temporal domains, which may be important for remembering spectrotemporal features of sounds, e.g., as in human voices and speech.

SIGNIFICANCE STATEMENTEpigenetic mechanisms have recently been implicated in memory and information processing. Here, we use a pharmacological inhibitor of histone deacetylase 3 (HDAC3) in a sensory model of learning to reveal, for the first time, its ability to enable unusually precise memory for amplitude modulated sound cues. In so doing, we uncover neural substrates for memorys "specificity" for temporal sound cues. Memory specificity was supported by auditory cortical changes in temporal coding, including greater response consistency and stronger phase-locking. HDAC3 appears to regulate effects across domains that determine specific cue saliency for behavior. Thus, epigenetic players may gate how sensory information is stored in long-term memory and can be leveraged to reveal the neural substrates of sensory details stored in memory.


title: Comparison of Type-1 and Type-2 Fuzzy Systems forMineralization of Bioprinted Bone

abstract: Bioprinting is an emerging tissue engineering method used to generate cell-laden scaffolds with high spatial resolution. Bioprinting parameters, such as pressure, nozzle size, and speed, have a large influence on the quality of the bioprinted construct. Moreover, cell suspension density, cell culture period, and other critical biological parameters directly impact the biological function of the final product. Therefore, an approximation model that can be used to find the values of bioprinting parameters that will result in optimal bioprinted constructs is highly desired. Here, we propose type-1 and type-2 fuzzy systems to handle the uncertainty and imprecision in optimizing the input values. Specifically, we focus on the biological parameters, such as culture period, that can be used to maximize the output value (mineralization volume). To achieve a more accurate approximation, we have compared a type-2 fuzzy system with a type-1 fuzzy system using two levels of uncertainty. We hypothesized that type-2 fuzzy systems may be preferred in biological systems, due to the inherent vagueness and imprecision of the input data. Here, our results demonstrate that the type-2 fuzzy system with a high uncertainty boundary (30%) is superior to type-1 and type-2 with low uncertainty boundary fuzzy systems in the overall output approximation error for bone bioprinting inputs.


title: Biomolecular solid-state NMR spectroscopy at highest field: the gain in resolution at 1200 MHz

abstract: Progress in NMR in general and in biomolecular applications in particular is driven by increasing magnetic-field strengths leading to improved resolution and sensitivity of the NMR spectra. Recently, persistent superconducting magnets at a magnetic field strength (magnetic induction) of 28.2 T corresponding to 1200 MHz proton resonance frequency became commercially available. We present here a collection of high-field NMR spectra of a variety of proteins, including molecular machines, membrane proteins and viral capsids and others. We show this large panel in order to provide an overview over a range of representative systems under study, rather than a single best performing model system. We discuss both carbon-13 and proton-detected experiments, and show that in 13C spectra substantially higher numbers of peaks can be resolved compared to 850 MHz while for 1H spectra the most impressive increase in resolution is observed for aliphatic side-chain resonances.

category: biophysics


title: Auxin-responsive (phospho)proteome analysis reveals regulation of cell cycle and ethylene signaling during rice crown root development

abstract:
The rice root system, which primarily consists of adventitious/crown roots (AR/CR) developed from the coleoptile base, is an excellent model system for studying shoot-to-root trans-differentiation process. We reveal global changes in protein and metabolite abundance, and protein phosphorylation in response to an auxin stimulus during CR development. Global proteome and metabolome analyses of developing crown root primordia (CRP) and emerged CRs uncovered that the biological processes associated with chromatin conformational change, gene expression, and cell cycle were translationally regulated by auxin signaling. Spatial gene expression pattern analysis of differentially abundant proteins disclosed their stage-specific dynamic expression pattern during CRP development. Further, our tempo-spatial gene expression and functional analyses revealed that auxin creates a regulatory feedback module during CRP development and activates ethylene biosynthesis exclusively during CRP initiation. Ethylene signaling promotes CR formation by repressing the cytokinin response regulator, OsRR2. Additionally, the (phospho)proteome analysis identified differential phosphorylation of the Cyclin-dependent kinase G-2 (OsCDKG;2), and cell wall proteins, in response to auxin signaling, suggesting that auxin-dependent phosphorylation may be required for cell cycle activation, and cell wall synthesis during root organogenesis. Thus, our study provides evidence for the translational and post-translational regulation during CRP trans-differentiation downstream of the auxin signaling pathway.

HighlightGlobal (phospho)proteome and metabolic profiling of rice CRP and CRs uncover differential proteins and metabolites associated with gene expression, cell cycle, ethylene signaling and cell wall synthesis during CR development.


title: Plant Metabolic Network: A multi-species resource of plant metabolic information

abstract:
Plant metabolism is a pillar of our ecosystem, food security, and economy. To understand and engineer plant metabolism, we first need a comprehensive and accurate annotation of all metabolic information across plant species. As a step towards this goal, we previously created the Plant Metabolic Network (PMN), an online resource of curated and computationally predicted information about the enzymes, compounds, reactions, and pathways that make up plant metabolism. Here we report PMN 15, which contains genome-scale metabolic pathway databases of 126 algal and plant genomes, ranging from model organisms to crops to medicinal plants, and new tools for analyzing and viewing metabolism information across species and integrating omics data in a metabolic context. We systematically evaluated the quality of the databases, which revealed that our semi-automated validation pipeline dramatically improves the quality. We then compared the metabolic content across the 126 organisms using multiple correspondence analysis and found that Brassicaceae, Poaceae, and Chlorophyta appeared as metabolically distinct groups. To demonstrate the utility of this resource, we used recently published sorghum transcriptomics data to discover previously unreported trends of metabolism underlying drought tolerance. We also used single-cell transcriptomics data from the Arabidopsis root to infer cell-type specific metabolic pathways. This work shows the continued growth and refinement of the PMN resource and demonstrates its wide-ranging utility in integrating metabolism with other areas of plant biology.

