Biology subjects

Mori, H.

Publications and source records attributed to Mori, H..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Enhanced tameness by Limosilactobacillus reuteri from gut microbiota of selectively bred mice

Domestication alters animal behaviour, primarily their tameness. In this study, we examine the effect of gut bacteria on mouse tameness. We previously conducted selective breeding for active tameness, defined as the motivation to approach a human hand, using genetically heterogeneous mice derived from eight wild inbred strains. We examined gut microbiota in the selectively bred mice by analysing faecal samples from 80 mice through shotgun metagenomic analysis. In the current study, we found that the selectively bred mice exhibit higher levels of active tameness as well as higher levels of blood oxytocin, which plays a key role in social behaviours. Selection for tameness did not substantially alter the taxonomic or functional diversity of the gut microbiota. However, we observed an increased abundance of Limosilactobacillus reuteri in the selected groups and higher pyruvate levels in their plasma. We isolated L. reuteri strains secreting extracellular pyruvate from mice faeces and administrated the cultured bacteria through drinking water. Mice treated with L. reuteri showed higher colonization of the bacteria in the gut, as well as higher levels of active tameness behaviour and blood oxytocin. Additionally, we generated 374 high-quality metagenome-assembled genomes (MAGs) of bacteria across 11 phyla. This collection includes 27 novel species level bacterial MAGs not previously known to exist in the mouse gut. This study elucidates the potential role of L. reuteri in the animal domestication process and explores the underlying mechanisms that may influence this process.

microbiology↗

bioRxiv2023

Age-associated aberrations of cumulus-oocyte interaction and microfilamentous structure in the zona pellucida decline female fertility

One of the major age-related declines in female reproductive function is the reduced quantity and quality of oocytes. Here we demonstrate that structural changes in the zona pellucida (ZP) decrease fertilization rates from 34- to 38-week-old female mice, equivalent to the mid-reproductive of human females. In middle-age ovaries, the decline in the number of transzonal projections was accompanied by a decrease in cumulus cell-oocyte interactions, resulting in a deterioration of the oocyte quality. Scanning electron microscopy showed the ZP surface microfilament structure transitioning from rugged to smooth with age, leading to decreased fertilization rates due to impaired sperm binding to the ZP. Moreover, the fertilization rate of middle-age mice was restored to a comparable level to that of young mice by destabilizing the ZP in the presence of glutathione. These results suggest that the age-related structural changes in the ZP is a key for successful fertilization at reproductive age.

developmental biology↗

bioRxiv2023

Tumor microenvironmental determinants of high-risk DCIS progression

ABSTRACT/SUMMARYDuctal carcinoma in situ (DCIS) constitutes an array of morphologically recognized intraductal neoplasms in the mammary ductal tree defined by an increased risk for subsequent invasive carcinomas at or near the site of biopsy detection. However, only 15-45% of untreated DCIS cases progress to invasive cancer, so understanding mechanisms that prevent progression is key to avoid overtreatment and provides a basis for alternative therapies and prevention. This study was designed to characterize the tumor microenvironment and molecular profile of high-risk DCIS that grew to a large size but remained as DCIS. All patients had DCIS lesions >5cm in size with at least one additional high-risk feature: young age (<45 years), high nuclear grade, hormone receptor negativity, HER2 positivity, the presence of comedonecrosis, or a palpable mass. The tumor immune microenvironment was characterized using multiplex immunofluorescence to identify immune cells and their spatial relationships within the ducts and stroma. Gene copy number analysis and whole exome DNA sequencing identified the mutational burden and driver mutations, and quantitative whole-transcriptome/gene expression analyses were performed. There was no association between the percent of the DCIS genome characterized by copy number variants (CNAs) and recurrence events (DCIS or invasive). Mutations, especially missense mutations, in the breast cancer driver genes PIK3CA and TP53 were common in this high-risk DCIS cohort (47% of evaluated lesions). Tumor infiltrating lymphocyte (TIL) density was higher in DCIS lesions with TP53 mutations (p=0.0079) compared to wildtype lesions, but not in lesions with PIK3CA mutations (p=0.44). Immune infiltrates were negatively associated with hormone receptor status and positively associated with HER2 expression. High levels of CD3+CD8-T cells were associated with good outcomes with respect to any subsequent recurrence (DCIS or invasive cancer), whereas high levels of CD3+Foxp3+ Treg cells were associated with poor outcomes. Spatial proximity analyses of immune cells and tumor cells demonstrated that close proximity of T cells with tumor cells was associated with good outcomes with respect to any recurrence as well as invasive recurrences. Interestingly, we found that myoepithelial continuity (distance between myoepithelial cells surrounding the involved ducts) was significantly lower in DCIS lesions compared to normal tissue (p=0.0002) or to atypical ductal hyperplasia (p=0.011). Gene set enrichment analysis identified several immune pathways associated with low myoepithelial continuity and a low myoepithelial continuity score was associated with better outcomes, suggesting that gaps in the myoepithelial layer may allow access/interactions between immune infiltrates and tumor cells. Our study demonstrates the immune microenvironment of DCIS, in particular the spatial proximity of tumor cells and T cells, and myoepithelial continuity are important determinants for progression of disease.

