Cell Stem Cell 23, 780781 (2018). 58, 118131 (2021). T his RNA-Seq atlas extends upon the analyses of previous gene expression atlases performed using Affymetrix GeneChip technology and describes new methods that compensate for the increase in transcriptome data obtained from next generation sequencing. JAMA Neurol. Neurol. MBP-cholesterol colocalization was quantified using Imaris. To ensure robust nervous system function, mechanisms must exist to accommodate variation in neurite positioning during circuit formation. 2 APOE-associated and lipid pathway changes in APOE4 and AD. (G) bmp6 channel only. W.W, M.W., Q.W. Transcriptome Shotgun Assembly Sequence Database What is the Transcriptome Shotgun Assembly (TSA) Database? In some cases, these compounds are transported to the oocyte via connexin-mediated gap junctions (Su et al., 2008). 3, 89ra57 (2011). Chang, C. C. et al. Song, Q. et al. USA 97, 1103211037 (2000). While these enzymes catalyze similar reactions, in zebrafish Hsd17b3 prefers androstenedione as a substrate while Hsd17b1 prefers estrone (Mindnich et al., 2004; Mindnich et al., 2005; Figure 9A, Figure 9figure supplement 1). Human apoE isoforms differentially regulate brain amyloid- peptide clearance. Bioinformatics 27, 15711572 (2011). Expressed sequence tag or EST assembly was an early strategy, dating from the mid-1990s to the mid-2000s, to assemble individual genes rather than whole genomes. CAS Oxford Nanopore Technologies products are not intended for use for health assessment or to diagnose, treat, mitigate, cure, or prevent any disease or condition. Chu, E., Mychasiuk, R., Hibbs, M. L. & Semple, B. D. Dysregulated phosphoinositide 3-kinase signaling in microglia: shaping chronic neuroinflammation. (c) eQTLs of MRPL45 colocalized with GWAS signals of somatic cell score (SCS) in blood. Neurosci. A.T. acknowledged funding from the Biotechnology and Biological Sciences Research Council through program grants BBS/E/D/10002070 and BBS/E/D/30002275, Medical Research Council research grant MR/P015514/1 and HDRUK award HDR-9004. Cesar, A. S. M. et al. Extended Data Fig. was supported by the National Science Foundation of China-Pakistan Science Foundation Joint Project (31961143009) and National Key R&D Program of China (2021YFD1200900 and 2021YFD1200903). and JavaScript. Proc. Y. Yu. Article Nanopore sequencing offers advantages in all areas of research. To obtain This makes the Discussion section repetitive, as it mainly recapitulates points already made in the results. WebMedicinal Plants Genomics Resource (MPGR) is an NIH funded project that provides transcript assembly sequences and annotation for those sequences from fourteen plants that are known or believed to contain natural compounds that are of interest to medical researchers. Moreno-Jimenez, E. P. et al. However, pre-follicle cells have not been molecularly identified in any teleost (Beer and Draper, 2013; Cao et al., 2019b). Ye, S. et al. WebPicard. Two of these libraries were prepared using a dissociation method that favored isolation of single somatic cells. (e) Distribution of read lengths. Further mutational analysis is required to test these relationships. Bars represent means from four biological replicates. Some genes, like col1a1a, are expressed in all stromal cells, while others, such as stm, are only expressed in a specific subcluster. Significant TWAS results from S-MultiXcan (Bonferroni corrected P<4106) aggregated across tissues. 1 Data summary of publicly available RNA-Seq data. The two barley genotypes XZ5 and Tadmor were also used for transcriptome profiling as described by Qiu et al.. A greenhouse hydroponic experiment was designed with these two genotypes for control and PEG-induced water deficit treatment (20 % PEG-6000) with an equivalent osmotic potential of 0.53 MPa . All references to smFISH have been changed to HRC RNA-FISH.. However, after several attempts we have not observed convincing signal. Autocrine Mfge8 signaling prevents developmental exhaustion of the adult neural stem cell pool. Results, Page10, line24-28, and page14, line20-23. 7, 841846 (2004). Microglia shape adult hippocampal neurogenesis through apoptosis-coupled phagocytosis. We obtained germ cells with an average of 2510 genes/cell, 11,294 transcripts/cell and somatic cells with an average of 854 genes/cell, 4812 transcripts/cell. Gao, Y. et al. & Kellis, M. A multiresolution framework to characterize single-cell state landscapes. The All-but-One conditional analysis across the individual GWAS signals shows that only rs135928140 in strength GWAS is colocalized with cis-eQTLs of TIGAR in muscle, when conditioning on the remaining three signals. Finally, we used gene knockout experiments to determine the roles of foxl2l and wnt9b for oocyte development and sex determination and/or differentiation, respectively. Article Fast computation and applications of genome mappability. 