Biology subjects

Suzuki, M.

Publications and source records attributed to Suzuki, M..

At least 91 records · Page 5Linked to original sources

bioRxiv2023

Early-life stress and ovarian hormones alter transcriptional regulation in the nucleus accumbens resulting in sex-specific responses to cocaine

Early-life stress and ovarian hormones contribute to increased female vulnerability to cocaine addiction. Here we reveal molecular substrates in the key reward area, the nucleus accumbens, through which these female-specific factors affect immediate and conditioning responses to cocaine in mice. We find shared involvement of X chromosome and estrogen signaling gene regulation in enhanced conditioning responses seen after early-life stress and during the low-estrogenic state in females. During the low-estrogenic state, females respond to acute cocaine exposure by increasing the accessibility of neuronal chromatin enriched for the binding sites of {Delta}FosB, a transcription factor implicated in chronic cocaine response and addiction. Conversely, high-estrogenic females respond to cocaine by preferential closing of neuronal chromatin, providing a mechanism for limiting cocaine-driven chromatin and synaptic plasticity. We find that physiological estrogen withdrawal, exposure to early-life stress, and absence of the second X chromosome all nullify the protective effect of high-estrogenic state on cocaine conditioning in females. Our findings offer a molecular framework to understand sex-specific neuronal mechanisms underlying cocaine use disorder.

neuroscience↗

bioRxiv2023

Assessing the performance of the Cell Painting assay across different imaging systems

Quantitative microscopy is a powerful method for performing phenotypic screens from which image-based profiling can extract a wealth of information, termed profiles. These profiles can be used to elucidate the changes in cellular phenotypes across cell populations from different patient samples or following genetic or chemical perturbations. One such image-based profiling method is the Cell Painting assay, which provides morphological insight through the imaging of eight cellular compartments. Here, we examine the performance of the Cell Painting assay across multiple high-throughput microscope systems and find that all are compatible with this assay. Furthermore, we determine independently for each microscope system the best performing settings, providing those who wish to adopt this assay an ideal starting point for their own assays. We also explore the impact of microscopy setting changes in the Cell Painting assay and find that few dramatically reduce the quality of a Cell Painting profile, regardless of the microscope used.

cell biology↗

bioRxiv2022

Significant SAR11 removal by a hard-bottom community

Microorganisms are a key component of the marine food webs through the microbial loop. In previous work, we have shown that some bacteria, including Candidatus Pelagibacter spp (SAR11)--the most abundant bacterium in the ocean--can evade filtration by benthic and planktonic tunicates. Here we tested whether differential removal of microbial taxa by benthic filter-feeders can be observed in the distribution and abundance of microbial taxa from hard-bottom subtidal communities, a common coastal habitat in the Eastern Mediterranean Sea towards the open sea. The abundance of microbial groups along crosss-hore transects was measured based on combined flow cytometry and SSU rRNA gene metabarcoding. Our results show that most groups were depleted (up to 50%) over the hard-bottom compared to the open sea, but unexpectedly we did not observe a clear differential removal of different taxa, SAR 11 notably. This study indicates a strong top-down control of the abundance of pelagic microorganisms over shallow hard-bottom where suspension feeders are common.

microbiology↗

bioRxiv2022

Rapid and collective determination of the complete "hot-spring frog" mitochondrial genome containing long repeat regions using Nanopore sequencing

The mitochondrial genome (mt-genome) is one of the promising molecular markers for phylogenetics and population genetics. Recently, various mt-genomes have been determined rapidly by using massively parallel sequencers. However, the control region (CR, also called D-loop) in mt-genomes remain difficult to precisely determine due to the presence of repeat regions. Here, using Nanopore sequencing, we succeeded in rapid and collective determination of complete mt-genome of the hot-spring frog, Buergeria japonica, and found that its mt-genome size was 22,274 bp including CR (6,929 bp) with two types of tandem repeat motifs forming repeat regions. Comparison of assemble strategies revealed that the long- and short-read data combined together enabled efficient determination of the CR, but the short-read data alone did not. The B. japonica CR was longer than that of a congenic species inhabiting cooler climate areas, Buergeria buergeri, because of the long repeat regions in the former. During the thermal adaptation of B. japonica, the longer repeat regions in its CR may have accumulated within a short period after divergence from B. buergeri.