cancer biology↗

bioRxiv2023

Genetically stable kill-switch using ''demon and angel'' expression construct of essential genes

Genetic instability of synthetic genetic devices is a key obstacle for practical use. This problem is particularly critical in kill-switches for conditional host killing. Here, we propose a genetically stable kill-switch based on a "demon and angel" expression construct of a toxic essential gene. The kill-switch conditionally overexpresses the toxic essential gene. Additionally, the identical essential gene is deleted in the genome. The essential gene is expressed at a low level to maintain host survival in the OFF state and kills the host by the overexpression in the ON state. The single expression construct is responsible for both killing the hosts and maintaining viability, reducing the emergence of loss-of-function mutants. We constructed the kill-switch using the toxic essential gene encoding tyrosyl-tRNA synthetase, tyrS, in Escherichia coli. The bacteria harboring the kill-switch were conditionally suicidal over 300 generations. Toxic overexpression of essential genes has also been found in other organisms, suggesting that the "demon and angel" kill switch is scalable to various organisms. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/566782v1_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@8cb5d2org.highwire.dtl.DTLVardef@1f52190org.highwire.dtl.DTLVardef@6e9ef9org.highwire.dtl.DTLVardef@870853_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2023

SCD1 and monounsaturated lipids are required for autophagy and survival of adipocytes

Exposure of adipocytes to cool temperatures often found in the periphery of the body induces expression of Stearoyl-CoA Desaturase-1 (SCD1), an enzyme that converts saturated fatty acids to monounsaturated fatty acids. In this study, we employed Scd1 knockout cells and mouse models, along with pharmacological SCD1 inhibition, to investigate further the roles of SCD1 in adipocytes. Our study reveals that production of monounsaturated lipids by SCD1 is necessary for fusion of autophagosomes to lysosomes and that with a SCD1-deficiency, autophagosomes accumulate. In addition, SCD1-deficiency impairs lysosomal and autolysosomal acidification resulting in vacuole accumulation and eventual cell death. Blocking autophagosome formation or supplementation with monounsaturated fatty acids maintains vitality of SCD1-deficient adipocytes. Taken together, our results demonstrate that in vitro inhibition of SCD1 in adipocytes leads to autophagy-dependent cell death, and in vivo depletion leads to loss of bone marrow adipocytes.

molecular biology↗

bioRxiv2023

Tyk2-mediated signaling promotes the development of autoreactive CD8+ CTLs and autoimmune type 1 diabetes