4, 13131317 (1998). To further assess this developmental trajectory, we performed a dedicated trajectory analysis and inferred pseudotime using Monocle 3 (Cao et al., 2019a). Nat. WebDe-Novo Assembly; RNA-Seq Alignment; Quantify Expression Run and Save Workflows, Visualize Data, Genome Browser, Filter and Sort Large Tables, General Bioinformatics Tools, and More. Human hippocampal neurogenesis persists throughout aging. (c) Distribution of uncovered CpGs (< 5) along the entire genome. All rights reserved. Commun. WebThe branches of science known informally as omics are various disciplines in biology whose names end in the suffix -omics, such as genomics, proteomics, metabolomics, metagenomics, phenomics and transcriptomics.Omics aims at the collective characterization and quantification of pools of biological molecules that translate into the Neurosci. These cells are likely early oocyte progenitors (Figure 2E and G). Collaborative development of 2-hydroxypropyl--cyclodextrin for the treatment of Niemann-Pick type C1 disease. (d) The cis-eQTL cumulative proportion plot of |log2(aFC)| distribution across 9 tissues with variable sample sizes. Wen, X., Lee, Y., Luca, F. & Pique-Regi, R. Efficient integrative multi-SNP association analysis via deterministic approximation of posteriors. i, Pairwise correlations of cell-type-specific individual-level transcriptomic profiles (average expression values across cells of a given type, N=32 profiles per cell type). Bars represent the mean number of droplets per cell for each condition, error bars represent standard deviation. Mol. The workshop consisted of discussions, lectures, and hands-on tutorials, to cover topics important to getting started constructing 7, 500507 (2012). VAT will be added later in the checkout.Tax calculation will be finalised during checkout. Code resource at github website. Histol. Finally, we link gene expression in different tissues to 43 economically important traits using both transcriptome-wide association and colocalization analyses to decipher the molecular regulatory mechanisms underpinning such agronomic traits in cattle. The trypsin reaction was stopped by adding 500 l of 20 mg/ml trypsin inhibitor (Cat# 100612; MP Biomedicals) in L15. Cipriani, S. et al. (f) Distribution of cis-eQTLs around the TSS (1Mb up- and down-stream) across all 23 distinct tissues. Could the authors also report SL of the fish used at 40 and 50 dpf, so that others can reproduce accurately the stage? PubMed Extended Data Fig. We have changed this sentence to say: correlated with the trajectory determined by our initial germ cell re-cluster analysis, We have modified this sentence to make it more understandable. However, consistent with our hypothesis, CD34 is known to be expressed in diverse progenitor cell populations in mammals (e.g. Blood cells and a small number of germ cells were also removed from the whole ovary libraries. WebRNA-Seq Atlas of Glycine max: A guide to the soybean transcriptome. To address this, we identified the nanos3-expressing cells in our dataset. NEBULA is a fast negative binomial mixed model for differential or co-expression analysis of large-scale multi-subject single-cell data. Nat. (a) cis-eQTLs discovered in monocytes, and mammary gland show enrichments for top SNPs (top 10%) associated with age at first calving (AFC) (P=0.001, the two-sided permutation test with 1,000 times), and somatic cell score (SCS) (P=0.001, the two-sided permutation test with 1,000 times) respectively, compared to the null expectation (shown in gray) defined by Not eQTLs. Nat. Lin, Y.-T. et al. Quantification was performed using ImageJ software with the same threshold setting for all images and conditions. Hayes, B. J. Genome and Transcriptome Assemblies can be submitted through the Genomes and TSA portals, respectively. 