genomics↗

bioRxiv2022

An optimized approach for multiplexing single-nuclear ATAC-seq using oligonucleotide conjugated antibodies

BackgroundSingle-cell technologies to analyze transcription and chromatin structure have been widely used in many research areas to reveal the functions and molecular properties of cells at single-cell resolution. Sample multiplexing techniques are valuable when performing single-cell analysis, reducing technical variation and permitting cost efficiencies. Several commercially available methods are available and have been used in many scRNA-seq studies. On the other hand, while several methods have been published, the multiplexing techniques for single nuclear Assay for Transposase-Accessible Chromatin (snATAC)-seq assays remain under development. We developed a simple nucleus hashing method using oligonucleotide conjugated antibodies recognizing nuclear pore complex proteins, NuHash, to perform snATAC-seq library preparations by multiplexing. ResultsWe performed multiplexing snATAC-seq analyses on the mixture of human and mouse cell samples (two samples, 2-plex, and four samples, 4-plex) using NuHash. The demultiplexing accuracy of NuHash was high, and only ten out of 9,144 nuclei (2-plex) and 150 of 12,208 nuclei (4-plex) had discordant classifications between NuHash demultiplexing and discrimination using reference genome alignments. We compared results between snATAC-seq and deeply sequenced bulk ATAC-seq on the same samples and found that most of the peaks detected in snATAC-seq were also detected in deeply sequenced bulk ATAC-seq. The bulk ATAC-seq signal intensity was positively correlated with the number of cell subtype clusters detected in snATAC-seq, but not the subset of peaks detected in all clusters. These subsets of snATAC-seq peaks showed different distributions over different genomic features, suggesting that the peak intensities of bulk ATAC-seq can be used to identify different types of functional loci. ConclusionsOur multiplexing method using oligo-conjugated anti-nuclear pore complex proteins, NuHash, permits high accuracy demultiplexing of samples. The NuHash protocol is straightforward, it works on frozen samples, and requires no modifications for snATAC-seq library preparation.

genomics↗

bioRxiv2022

Gross anatomy of the Pacific hagfish, Eptatretus burgeri, with special reference to the coelomic viscera

Hagfish (Myxinoidea) are a deep-sea taxon of cyclostomes, the extant jawless vertebrates. Many researchers have examined the anatomy and embryology of hagfish to shed light on the early evolution of vertebrates; however, the diversity within hagfish is often overlooked. Hagfish have two lineages, Myxininae and Eptatretinae. Usually, textbook illustrations of hagfish anatomy reflect the morphology of the former lineage, especially Myxine glutinosa, with its single pair of external branchial pores. Here, we instead report the gross anatomy of an Eptatretinae, Eptatretus burgeri, which has six pairs of branchial pores, especially focusing on the coelomic organs. Dissections were performed on fixed and unfixed specimens to provide a guide for those doing organ- or tissue-specific molecular experiments. Our dissections revealed that the ventral aorta is Y-branched in E. burgeri, which differs from the unbranched morphology of Myxine. Otherwise, there were no differences in the morphology of the lingual apparatus or heart in the pharyngeal domain. The thyroid follicles were scattered around the ventral aorta, as has been reported for adult lampreys. The hepatobiliary system more closely resembled those of jawed vertebrates than those of adult lampreys, with the liver having two lobes and a bile duct connecting the gallbladder to each lobe. Overall, the visceral morphology of E. burgeri does not differ significantly from that of the known Myxine at the level of gross anatomy, except for the number of branchial pores.

zoology↗

bioRxiv2022

Sequencing and chromosome-scale assembly of the giant Pleurodeles waltl genome

The Iberian ribbed newt (Pleurodeles waltl) constitutes a central model for probing the basis of vertebrate regeneration. Here, we present the sequencing and chromosome-scale assembly of the 20.3Gb P. waltl genome, which exhibits the highest level of contiguity and completeness among giant genome assemblies. We uncover that DNA transposable elements are the major contributors to its expansion, with hAT transposons comprising a large portion of repeats. Several hATs are actively transcribed and differentially expressed during adult P. waltl limb regeneration, along with domesticated hAT transposons of the ZBED transcription factor family. Despite its size, syntenic relationships are conserved. As an example, we show the high degree of conservation of the regeneration-associated Tig1 locus with several neighbouring genes. Together, the P. waltl genome provides a fundamental resource for the study of regenerative, developmental and evolutionary principles.