Tyrosine kinase 2 (TYK2), a member of the JAK family, might be a susceptibility gene for type 1 diabetes (T1D), whereas its precise role in autoimmune T1D remains unknown. We showed Tyk2 deficiency and inhibition suppressed autoimmune T1D development in non-obese diabetic (NOD) mice. Defective IL-12 signaling due to Tyk2 deficiency in islet-autoreactive CD8+ CTLs during their priming reduced T-bet expression, leading to impaired Cxcr3 expression and effector functions against {beta}-cells. Tyk2 deficient CD8+ resident dendritic cells (rDC) exhibited reduced MHC I expression and impaired cross-priming of CTLs. In {beta}-cells, increased expressions of Fas, MHC I, and chemokines with age were attenuated by Tyk2 deficiency. We demonstrated that treatment with BMS-986165, a Tyk2 inhibitor, inhibited the development of CTLs and inflammation in {beta}-cells in vitro. BMS-986165 reduced the incidence of diabetes in NOD mice. Thus, we demonstrated that Tyk2-mediated signaling has a critical role in the development of autoreactive CD8+ CTLs, inflammation in {beta}-cells, and the pathogenesis of autoimmune T1D. SummaryWe demonstrated that Tyk2-mediated signaling plays a critical role in the development of autoreactive CD8+ CTLs, inflammation in {beta}-cells, and the pathogenesis of autoimmune T1D. These findings will lead to the development of safety and effective prevention strategies for T1D.

immunology↗

bioRxiv2023

Recognition of commensal bacterial peptidoglycans defines Drosophila gut homeostasis and lifespan

Commensal microbes in animals have a profound impact on tissue homeostasis, stress resistance, and ageing. We previously showed in Drosophila melanogaster that Acetobacter persici is a member of the gut microbiota that promotes ageing and shortens fly lifespan. However, the molecular mechanism by which this specific bacterial species changes lifespan and physiology remains unclear. The difficulty in studying longevity using gnotobiotic flies is the high risk of contamination during ageing. To overcome this technical challenge, we used a bacteria-conditioned diet enriched with bacterial products and cell wall components. Here, we demonstrate that an A. persici-conditioned diet shortens lifespan and increases intestinal stem cell (ISC) proliferation. Feeding adult flies a diet conditioned with A. persici, but not with Lactiplantibacillus plantarum, can decrease lifespan but increase resistance to paraquat or oral infection of Pseudomonas entomophila, indicating that the bacterium alters the trade-off between lifespan and host defence. A transcriptomic analysis using fly intestine revealed that A. persici preferably induces antimicrobial peptides (AMPs), while L. plantarum upregulates amidase peptidoglycan recognition proteins (PGRPs). The specific induction of these Imd target genes by peptidoglycans from two bacterial species is due to the stimulation of the receptor PGRP-LC in the anterior midgut for AMPs or PGRP-LE from the posterior midgut for amidase PGRPs. Heat-killed A. persici also shortens lifespan and increases ISC proliferation via PGRP-LC, but it is not sufficient to alter the stress resistance. Our study emphasizes the significance of peptidoglycan specificity in determining the gut bacterial impact on healthspan. It also unveils the postbiotic effect of specific gut bacterial species, which turns flies into a "live fast, die young" lifestyle. Author SummaryMicrobiota plays a vital role in our health, but it can also have a negative impact on the lifespan of certain model organisms, such as the fruit fly Drosophila melanogaster. Despite its impact, the molecular mechanism behind how gut bacteria limits host lifespan remains unclear. In this study, we investigated the mechanism that one specific species of microbiota shortens lifespan and disrupts gut homeostasis of the aged flies, using a "fermented" fly diet. We found that the specific effects of this bacterium on fly healthspan were due to its capability of stimulating a specific receptor in the gut that recognizes peptidoglycan, a component of bacterial cell wall. Paradoxically, the same bacterium also increases stress resistance and defence against oral infection of a pathogen. Our study provides insight into the mechanisms underlying how certain members of the microbiota can lead to a "life fast, die young" lifestyle.

immunology↗

bioRxiv2023

Identification and characterization of a proliferative cell population in estrogen receptor-positive metastatic breast cancer through spatial and single-cell transcriptomics