15, 13991406 (2012). Furthermore, it would be interesting to make parallels with other scRNAseq datasets from other organisms in the discussion. Astrocytes decrease adult neurogenesis during virus-induced memory dysfunction via IL-1. Thus, we were able to identify all previously observed and expected cell types in our dataset. k, Fraction of subjects lacking cells of a given type. Consistent with this hypothesis, wild-type testes at 50 dpf contained germ cells at all stages of spermatogenesis, including early spermatocytes and mature spermatozoa (Figure 5D), while the transitioning gonad contained only spermatocytes, but no mature spermatozoa (Figure 5G). Follicle cell proliferation is not surprising given that the surface area of the zebrafish oocyte increases 1400-fold during oogenesis. However, nonuniform expression of star was unexpected as StAR is thought to catalyze the rate-limiting step in steroid synthesis the transfer of cholesterol from the outer to the inner mitochondrial membrane (Miller and Auchus, 2011). (b) Significant GO terms for genes with tissue-specific hypomethylated promoters (P<0.05). From day 12 to 20 dpf, each tank of larvae is fed two drops concentrated brine shrimp nauplii, twice daily. PubMed To determine the location of pericytes in the ovary, we used plp1b as a marker because, in contrast to notch3 and pdgfrb, our data indicate that plp1b is specific to pericytes. ROSMAP is supported by NIA grants P30AG20262, R01AG15819, R01AG17917, U01AG46152, U01AG 61356 and P30AG72975. Given the expression of fgf24 in surface epithelial cells of the early gonad (Leerberg et al., 2017) and the role of claudin proteins in forming tight junctions between epithelial cells (Tsukita et al., 2019), it was plausible that subcluster 5 cells corresponded to the ovarian surface epithelium. In mammals, retinoic acid signaling is required to induce germ cell entry into meiosis (Bowles et al., 2006; Koubova et al., 2006). n = 70, N = 3. Xiang, R. et al. July 22, 2022. Notably, subcluster 3 cells also uniquely express iroquois homeobox 3a and 5a (irx3a and irx5a; Dildrop and Rther, 2004) whose orthologs in mammals are required for pre-granulosa cell development in the embryonic mouse ovary (Figure 4B and C, Figure 4figure supplement 2C and D; Fu et al., 2018; Kim et al., 2011). & Weisgraber, K. H. Apolipoprotein E structure: insights into function. Moreno-Jimenez, E. P., Terreros-Roncal, J., Flor-Garcia, M., Rabano, A. analyzed the snRNA-seq dataandscATAC-seq data. 200 l of 7 mg/ml trypsin (Cat# LS003708; Worthington) in L15 were added and incubated on an orbital rotator for 10 min or until a minimal amount of cell clumps was observed. Kamphorst, J. J., Fan, J., Lu, W., White, E. & Rabinowitz, J. D. Liquid chromatography-high resolution mass spectrometry analysis of fatty acid metabolism. Using subcluster analysis, we have defined subpopulations within these broad cell-type classifications and validated these subpopulations by determining where they reside in the ovary using hybridization chain reaction RNA fluorescent in situ hybridization (HCR RNA-FISH). Good point. Nature Genetics thanks Ben Hayes and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Acad. Glia 51, 173186 (2005). From early next year, we will no longer make accept/reject decisions after peer review. Article Estimation of linkage disequilibrium in a sample of the United Kingdom dairy cattle population using unphased genotypes. Methods 14, 10831086 (2017). was supported by appropriated project 8042-31000-002-00-D, Improving Dairy Animals by Increasing Accuracy of Genomic Prediction, Evaluating New Traits, and Redefining Selection Goals of ARS-USDA. (g) Correlation of effect sizes (fastQTL slope) of cis-eQTLs and aFC of matched loci tested by allelic specific expression (ASE) analysis in adipose (Spearmans rho = 0.75, the two-sided Students t-test: P<2.210308) and muscle (Spearmans rho = 0.68, the two-sided Students t-test: P=2.110162). 4 Genotyping and imputation of variants from RNA-Seq data. Cell Stem Cell 23, 2530 (2018). Prior to assessing doublets, we performed quality control on the dataset. USA 115, E610E619 (2018). the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in All raw data analyzed in this study are publicly available for download without restrictions from SRA (https://www.ncbi.nlm.nih.gov/sra/) and BIGD (https://bigd.big.ac.cn/bioproject/) databases. Nat. We agree. However, unlike mammals, zebrafish females can produce new follicles throughout adult life from a population of self-renewing GSCs and pre-follicle progenitor cells, thus allowing the study of these unique cell types in a tractable vertebrate research animal. Nature Genetics Tissue was collected from each fish during dissection for genotyping post-dissection. Altogether, these data argue that stm-expressing cells form the OCE and may also play a role in GSC niche formation and/or function. Sequence long targeted regions and expand on the limitations of traditional targeted sequencing approaches. Psychiatry 54, 10251034 (2003). Wild-type adult female zebrafish (D) has characteristic light-yellow pigmentation on ventral belly and a prominent anal papilla (highlighted with red dashed lines) (D). 