developmental biology↗

bioRxiv2022

Loss of the MLL3 tumor suppressor accelerates breast tumor onset via HIF1a-induced CCL2-mediated recruitment of CCR2+ regulatory T cells

While essential gatekeepers of immune homeostasis, Foxp3+ regulatory T (Treg) cells infiltrating tumors acquire distinct phenotypes and become highly immunosuppressive, promoting tumor immune escape and growth. How this occurs and relates to tumor-driver mutations is largely uncharacterized. Herein, we created a mouse mammary stem cell-based tumor model using CRISPR gene editing in which we introduced known human cancer-driver mutations. These included functional loss of the MLL3 histone methyltransferase and p53, and constitutive PI3-kinase activation, recapitulating the genetic makeup of aggressive breast cancers. We show that MLL3 loss fosters tumorigenesis by promoting the rapid establishment of an immunosuppressive microenvironment through induction of HIF1, which increases the secretion of the chemokine CCL2 by tumor cells and the recruitment of higher numbers of Foxp3+ Treg cells via CCR2. Greater infiltration of Treg cells also correlates with MLL3 downregulation and mutations in human breast cancer biopsies. Interestingly, HIF1 enforces the differentiation of tumor-infiltrating Treg cells into highly immunosuppressive ICOShiGITRhi Blimp-1hi effector Treg cells that enable rapid tumor escape. Monoclonal antibody targeting of ICOS or GITR inhibits tumorigenesis in most mice even two months after the cessation of treatment as well as the growth of established tumors, suggesting possible therapeutic opportunities for MLL3-mutant breast cancers.

immunology↗

bioRxiv2022

A Drosophila model of diabetic neuropathy reveals a crucial role of proteasome activity in the glia

Diabetic peripheral neuropathy (DPN) is the most common chronic, progressive complication of diabetes mellitus. The main symptom is sensory loss; the molecular mechanisms are not fully understood. We found that Drosophila fed a high-sugar diet, which induces diabetes-like phenotypes, exhibit impairment of noxious heat avoidance. The impairment of heat avoidance was associated with shrinkage of the leg neurons expressing the Drosophila transient receptor potential channel Painless. Using a candidate genetic screening approach, we identified proteasome modulator 9 as one of the modulators of impairment of heat avoidance. We further showed that proteasome inhibition in the glia reversed the impairment of noxious heat avoidance, and heat-shock proteins and exosome secretion in the glia mediated the effect of proteasome inhibition. Our results establish Drosophila as a useful system for exploring molecular mechanisms of diet-induced peripheral neuropathy and propose that the glial proteasome is one of the candidate therapeutic targets for DPN.

neuroscience↗

bioRxiv2022

Strand-biased circularizing integrative elements spread tmexCD-toprJ gene clusters encoding RND-type multidrug efflux pumps by repeated transpositions

Antimicrobial resistance genes (ARGs) are associated with mobile genetic elements (MGEs) that conscript useful genes into the human-microbe and microbe-microbe battlefields. Thus, under intense selective pressure, ARGs have been constantly adapting and evolving, spreading among microbes. tmexCD-toprJ gene clusters, which encode resistance-nodulation-cell division (RND)-type efflux pumps, confer multidrug-resistance to clinically important antimicrobials, including tigecycline. Noteworthily, these gene clusters have emerged in gram-negative bacteria in humans, animals, and the environment worldwide by MGE-mediated transfer. Here we show a hidden MGE, strand-biased circularizing integrative element (SE), that is recently recognized to mediate transpositions of ARGs, associated with the spread of tmexCD-toprJ gene clusters. We identified multidrug-resistant isolates of Aeromonas species in a water environment in Vietnam that harbored multiple copies of tmexCD-toprJ in their chromosomes that were associated with SEs. In particular, Aeromonas hydrophila NUITM-VA1 was found to harbor two copies of a novel variant of tmexC3.3D3.3-topJ1 within cognate SEs, whereas Aeromonas caviae NUITM-VA2 harbored four copies of a novel variant of tmexC2D2.3-topJ2 within cognate SEs. Based on the nature of SE to incorporate a neighboring sequence into the circular form and reinsert it into target sites during transposition, we identified the order of intragenomic movements of tmexCD-toprJ gene clusters. Altogether, our findings suggest that most known subgroups of tmexCD-toprJ and their subvariants underwent transpositions among bacterial chromosomes and plasmids via SEs. Hence, a tmexCD-toprJ gene cluster ancestor may have been initially mobilized via SE, subsequently spreading among bacteria and evolving in new hosts.