BackgroundIntratumor heterogeneity is a hallmark of most solid tumors, including breast cancers. We applied spatial transcriptomics and single-cell RNA-sequencing technologies to profile spatially resolved cell populations within estrogen receptor-positive (ER+) metastatic breast cancers and elucidate their importance in estrogen-dependent tumor growth. MethodsSpatial transcriptomics and single-cell RNA-sequencing were performed on two patient-derived xenografts (PDXs) of "ER-high" metastatic breast cancers with opposite estrogen-mediated growth responses: estrogen-suppressed GS3 (80-100% ER) and estrogen-stimulated SC31 (30-75% ER) models. The analyses included samples treated with and without 17{beta}-estradiol. The findings were validated via scRNA-seq analyses on "ER-low" estrogen-accelerating PDX, GS1 (5% ER). The results from our spatial and single-cell analyses were further supported by the analysis of a publicly available single cell dataset and a protein-based dual immunohistochemical (IHC) evaluation using three important clinical markers [i.e., ER, progesterone receptor (PR), and Ki67]. The translational implication of these results was assessed by clinical outcome analyses on public breast cancer cohorts. ResultsOur novel space-gene-function study revealed a "proliferative" cell population in addition to three major spatially distinct compartments within ER+ metastatic breast cancers. These compartments showed functional diversity (i.e., estrogen-responsive, proliferative, hypoxia-induced, and inflammation-related). The "proliferative (MKI67+)" population, not "estrogen-responsive" compartment, was crucial for estrogen-dependent tumor growth, leading to the acquisition of luminal B features. The cells with induction of typical estrogen-responsive genes such as PGR were not directly linked to estrogen-dependent proliferation. Additionally, the dual IHC analyses demonstrated the distinct contribution of the Ki67+ proliferative cells toward estrogen-mediated growth and their response to palbociclib, a CDK4/6 inhibitor. The gene signatures developed from the proliferative, hypoxia-induced, and inflammation-related compartments were significantly correlated with worse clinical outcomes, while patients with the high estrogen-responsive scores showed better prognosis, confirming that the estrogen-responsive compartment would not be directly associated with estrogen-dependent tumor progression. ConclusionsFor the first time, our study elucidated a "proliferative" cell population distinctly distributed in ER+ metastatic breast cancers. They contribute differently toward progression of these cancers, and the gene signature in the "proliferative" compartment is an important determinant of luminal cancer subtypes.

cancer biology↗

bioRxiv2023

A multi-kingdom genetic barcoding system for precise target clone isolation

Clonal heterogeneity underlies diverse biological processes, including cancer progression, cell differentiation, and microbial evolution. Cell tagging strategies with DNA barcodes have recently enabled analysis of clone size dynamics and clone-restricted transcriptomic landscapes of heterogeneous populations. However, isolating a target clone that displays a specific phenotype from a complex population remains challenging. Here, we present a new multi-kingdom genetic barcoding system, CloneSelect, in which a target cell clone can be triggered to express a reporter gene for isolation through barcode-specific CRISPR base editing. In CloneSelect, cells are first barcoded and propagated so their subpopulation can be subjected to a given experiment. A clone that shows a phenotype or genotype of interest at a given time can then be isolated from the initial or subsequent cell pools stored throughout the experimental timecourse. This novel CRISPR-barcode genetics platform provides many new ways of analyzing and manipulating mammalian, yeast, and bacterial systems. TeaserA multi-kingdom CRISPR-activatable barcoding system enables the precise isolation of target barcode-labeled clones from a complex cell population.

synthetic biology↗

bioRxiv2022

Efficient Gene Transduction in Pigs and Macaques with the Engineered AAV Vector AAV.GT5 for Hemophilia B Gene Therapy