20, 1149 (2019). Get the most important science stories of the day, free in your inbox. (A) Dot plot showing the relative expression of select genes in the stromal cell subclusters. An allele consisting of a 57 bp deletion was identified, which removed the 3 end of the first exon and 5 end of the first intron, leading to missplicing and a frameshift leading to a premature stop codon. Habib, N. et al. S.L., Y.G., O.C.-X., S.W., L.F., R.X., W.C., B.L., C.X., Y. Yao, Z.Y. Paredes, M. F. et al. We therefore propose that these cells are stromal progenitor cells. and X.W. Masuda, T., Sankowski, R., Staszewski, O. This will be expanded in the future to include other types of GenBank submissions. Mizrak, D. et al. i, Gene expression distribution in iPSC-derived oligodendroglia. https://doi.org/10.1038/s41588-022-01153-5, DOI: https://doi.org/10.1038/s41588-022-01153-5. c, Representative images of Bodipy-cholesterol staining, with markers for microglia (IBA1), astrocytes (GFAP) and oligodendrocytes (OLIG2) in the prefrontal cortex of APOE4-carriers (n=four individuals). Specifically, pathway 1 produces testosterone as an intermediate while pathway 2 produces estrone (E1; Figure 9A). Because sex ratios in any cross can vary, it is important to compare sex ratios in mutants to their wild-type siblings. What is the Nanopore Community? We reasoned that the ovarian cavity and its associated membrane would be more pronounced when the body cavity was devoid of mature oocytes. Unlike the development of many organs, such as the brain and heart that form during the first few days of development, zebrafish gonads do not begin to form until the larval stage (5 days post-fertilization). & Abecasis, G. R. METAL: fast and efficient meta-analysis of genomewide association scans. Phylogenetic trends are important to consider in the assessment of transcriptome From FastQ data to high confidence variant calls: the Genome Analysis Toolkit best practices pipeline. Boldrini, M. et al. For example, lhx9 appeared to have higher expression in cells distal to the subcluster 1 in the UMAP while gsdf and bone morphogenetic protein 6 (bmp6; Smith et al., 2006) appeared higher in cells proximal to subcluster 1 (Figure 4A and C). To access the data, a data use agreement is needed. The cells were centrifuge for 3 min at 300 g and the supernatant carefully removed. A gja11-specific template was then generated by PCR using Phusion polymerase (Cat# M0530L; New England Biolabs) and the following primer pair: fwd, 5-CCCTGAGCAGTCTTTTCGAGCCT-3; rev, 5-taattaatacgactcactataggGTGCTTAAAGCCAGGCGGTCA-3. As a first step, we performed nonzero matrix factorization (NMF) to find gene modules, which are sets of genes that are co-expressed within cell clusters and subclusters and may therefore identify co-regulated genes (Figure 2figure supplement 5; Brunet et al., 2004; Farrell et al., 2018; Siebert et al., 2019). However, while 95% of wnt4 mutants develop as males, we found that ~5% can develop as females. Genes are not significant cis-sGenes in any tissues. Non-human and environmental metagenomics data should go to the Metagenome archive. Bars represent means. was supported by NIH grant RF1-AG0540124 and the MIT BCS Henry E. Singleton Graduate Student Fellowship; D.v.M. Li, Y. I. et al. All authors edited and proofread the manuscript. Bars represent mean intensity from all mice for each genotype, and error bars represent standard deviation. The unipotent GSCs can divide both to replenish their numbers while also producing cells that differentiate into sex-specific gametes, either eggs or sperm. Zebrafish therefore provide a unique opportunity to study female GSCs in a vertebrate. Whether adult hippocampal neurogenesis (AHN) persists in adult and aged humans continues to be extensively debated. (C) Gene expression UMAP plots of select genes. This gene was chosen because it is specific to this cell population and appeared to have relatively high expression. Cells expressing the indicated gene are colored purple, and the relative intensity indicates relative expression levels (intensity scale for each plot is on the right). Wild-type allele yields a 238 bp product, wnt9b(fb209) allele yields a 181 bp product. Here, to provide a better understanding of AHN in primates and to reveal more novel markers for distinct cell types, droplet-based single-nucleus RNA sequencing (snRNA-seq) is used to investigate the cellular heterogeneity and molecular characteristics of the hippocampi in macaques across the lifespan and in aged humans. On page 20, paragraph 2, "Unfortunately, we have yet to identify where these cells reside in the ovary." Liu, H. et al. Data points represent the mean