microbiology↗

bioRxiv2022

Emergence of plasmid-mediated RND-type efflux pump gene cluster tmexCD-toprJ in Shewanella xiamenensis in a water environment

The emergence of the mobile resistance-nodulation-division (RND)-type efflux pump tmexCD-toprJ gene cluster that confers multidrug resistance (MDR), including tigecycline resistance, in gram-negative bacteria poses a global public health threat. However, the spread of such clinically important antimicrobial resistance genes (ARGs) in the natural environment has not yet been well investigated. In this study, we investigated MDR aquatic bacteria in Vietnam. A carbapenem- and tigecycline-resistant Shewanella xiamenensis isolate NUITM-VS2 was obtained from urban drainage in Hanoi, Vietnam, in October 2021. S. xiamenensis NUITM-VS2 showed resistance to most antimicrobials tested, including tigecycline, tetracyclines, carbapenems, cephalosporins, fluoroquinolone, and aminoglycosides. Whole-genome analysis was performed by long- and short-read sequencing, resulting in the complete genome sequence consisting of one chromosome and five plasmid sequences. ARGs and plasmid replicons in the genome were detected using ResFinder with the custom ARG database, including all known tigecycline resistance genes, and PlasmidFinder, respectively. A 152.2-kb IncC plasmid, pNUITM-VS2_2, co-carried two mobile tigecycline resistance genes, tet(X4) and tmexC3.2D3.2-toprJ1. In addition, a 24.8-kb untypeable plasmid, pNUITM-VS2_4, carried the carbapenemase gene blaNDM-1. pNUITM-VS2_2 was transferred to Escherichia coli by conjugation, which simultaneously conferred high-level resistance against many antimicrobials, including tigecycline. To the best of our knowledge, this is the first report of the detection of the mobile RND-type efflux pump gene cluster tmexCD-toprJ in Shewanella species. Our results provide genetic evidence of the complexity of the dynamics of clinically important ARGs among aquatic bacteria, which could be important reservoirs for ARGs in the natural environment.

microbiology↗

bioRxiv2022

Enterococcal linear plasmids adapt to Enterococcus faecium and spread within multidrug-resistant clades.

Antimicrobial resistance (AMR) of bacterial pathogens, including enterococci, is a global concern, and plasmids are crucial for spreading and maintaining AMR genes. Plasmids with linear topology were recently identified in clinical multidrug-resistant enterococci. The enterococcal linear-form plasmids, such as pELF1, confer resistance to clinically important antimicrobials, including vancomycin; however, little information exists about their epidemiological and physiological effects. In this study, we identified several lineages of enterococcal linear plasmids that are structurally conserved and occur globally. pELF1-like linear plasmids show plasticity in acquiring and maintaining AMR genes, often via transposition with the mobile genetic element IS1216E. This linear plasmid family has several characteristics enabling long-term persistence in the bacterial population, including high horizontal self-transmissibility, low-level transcription of plasmid-encoded genes, and a moderate effect on the Enterococcus faecium genome alleviating fitness cost and promoting vertical inheritance. Combined with its broad host range, the linear plasmid is an important factor in the spread and maintenance of AMR genes among enterococci.

microbiology↗

bioRxiv2022

Castration alters the circadian clock and Ucp1 expression in subcutaneous white adipose tissue of mice