Gene therapy for hemophilia using adeno-associated virus (AAV) vectors allows long-term coagulation factor expression. We examined the potential of a novel engineered liver-tropic AAV3B-based vector AAV.GT5 for hemophilia B gene therapy. In vitro transduction with AAV.GT5 in human hepatocytes was more than 100 times higher than with AAV-Spark100, while in vivo transduction efficacy into the liver and the increase in coagulation factor IX (FIX) antigen following intravenous injection of these vectors were similar in PXB mice (chimeric mice with a humanized liver) and macaques. The discrepancy was due to the low recovery and short half-life of AAV.GT5 in blood, depending on the positive charge of the heparin-binding site in the original AAV3B. The intra-hepatic vascular administration of AAV.GT5, but not AAV-Spark100, enhanced vector transduction into the liver and reduced vector distribution to the kidney in pigs. In macaques, the intra-hepatic artery injection of AAV.GT5 yielded a comparable increase in FIX antigen with a one-third dosage of peripheral venous administration. Two of four macaques who received AAV.GT5 intravenously did not develop neutralizing antibodies (NAbs) against AAV.GT5, while AAV-Spark100 induced serotype-specific NAbs in all four macaques. The NAb produced after the administration was relatively specific to the serotype and less responsive to the other serotype. As a result, the administration of AAV.GT5 successfully boosted FIX expression in one animal previously given AAV-Spark100. Thus, AAV.GT5 has different biodistribution and immunogenic characteristics compared with AAV-Spark100, and the intra-hepatic vascular administration may lessen the vector dose and avoid vector distribution to other organs. Key PointsO_LIThe AAV.GT5 vector has a strong transduction efficacy in human hepatocytes but has a faster clearance after systemic administration. C_LIO_LIIntra-hepatic vascular administration of the AAV.GT5 vector is an effective liver transduction method for hemophilia gene therapy. C_LI

genetics↗

bioRxiv2022

Analysis of IGHA1 and other salivary proteins post half marathon in female participants

BackgroundHigh intensity exercise, such as in marathons and triathlons, suppresses transient local and systemic immunity. Much is known about the systemic immunosuppression response, but little is known about its local response in the oral cavity, lungs, bronchial tubes, and skin. The oral cavity is an entrance for bacteria or viruses to enter the body. Saliva covers the epidermis of the oral cavity and plays an important role in the local stress response by preventing infection. In this study, we examined the properties of saliva secreted during the local stress response induced by a half marathon (HM) using quantitative proteomic analysis. MethodsThe Exercise group (ExG), 19 healthy female university students participated in (HM) race. The non-exercise group (NExG), 16 healthy female university students had not participated in the ExG. In ExG, saliva samples were collected 1h pre and 2h and 4h post HM. NExG were collected in same time interval. Saliva volume, protein concentration and relative IGHA1 expression were analyzed. In addition, 1h pre and 2h post HM saliva were analyzed by iTRAQ. The identified factors of iTRAQ, analyzed for the ExG and NExG by western blotting. ResultsWe identified kallikrein 1 (KLK1), immunoglobulin kappa chain (IgK) and cystatin S (CST4) as suppression factors, as well as immunoglobulin heavy constant alpha 1 (IGHA1), which has been reported as an immunological stress marker. KLK1 (p = 0.011), IGK (p = 0.002), and CST4 (p = 0.003) were suppressed 2 h post-HM compared to pre-HM, and KLK1 (p = 0.004) and CST4 (p = 0.006) were suppressed 4 h post-HM. There was also a positive correlation between IGHA1, IGK, and CST4 2 and 4 h post-HM. In addition, KLK1 and IGK after 2 h post-HM showed a positive correlation. ConclusionOur study demonstrated that the salivary proteome is regulated and antimicrobial proteins are suppressed post-HM. These results suggest that oral immunity was transiently suppressed post-HM. The positive correlation of each protein at 2 and 4 h post-HM suggests that the suppressed state was similarly regulated up to 4 h after a HM. The proteins identified in this study may have applications as stress markers for recreational runners and individuals who perform high- and moderate-intensity exercises in their daily lives.