One-sentence SummaryThe Plant Metabolic Network is a collection of databases containing experimentally-supported and predicted information about plant metabolism spanning many species.


title: Acclimation of Chlamydomonas reinhardtii to Nitric Oxide Stress Related to Respiratory Burst Oxidase-Like 2

abstract:
The acclimation mechanism of Chlamydomonas reinhardtii to nitric oxide (NO) was studied by exposure to S-nitroso-N-acetylpenicillamine (SNAP), a NO donor. Treatment with 0.1 or 0.3 mM SNAP transiently inhibited photosynthesis within 1 h, followed by a recovery without growth impairment, while 1.0 mM SNAP treatment caused irreversible photosynthesis inhibition and mortality. The SNAP effects are avoided in the presence of the NO scavenger, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-l-oxyl-3-oxide (cPTIO). RNA-seq, qPCR, and biochemical analyses were conducted to decode the metabolic shifts under sub-lethal NO stress by exposure to 0.3 mM SNAP in the presence or absence of 0.4 mM cPTIO. These findings revealed that the acclimation to NO stress comprises a temporally orchestrated implementation of metabolic processes: 1. trigger of NO scavenging elements to reduce NO level; 2. prevention of photo-oxidative risk through photosynthesis inhibition and antioxidant defense system induction; 3. acclimation to nitrogen and sulfur shortage; 4. degradation of damaged proteins through protein trafficking machinery (ubiquitin, SNARE, and autophagy) and molecular chaperone system for dynamic regulation of protein homeostasis. NO increased NADPH oxidase activity and respiratory burst oxidase-like 2 (RBOL2) transcript abundance, which were not observed in the rbol2 insertion mutant. Changes in gene expression in the rbol2 mutant and increased mortality under NO stress demonstrate that NADPH oxidase (RBOL2) is involved in the modulation of some acclimation processes (NO scavenging, antioxidant defense system, autophagy, and heat shock proteins) for Chlamydomonas to cope with NO stress. Our findings provide insight into the molecular events underlying acclimation mechanisms in Chlamydomonas to sub-lethal NO stress.

One-sentence SummaryAcclimation machinery is triggered in Chlamydomonas reinhardtii cells against sub-lethal nitric oxide stress.


title: A morpho-transcriptomic map of brassinosteroid action in the Arabidopsis root

abstract: The effects of brassinosteroid signaling on shoot and root development have been characterized in great detail but did not identify a simple consistent positive or negative impact on a basic cellular parameter that would comprehensively explain the phenotype of brassinosteroid-related mutants. Here we combined digital 3D single-cell shape analysis and single-cell mRNA sequencing to characterize root meristems and mature root segments of brassinosteroid-blind mutants and wildtype. These data demonstrate that brassinosteroid signaling neither affects cell volume nor cell proliferation capacity. Instead, brassinosteroid signaling is essential for the precise orientation of cell division planes and the extent and timing of anisotropic cell expansion. Moreover, we found that the cell-aligning effects of brassinosteroid signaling can propagate to normalize the anatomy of both adjacent and distant brassinosteroid-blind cells through non-cell-autonomous functions, which are sufficient to restore overall root growth vigor. Finally, single-cell transcriptome data discern directly brassinosteroid-responsive genes from genes that can react to non-cell-autonomous brassinosteroid-dependent signals and highlight arabinogalactans as sentinels of brassinosteroid-dependent anisotropic cell expansion.


title: Production of Volatile Moth Sex Pheromones in Transgenic Nicotiana benthamiana Plants

abstract: Plant-based bio-production of insect sex pheromones has been proposed as an innovative strategy to increase the sustainability of pest control in agriculture. Here we describe the engineering of transgenic plants producing (Z)-11-hexadecen-1-ol (Z11-16OH) and (Z)-11-hexadecenyl acetate (Z11-16OAc), two main volatile components in many Lepidoptera sex pheromone blends. We assembled multigene DNA constructs encoding the pheromone biosynthetic pathway and stably transformed them in Nicotiana benthamiana plants. The constructs comprised the Amyelois transitella Atr{Delta}11 desaturase gene, the Helicoverpa armigera farnesyl reductase HarFAR gene, and the Euonymus alatus diacylglycerol acetyltransferase EaDAct gene in different configurations. All the pheromone-producing plants showed dwarf phenotypes, whose severity correlated with pheromone levels. All but one of the recovered lines produced high levels of Z11-16OH but very low levels of Z11-16OAc, probably as a result of recurrent truncations at the level of the EaDAct gene. Only one plant line (SxPv1.2) was recovered harbouring an intact pheromone pathway and producing moderate levels of Z11-16OAc (11.8 {micro}g g-1 FW), next to high levels of Z11-16OH (111.4 {micro}g g-1). Z11-16OAc production was accompanied in SxPv1.2 by a partial recovery of the dwarf phenotype. SxPv1.2 was used to estimate the rates of volatile pheromone release, which resulted in 8.48 ng g-1 FW per day for Z11-16OH and 9.44 ng g-1 FW per day for Z11-16OAc. Our results suggest that pheromone release acts as a limiting factor in pheromone bio-dispenser strategies and establish a roadmap for biotechnological improvements.

category: synthetic biology