Castration increases the risk of metabolic syndromes in mammals. In addition, recent studies have demonstrated that castration increases the expression of Ucp1 in subcutaneous white adipose tissue (scWAT). However, how castration leads to thermogenic aberration remains poorly understood. Here, we found that castrated mice showed elevated Ucp1 expression in scWAT in response to cold exposure with an increase in body temperature in the light phase and a reduction in body weight gain with fat reduction despite showing no alterations in food intake, locomotor activity in a novel environment, and gastrocnemius muscle weight. We also analyzed the circadian behavioral rhythm and the expression rhythm of clock genes in the suprachiasmatic nucleus (SCN) and scWAT. We found that castrated mice showed fluctuations in circadian locomotor activity and a decrease in the expression rhythm of clock genes in the SCN (clock) and scWAT (Bmal1 and Per2). Moreover, the expression of Ucp1 in scWAT was rhythmic, and it was increased in castrated mice, while the expression of clock genes was decreased. These results indicate that castration may impact browning in scWAT through variations in the circadian clock and Ucp1 expression.

physiology↗

bioRxiv2022

Brain and Organoid Manifold Alignment (BOMA), a machine learning framework for comparative gene expression analysis across brains and organoids

Organoids have become valuable models for understanding cellular and molecular mechanisms in human development including brains. However, whether developmental gene expression programs are preserved between human organoids and brains, especially in specific cell types, remains unclear. Importantly, there is a lack of effective computational approaches for comparative data analyses between organoids and developing humans. To address this, by considering the public data availability and research significance, we developed a machine learning framework, Brain and Organoid Manifold Alignment (BOMA) for comparative gene expression analysis of brains and organoids, to identify conserved and specific developmental trajectories as well as developmentally expressed genes and functions, especially at cellular resolution. BOMA first performs a global alignment and then uses manifold learning to locally refine the alignment, revealing conserved developmental trajectories between brains and organoids. Using BOMA, we found that human cortical organoids better align with certain brain cortical regions than other non-cortical regions, implying organoid-preserved developmental gene expression programs specific to brain regions. Additionally, our alignment of non-human primate and human brains reveals highly conserved gene expression around birth. Also, we integrated and analyzed developmental scRNA-seq data of human brains and organoids, showing conserved and specific cell trajectories and clusters. Further identification of expressed genes of such clusters and enrichment analyses reveal brain- or organoid-specific developmental functions and pathways. Finally, we experimentally validated important specific expressed genes using immunofluorescence. BOMA is open-source available as a web tool for general community use.

bioinformatics↗

bioRxiv2022

Cardiac-Gated Spectroscopic Photoacoustic Imaging for Ablation-Induced Necrotic Lesion Visualization: In Vivo Demonstration in a Beating Heart

Radiofrequency (RF) ablation is a minimally invasive therapy for heart arrhythmia, including atrial fibrillation (A-fib), which creates lesions using an electric current to isolate the heart from abnormal electrical signals. However, conventional RF procedures do not involve intraoperative monitoring of the area and extent of ablation-induced necrosis, making the assessment of the procedure completeness challenging. Previous studies have suggested that spectroscopic photoacoustic (sPA) imaging is capable of differentiating ablated tissue from its non-ablated counterpart based on PA spectrum variation. Here, we aim to demonstrate the applicability of sPA imaging in an in vivo environment, where the cardiac motion presents, and introduce a framework for mapping the necrotic lesion using cardiac-gated sPA imaging. We computed the degree of necrosis, or necrotic extent (NE), by dividing the quantified ablated tissue contrast by the total contrast from both ablated and non-ablated tissues, visualizing it as continuous colormap to highlight the necrotic area and extent. To compensate for tissue motion during the cardiac cycle, we applied the cardiac-gating on sPA data, based on the image similarity. The in vivo validation of the concept was conducted in a swine model. As a result, the ablation-induced necrotic lesion at the surface of the beating heart was successfully depicted throughout the cardiac cycle through cardiac-gated sPA (CG-sPA) imaging. The results suggest that the introduced CG-sPA imaging system has great potential to be incorporated into clinical workflow to guide ablation procedures intraoperatively.

bioengineering↗

bioRxiv2022

Pulmonary dust foci as rat pneumoconiosis lesion induced by titanium dioxide nanoparticles in 13-week inhalation study