physiology↗

bioRxiv2022

Multimodal brain age estimates relate to Alzheimer disease biomarkers and cognition in early stages: a cross-sectional observational study

BackgroundEstimates of "brain-predicted age" quantify apparent brain age compared to normative trajectories of neuroimaging features. The brain age gap (BAG) between predicted and chronological age is elevated in symptomatic Alzheimer disease (AD), but has not been well explored in preclinical AD. Prior studies have typically modeled BAG with structural magnetic resonance imaging (MRI), but more recently other modalities, including functional connectivity (FC) and multimodal MRI, have been explored. MethodsWe trained three models to predict age from FC, volumetric (Vol), or multimodal MRI (Vol+FC) in 390 control participants (18-89 years old). In independent samples of 144 older adult controls, 154 preclinical AD participants, and 154 cognitively impaired (CI; CDR > 0) participants, we tested relationships between BAG and AD biomarkers of amyloid, tau, and neurodegeneration, as well as a global cognitive composite. ResultsAll models predicted age in the control training set, with the multimodal model outperforming the unimodal models. All three BAG estimates were significantly elevated in CI compared to controls. FC-BAG and Vol+FC-BAG were marginally reduced in preclinical AD participants compared to controls. In CI participants only, elevated Vol-BAG and Vol+FC-BAG were associated with more advanced AD pathology and lower cognitive performance. ConclusionsBoth FC-BAG and Vol-BAG are elevated in CI participants. However, FC and volumetric MRI also capture complementary signals. Specifically, FC-BAG may capture a unique biphasic response to preclinical AD pathology, while Vol-BAG may capture pathological progression and cognitive decline in the symptomatic stage. A multimodal age-prediction model captures these modality-specific patterns, and further, improves sensitivity to healthy age differences. FundingThis work was supported by the National Institutes of Health (P01-AG026276, P01-AG03991, P30-AG066444, 5-R01-AG052550, 5-R01-AG057680, 1-R01-AG067505, 1S10RR022984-01A1, U19-AG032438), the BrightFocus Foundation (A2022014F), and the Alzheimers Association (SG-20-690363-DIAN).

neuroscience↗

bioRxiv2022

Metagenomic thermometer

Various microorganisms exist in environments, and each of which has an optimal growth temperature (OGT). The relationship between genomic information and OGT of each species has long been studied, and one such study revealed that OGT of prokaryotes can be accurately predicted based on the fraction of seven amino acids (IVYWREL) among all encoded amino-acid sequences in its genome. Extending this discovery, the fraction of IVYWREL among all amino acids encoded in the metagenomic DNA in an environment may reflect the environmental temperature to which the microorganisms have adapted. In the present study, we developed a Metagenomic Thermometer as a means of predicting environmental temperature based on metagenomic sequences. Analysis of 41 metagenomic samples obtained from hot springs of varying temperature revealed that the Metagenomic Thermometer could indeed predict environmental temperature with high accuracy (R2 = 0.75). Although the taxonomic composition of microbial community was substantially affected by pH, the performance of the Metagenomic Thermometer was refractory to changes in community composition. The Metagenomic Thermometer was also applied to 616 human gut metagenomic samples, yielding a reasonably accurate value (i.e., 36.22 {+/-} 1.98{degrees}C) for human body temperature. Our results suggest that environmental temperature can be predicted based on metagenomic sequence data.

bioinformatics↗

bioRxiv2022

CAPG is required for Ebola virus infection by controlling virus egress from infected cells.