of four images from one biological replicate. To learn how to use Advanced Search Builder please refer to Search in SRA. (c) Tissue-sharing patterns of cis-sQTL. 47, D1005D1012 (2019). Extended Data Fig. In future development of de novo transcriptome assembly software, the incorporation of phylum-specific information may be useful in improving the overall quality of assemblies for different taxa. The black line is the smoothed curve fitted by a generalized additive model using geom_smooth function from ggplot2 (v3.3.6) in R (v3.4.1). p values were calculated using an unpaired, two-tailed students t-test. Hu, Z. L., Park, C. A. A list of genes with enriched expression in each cluster can be found in Supplementary file 1. Neuroscientist 17, 7993 (2011). The suspension was monitored every 10 min until no or a minimal number of cell clumps were observed (~30 min). New users get started quicker with access to Nanopore Learning.Discussion and collaboration drives experimental success with full access to documentation and support and as a member of our community you will help shape the 50, 151158 (2018). We have modified as requested. Finally, the motif enrichment analysis also identified multiple genes with known functions in germ cell regulation that contain Bhlhe23 binding motifs, such as cnot1, which encoded a component of the CCR4/Not deadenylase complex that functions with Nanos proteins to regulate translation of select targets in GSCs (Suzuki et al., 2012) In addition, dnmt3bb.1, tsmb4x, and mcm6 are expressed in GSC and have putative Bhlhe23 sites within 2 kb of their transcription start sites, suggesting that Bhlhe23 may regulate the expression of multiple genes (Figure 2figure supplement 5F). Transcriptome assembly validation was done using Busco, Detonate and Transrate. PubMed Central WebTranscript survey sequence assemblies should go to the Transcriptome Shotgun Assembly (TSA) archive. In the interest of shortening the paper and reducing redundancy, Results and Discussion could be combined into one section, with a final "Conclusion" section at the end for any additional points not discussed in the 'Results and Discussion" section. Thank you. Mean fluorescence intensity quantified using FIJI ImageJ software. Nat. Cell Research As previously reported, nanos3 is expressed at high levels in early oocytes, but we also detected expression in the apparent GSC subpopulation, confirming that nanos3 is expressed in GSCs as suggested by the nanos3 mutant phenotype (Figure 2B and C). all of the known stromal cell subtypes have clear mammalian counterparts, except the ovarian cavity epithelium (OEC). Axonal area was calculated by measuring the area immunoreactive to neurofilament, and normalized to APOE3/3. BMC Biol. Boxes are exons, UTR sequences are shaded. Cell Res 32, 729743 (2022). The standard length (SL) of the fish is a much better way to report the stage for zebrafish >5 dpf, because it more accurately reflects the developmental stage than dpf. What is the Nanopore Community? For example, the transcription factors odd-skipped related 1 (osr1; Tena et al., 2007) and transcription factor 21 (tcf21; Knight et al., 2008) are expressed specifically in stromal cell subcluster 3 cells. This reference dataset is available in a processed and interactively browsable form through the Broad Institute Single Cell Portal (available here) and will serve as a resource to greatly enhance future studies of ovarian development and function in unprecedented detail. Manolis Kellis or Li-Huei Tsai. From 20 to 30 dpf, each tank of larvae is fed three drops concentrated brine shrimp nauplii twice daily. Previous studies have shown that cxcl12a is essential for directing migrating PGCs to the forming gonad during embryogenesis (Doitsidou et al., 2002). A1217701; Thermo Fisher) in L15 and incubated on an orbital rotator at room temperature for 15 min. Watch the keynote and other presentations from our 2022 event! Remer, J. et al. Tools and resources for plant genomics. Briefly, the expression matrices generated from Cell Ranger (germ cell library zx1_40gc, whole ovary libraries zx2_40ov and zx4_40ov) were first processed with SoupX (v0.3.1; Young and Behjati, 2020) to remove ambient RNA. Ddx4 indirect immunofluorescence (green) labels all germ cells. Wnt-mediated activation of NeuroD1 and retro-elements during adult neurogenesis. Gold, B. T., Powell, D. K., Andersen, A. H. & Smith, C. D. Alterations in multiple measures of white matter integrity in normal women at high risk for Alzheimers disease. A., Dermitzakis, E. T. & Delaneau, O. (F, F) Higher-magnification views of regions boxed in (F) showing that lhx9+ cells (arrows) surround the germ cells. Thank you for catching that! BMC Bioinform. The National Center for Genome Analysis Support (NCGAS) offered this three-day workshop on HPC usage and de novo transcriptome assembly. Locked-down, research-validated devices for applied sequencing applications. The area of neuronal axon (immunoreactive against Neurofilament) positive for MBP was calculated using Imaris, and each experimental condition was compared to the APOE3/3 control condition. Clustering analyses were conducted using Seurat (v3.1.0; Stuart et al., 2019) and the SCTransform workflow to normalize, identify variable genes, and perform scaling of the data. 31, 2030 (2011). (a) The number of publicly available RNA-Seq samples increases rapidly over years by fitting a second order polynominal model. Box plots depict the interquartile range (IQR), whiskers depict 1.5 IQR. We found that in heterozygous animals GFP was expressed in a subset of premeiotic germ cells in the ovary (Figure 3B), identical to the pattern determined using HCR RNA-FISH (Figure 2EH). The work was enabled in part by a community data portal of the U.S. [Read More], JGI is a DOE Office of Science User Facility managed by Lawrence Berkeley National Laboratory 1997-2022 The Regents of the University of California, NEWS: A New Actinobacterial Chapter in the Genomic Encyclopedia of Bacteria and Archaea, LISTEN: A Tour of the JGIs Sequencing Pipeline, SCIENCE HIGHLIGHT: Model fern reveals insight into DNA thievery in ferns, NEWS: A Better Way to Find RNA Virus Needles in the Proverbial Database Haystacks. g, Fold change for unfolded protein response genes in APOE4/4 vs APOE3/3 oligodendroglia (N=3 per genotype) from panel e (adjusted p-value<0.05, negative binomial distribution). |log2(aFC)|, that is, the log2 transformed allelic fold change, which is used to measure the effect size of cis-eQTL. (e) Distribution of cis-eQTLs around TSS (1Mb up- and down-stream) in adipose. Sequence-based association analysis reveals an MGST1 eQTL with pleiotropic effects on bovine milk composition. This data has allowed us to identify all known stages of germ cells, from GSCs to pre-follicle stage oocytes (stage IA; Selman et al., 1993), as well as the somatic cell populations, including follicle, theca, and stromal cells. Blanchard, J.W., Akay, L.A., Davila-Velderrain, J. et al. Theca cells are found around and between oocytes but are otherwise not well characterized in zebrafish. Profile and regulation of apolipoprotein E (ApoE) expression in the CNS in mice with targeting of green fluorescent protein gene to the ApoE locus. Trapnell, C. et al. 7, 89102 (2019). A number of the same points are made in the Discussion as in the Results section, including, for example, the conclusions made about E2 production. This is addressed in answer to the following question. Somatic ovary cells were dissociated using a modification of Elkouby and Mullins, 2017. Nat. Springer Nature or its licensor holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law. Ottinger, E. A. et al. was supported in part by AFRI grant numbers 2020-67015-31398 and 2021-67015-33409 from the NIFA. PLoS Genet. Top. Acta Neuropathologica Communications ISSN 1061-4036 (print). Wang, W., Wang, M., Yang, M. et al. Urban, N., Blomfield, I. M. & Guillemot, F. Quiescence of adult mammalian neural stem cells: a highly regulated rest. WebEST Assemblers. Cell Syst. 6 Lipidomics on iPSC-derived oligodendroglia. Regardless, cxcl21a provides a convenient marker for determining the location of this cell population in the ovary. Therefore, a multi-assembler approach, followed by a reduction step, is often sought to generate an improved representation of the assembly. To identify novel markers of oocyte progenitor cells, we searched for genes with enriched expression in this subpopulation (subcluster 2 in Figure 2A; Supplementary file 2). Neuronal ApoE upregulates MHC-I expression to drive selective neurodegeneration in Alzheimers disease. And a lot of content courtesy of Matt MacManes (@macmanes) Overview. Appl. As to the point about the definition of early germ cells, we have better defined what germ cell-stage the various marker genes identify when these markers are first introduced in the germ cell section (page 7, lines 12-15). ROSMAP resources can be requested at https://www.radc.rush.edu. Whole fish were fixed overnight in neutral alcoholic formalin fixative (29.5% ethanol, 10% formalin, 50 mM sodium phosphate buffer, pH 7.2). Transcriptome profiling. Histological images were collected using a Zeiss Axiophot microscope equipped with a Leica DFC 500 camera. a, Co-localization of immunohistochemistry