BackgroundMost toxicological studies on titanium dioxide (TiO2) particles to date have concentrated on carcinogenicity and acute toxicity, with few studies focusing of pneumoconiosis, which is a variety of airspace and interstitial lung diseases caused by particle-laden macrophages. The present study examined rat pulmonary lesions associated with pneumoconiosis after inhalation exposure to TiO2 nanoparticles (NPs). MethodsMale and female F344 rats were exposed to 6.3, 12.5, 25, or 50 mg/m3 anatase type TiO2 NPs for 6 hours/day, 5 days/week for 13 weeks using a whole-body inhalation exposure system. After the last exposure the rats were euthanized and blood, bronchoalveolar lavage fluid, and all tissues including lungs and mediastinal lymph nodes were collected and subjected to biological and histopathological analyses. ResultsNumerous milky white spots were present in the lungs after exposure 25 and 50 mg/m3 TiO2 NPs. Histopathological analysis revealed that the spots were alveolar lesions, characterized predominantly by the agglomeration of particle-laden macrophages and the presence of reactive alveolar epithelial type 2 cell (AEC2) hyperplasia. We defined this characteristic lesion as pulmonary dust foci (PDF). The PDF is an inflammatory niche, with decreased vascular endothelial cells in the interstitium, and proliferating AEC2 transformed into alveolar epithelial progenitor cells. The AEC2 in the PDF had acquired DNA damage. Based on PDF induction, the lowest observed adverse effect concentration for pulmonary disorders in male and female rats in this study was 12.5 mg/m3 and 6.3 mg/m3, respectively. The no observed adverse effect concentration for male rats was 6.3 mg/m3. There was a sex difference in lung lesion development, with females showing more pronounced lesion parameters than males. ConclusionsInhalation exposure to TiO2 NPs caused PDF, an air-space lesion which is an alveolar inflammatory niche containing particle-laden macrophages and proliferating AEC2. This PDF histopathologically resembles some pneumoconiosis lesions (pulmonary siderosis and hard metal pneumoconiosis) in workers and lung disease in smokers, suggesting that it is an early pneumoconiosis lesion caused by exposure to TiO2 NPs in rats and a common alveolar reaction in mammals.

pathology↗

bioRxiv2022

Density-dependent attributes of schooling in small pelagic fishes

Many small pelagic fishes obligately form schools; some of these schools attain a remarkable size. Although the school is a fundamental and important ecological unit and is the site of biological interactions such as competition and predation, information on schooling processes in the field remains scarce. Here, we examined the quantitative relationships between population density and school size, the number of schools, and other school characteristics (i.e., packing density, volume, and cross-sectional area) in three species of small pelagic fishes: Japanese anchovy Engraulis japonicus, Japanese sardine Sardinops melanostictus, and chub mackerel Scomber japonicus. We found that school size increased almost linearly with population density, whereas the number of schools and other characteristics increased non-linearly with population density, whereby the rate of increase slowed radically as population density increased. These results indicate that, at low population densities, an increase in density causes an increase in both school size and the number of schools, whereas at higher population densities, an increase in density triggers the formation of larger schools rather than more schools. Furthermore, we found that the shapes of the relationships of all school characteristics with population density differed among species. Our results indicate that the schooling behaviour of small pelagic fishes is density-dependent, and responses to changes in density are species-specific. Our results provide insight into how biological interactions such as intra- and inter-specific competition and predator-prey interactions mediate the density-dependent processes that underlie the population dynamics and community structure of small pelagic fishes in marine ecosystems.

zoology↗

bioRxiv2022

Tuning of motor outputs produced by spinal stimulation during voluntary control of torque directions in monkeys

Spinal stimulation is a promising method to restore motor function after impairment of descending pathways. While paresis, a weakness of voluntary movements driven by surviving descending pathways, can benefit from spinal stimulation, the effects of descending commands on motor outputs produced by spinal stimulation are unclear. Here, we show that descending commands amplify stimulus-evoked joint torque and the function of intraspinal elements. During the wrist torque tracking task, optimized currents of spinal stimulation over the cervical enlargement facilitated and/or suppressed activities of forelimb muscles. Magnitudes of these effects were dependent on directions of voluntarily-produced torque and positively correlated with levels of voluntary muscle activity. Furthermore, the directions of evoked wrist torque corresponded to the directions of voluntarily-produced torque. These results suggest that spinal stimulation is beneficial in cases of partial lesion of descending pathways by compensating for reduced descending commands through activation of excitatory and inhibitory synaptic connections to motoneurons.

neuroscience↗