Replication of Ebola virus (EBOV) is dependent upon actin functionality, especially at cell entry through macropinocytosis and at release of virus from cells. Previously, major actin-regulatory factors such as Rac1 and ARP2/3, involved in actin nucleation were shown important in both steps. However, downstream of nucleation, many other cell factors, are needed to control actin dynamics. How these regulate EBOV infection remains largely unknown. Here, we identified the actin-regulating protein, CAPG, as important for EBOV replication. Notably, knockdown (KD) of CAPG specifically inhibited viral infectivity and yield of infectious particles. Mechanistic analysis revealed a requirement of CAPG for virus production from infected cells. Proximity ligation and split GFP reconstitution assays revealed strong association of CAPG with VP40 that was mediated through the S1 domain of CAPG. Overall, CAPG is a novel host factor regulating EBOV infection through connecting actin filament stabilization to viral egress from cells.

microbiology↗

bioRxiv2022

Neuroimaging within the Dominantly Inherited Alzheimer's Network (DIAN): PET and MRI

The Dominantly Inherited Alzheimer Network (DIAN) Observational Study is an international collaboration studying autosomal dominant Alzheimer disease (ADAD). This rare form of Alzheimer disease (AD) is caused by mutations in the presenilin 1 (PSEN1), presenilin 2 (PSEN2), or amyloid precursor protein (APP) genes. As individuals from these families have a 50% chance of inheriting the familial mutation, this provides researchers with a well-matched cohort of carriers vs non-carriers for case-control studies. An important trait of ADAD is that the age at symptom onset is highly predictable and consistent for each specific mutation, allowing researchers to estimate an individuals point in their disease time course prior to symptom onset. Although ADAD represents only a small proportion (approximately 0.1%) of all AD cases, studying this form of AD allows researchers to investigate preclinical AD and the progression of changes that occur within the brain prior to AD symptom onset. Furthermore, the young age at symptom onset (typically 30-60 years) means age-related comorbidities are much less prevalent than in sporadic AD, thereby allowing AD pathophysiology to be studied independent of these confounds. A major goal of the DIAN Observational Study is to create a global resource for AD researchers. To that end, the current manuscript provides an overview of the DIAN magnetic resonance imaging (MRI) and positron emission tomography (PET) protocols and highlights the key imaging results of this study to date.

neuroscience↗

bioRxiv2022

Lipolysis of bone marrow adipocytes is required to fuel bone and the marrow niche during energy deficits

To investigate roles for bone marrow adipocyte (BMAd) lipolysis in bone homeostasis, we created a BMAd-specific Cre mouse model in which we knocked out adipose triglyceride lipase (ATGL, Pnpla2). BMAd-Pnpla2-/- mice have impaired BMAd lipolysis, and increased size and number of BMAds at baseline. Although energy from BMAd lipid stores is largely dispensable when mice are fed ad libitum, BMAd lipolysis is necessary to maintain myelopoiesis and bone mass under caloric restriction. BMAd-specific Pnpla2 deficiency compounds the effects of caloric restriction on loss of trabecular bone, likely due to impaired osteoblast expression of collagen genes and reduced osteoid synthesis. RNA sequencing analysis of bone marrow adipose tissue reveals that caloric restriction induces dramatic elevations in extracellular matrix organization and skeletal development genes, and energy from BMAd is required for these adaptations. BMAd-derived energy supply is also required for bone regeneration upon injury, and maintenance of bone mass with cold exposure.

cell biology↗

bioRxiv2021

AP2/ERF transcription factor NbERF-IX-33 is involved in the regulation of phytoalexin production for the resistance of Nicotiana benthamiana to Phytophthora infestans.