against lipid-droplet associated protein perilipin-1 (PLIN1) with Bodipy-cholesterol staining in the human prefrontal cortex from APOE3/3 and APOE3/4 individuals (n=3 imaged per genotype). Transcriptome assembly is a crucial first step for studying the molecular basis of phenotypes of interest using RNA-Sequencing (RNA-Seq). Here, to gain more comprehensive insights into the impact of APOE4 on the human brain, we performed single-cell transcriptomics profiling of post-mortem human brains from APOE4 carriers compared with non-carriers. Cell Neurosci. In zebrafish, the ability to produce oocytes is a requirement for female development, thus loss of oocyte production, which occurs in foxl2l mutants, results in all-male development (Rodrguez-Mar et al., 2010; Shive et al., 2010). From days 10 to 30, the tanks are maintained with a slow continuous flow of system water. Some genes, like gsdf, are expressed in all follicle cells, while others, such as lhx9, are only expressed in distinct subclusters. was supported by NIH grant U01-NS110453; Y.-T.L. The P value is obtained by the two-sided Students t-test. Troy+ brain stem cells cycle through quiescence and regulate their number by sensing niche occupancy. Cell viability and number were determined using propidium iodine (Cat# P1304MP; Thermo Fisher) and Hoechst 33342 (Cat# H3570; Thermo Fisher) staining on a Fuchs-Rosenthal hemocytometer (Cat# DHC-F01; Incyto). The plants utilized by MPGR are Atropa belladonna, Camptotheca Protoc. Xiang, R. et al. WebEST Assemblers. Resolve complex structural variants and repetitive regions, Simplify de novo genome assembly and improve existing reference genomes, Enhance metagenomic identification of closely related species and distinguish plasmid from genome, Sequence entire microbes in single reads in real-time, Explore epigenetic modifications using direct, long-read DNA sequencing. Bars represent means, error bars represent standard deviation, and p values were calculated using one-way ANOVA with Bonferroni correction. Following quality control, data cleaning, and the removal of red blood cells, doublets, and ambient RNA, we recovered a total of 25,089 single-cell transcriptomes that comprise 10,658 germ cells and 14,431 somatic cells (Figure 1figure supplement 2; see Materials and methods). Ongen, H., Buil, A., Brown, A. Fluorescence RNA in situ hybridization, HCR fluorescence RNA in situ hybridization, and immunohistochemical staining samples were mounted with ProLong Diamond Antifade Mountant and imaged with either Olympus FV1000 laser scanning confocal microscope or Zeiss LSM 880 Airyscan microscope. We used GSC-specific nanos2 expression to identify the root of the trajectory and measured pseudo-time. Commun. Boxplots indicate median, 25th and 75th percentiles. Find out more about analysing nanopore DNA sequencing data and access our online tutorials. The Science Synaptic pruning by microglia is necessary for normal brain development. Nat. All zebrafish used in this study were raised in a recirculating aquatic system (Aquaneering, Inc, San Diego, CA), with the following water parameters: water temperature, 2829C; pH, 7.07.2 (maintained with Sodium Bicarbonate); conductivity, 600700 S (maintained with Crystal Sea Marine Mix, Aquaneering, CA); ammonia, 0 ppm; nitrate, 4060 ppm; water exchange, 10% total volume/day; water source, deionized water. a, Curation process for APOE-associated pathway database. Whole genome sequence assemblies should be submitted to WGS. CAS In the adult ovary, early germ cells localize to a discrete region on the surface of the ovary, called the germinal zone, which has been proposed to be the GSC niche (Beer and Draper, 2013). Gene expression analysis argues that cells within subcluster 4 are a mitotic subpopulation of subcluster 0 cells as these cells have nearly identical gene expression with cells in subcluster 0 but also express high levels of genes necessary for mitosis, such as pcna (Figure 4B and C). Perhaps surprisingly, no adapter sequences were found in the assembly of the untrimmed version of this dataset. Thus, forward genetic screens have identified very few genes required for gonad development. Aizarani, N. et al. Genet. The lengthy results would be more concise if some of those points were addressed only in the discussion and this would greatly improve the legibility of the manuscript. APOE4/4 group comprised 3 male and 5 female subjects, all with an AD diagnosis. Tg(kdrl:dsRed) is expressed in vascular endothelial cells, while acta2:egfp is expressed in vSMC (Kikuchi et al., 2011; Whitesell et al., 2014). A genome reference file was generated with Cell Rangers mkref