Plants recognize molecular patterns unique to a certain group of microbes to induce effective resistance mechanisms. Elicitins are secretory proteins produced by plant pathogenic oomycete genera including Phytophthora and Pythium. Treatment of INF1 (an elicitin produced by P. infestans) induces a series of defense responses in Nicotiana species, including reactive oxygen species (ROS) production, transient induction of ethylene production, hypersensitive cell death and accumulation of the sesquiterpenoid phytoalexin capsidiol. In this study, we analyzed the expression profiles of N. benthamiana genes after INF1 treatment by RNAseq analysis. Based on their expression patterns, N. benthamiana genes were categorized into 20 clusters and 4,761 (8.3%) out of 57,140 genes were assigned to the clusters for INF1-induced genes. All genes encoding enzymes dedicated to capsidiol production, 5-epi-aristolochene (EA) synthase (NbEAS, 10 copies) and EA dehydrogenase (NbEAH, 6 copies), and some genes for ethylene production, such as 1-aminocyclopropane 1-carboxylate (ACC) synthase (NbACS) and ACC oxidase (NbACO), were significantly upregulated by INF1 treatment. Analysis of NbEAS1 and NbEAS4 promoters revealed that AGACGCC (GCC box-like motif) is the essential cis-element required for INF1-induced expression of NbEAS genes. Given that the GCC box is known to be targeted by ERF (ethylene-responsive factor) transcription factors, we created a complete list of N. benthamiana genes encoding AP2/ERF family transcription factors, and identified 45 out of 337 AP2/ERF genes in the clusters for INF1-induced genes. Among INF1-induced NbERF genes, silencing of NbERF-IX-33 compromised resistance against P. infestans and INF1-induced production of capsidiol. Recombinant NbERF-IX-33 protein can bind to the promoter sequence of NbEAS4, suggesting that NbERF-IX-33 is a transcription factor directly regulating the expression of genes for phytoalexin production.

plant biology↗

bioRxiv2021

The RNA helicase DDX6 controls early mouse embryogenesis by repressing aberrant inhibition of BMP signaling through miRNA-mediated gene silencing

The evolutionarily conserved RNA helicase DDX6 is a central player of post-transcriptional regulation, but its role during embryogenesis remains elusive. We here demonstrated that DDX6 enables proper cell lineage specification from pluripotent cells by analyzing Ddx6 KO mouse embryos and in vitro epiblast-like cell (EpiLC) induction system. Our study unveiled a great impact of DDX6-mediated RNA regulation on signaling pathways. Deletion of Ddx6 caused the aberrant transcriptional upregulation of the negative regulators of BMP signaling, which accompanied with enhanced Nodal signaling. Ddx6{bigtriangleup}/{bigtriangleup} pluripotent cells acquired higher pluripotency with a strong inclination toward neural lineage commitment. During gastrulation, abnormally expanded Nodal expression in the primitive streak likely promoted endoderm cell fate specification while inhibiting mesoderm development. We further clarified the mechanism how DDX6 regulates cell fate determination of pluripotent cells by genetically dissecting major DDX6 pathways: processing body (P-body) formation, translational repression, mRNA decay, and miRNA-mediated silencing. P-body-related functions were dispensable, but the miRNA pathway was essential for the DDX6 function. DDX6 may prevent aberrant transcriptional upregulation of the negative regulators of BMP signaling by repressing translation of certain transcription factors through the interaction with miRNA-induced silencing complexes (miRISCs). Overall, this delineates how DDX6 affects development of the three primary germ layers during early mouse embryogenesis and the underlying mechanism of DDX6 function. Author summaryGene expression occurs through the two steps: transcription (DNA to RNA) and translation (RNA to protein). Cells have very sophisticated regulatory processes working on various levels for the accurate gene expression. Post-transcriptional regulation, which includes all RNA-related controls, is crucial because it enables fine-tuning and rapid alteration of gene expression. RNA- binding proteins and non-coding RNAs are the two main players of post-transcriptional regulation. DDX6, the subject of our study, is an RNA-binding protein, more specifically an RNA helicase, which can unwind or rearrange RNA secondary structures. Its diverse molecular and cellular functions have been reported, but its embryogenic role is unknown. Here, we describe DDX6 function during early mouse embryogenesis and the underlying mechanism using genetic methodology. DDX6 enables proper cell lineage specification of pluripotent stem cells by mainly regulating BMP signaling through miRNA-mediated gene silencing. As DDX6- mediated RNA regulation affected signaling pathways, the loss of Ddx6 had a wide impact on developmental processes from pluripotency to embryo patterning. In addition, we identified which DDX6 molecular function is essential during early embryogenesis by genetically dissecting its main pathways.

genetics↗