function using the GRCz11 zebrafish genome obtained from the Ensembl Genome Browser with alternative loci scaffolds removed and the filtered GTF file. Publishers note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. CAS All.SNPs are those 31,377,923 imputed variants common in the two imputation processes (that is, the genotype imputation based on RNA-Seq SNPs and that based on SNP array). Bars represent mean across all wells, error bars represent standard deviation. USA 116, 1939819408 (2019). P values calculated using unpaired, two-tailed students t-test. Get immediate access to your DNA sequencing results with real-time data streaming. vSMC are characterized by expression of -smooth muscle actin (acta2) and transgelin (tagln, formally known as SM22; Bahrami and Childs, 2018). & Daetwyler, H. D. 1000 Bull Genomes Project to map simple and complex genetic traits in cattle: applications and outcomes. Lambert, J. C. et al. Commun. Prior to imaging, sections were stained 1 hr with 1 g/ml 4,6-diamidino-2-phenylindole (DAPI) in PBS, cleared in glycerol-PBS gradient (30, 50, and 70%) for 10 min each, and then mounted with ProLong Diamond Antifade Mountant. WebArchived workshop: De novo Assembly of Transcriptomes using HPC resources. Muscle_G1 (n=435) and Muscle_G2 (n=435) are two replicates of muscle samples by dividing the whole muscle samples randomly into two groups. fk, Pathway activity scores for Cholesterol biosynthesis III (via desmosterol) stratified by APOE genotype and/or AD pathology (p-values, linear model, unadjusted, n=6 per group). A revised airway epithelial hierarchy includes CFTR-expressing ionocytes. Detailed information of 144 whole-genome bisulfite sequence (WGBS) data. Fast, sensitive and accurate integration of single-cell data with Harmony. Gene expression UMAP plots of select genes. (b) eQTLs of CACNB3 colocalized with GWAS signals of protein percentage in blood. Scale bar in (C) for (CG) 50 m. Unraveling human adult hippocampal neurogenesis. Expect-cells was set to 10,000 based on estimated cell recovery. Dean, D. C. 3rd et al. More accurately identify microbes and their abundance with real-time results. A neuronal molecular switch through cell-cell contact that regulates quiescent neural stem cells. Tissue specificity of transcriptome and methylome. The USDA is an equal opportunity provider and employer. At 8 dpf, an additional 250 ml 4 ppt seawater is added to each tank. Sample extraction and library preparation can also be automated using VolTRAX, a portable, USB-powered device saving hands-on time and minimising the potential for user error. (f) has the same color key as (d). The optimal K was identified as 36 for the germ cell dataset. To obtain Bioinformatics 43, 11.10.111.10.33 (2013). Human adult neurogenesis: evidence and remaining questions. Artificial-intelligence tools are becoming increasingly adept at autogenerating text, raising concerns around authorship and research integrity. The NHGRI-EBI GWAS Catalog of published genome-wide association studies, targeted arrays and summary statistics 2019. Abbott, L. C. & Nigussie, F. Adult neurogenesis in the mammalian dentate gyrus. A number of tools have been developed for transcriptome assembly. The shaded area around the lines represents the 95% confidence interval for the fitted values (the lines). Disord. f, TEM on myelinated axon from iPSC-derived neuron and oligodendrocyte co-culture, suggesting presence of myelin rings. Transcriptomic taxonomy and neurogenic trajectories of adult human, macaque, and pig hippocampal and entorhinal cells. & He, Q. Y. clusterProfiler: an R package for comparing biological themes among gene clusters. Fast and efficient QTL mapper for thousands of molecular phenotypes. Fang, L. et al. Distributions represent distances between each post-mortem cell type (x-axis) and iPSC oligodendroglia in scaled gene space (N=32 per post-mortem cell type, N=6-8 for iPSC cell types). Article Data is provided in standard FASTQ and FAST5 formats suitable for analysis using a range of downstream tools, including the EPI2ME platform, which provides easy access to a growing number of real-time analysis workflows. conceived and designed the project. Mohammadi, P., Castel, S. E., Brown, A. WebEukaryotic DNA is wrapped around a core of histone proteins (), hierarchically organized into chromatin and having limited accessibility to external factors.Chromatin accessibility to regulatory factors is dynamically regulated in a cell typespecific manner (2, 3).Various genome-wide methods have been developed to identify active regulatory regions and a, Study cohort description by APOE genotype groups: APOE3/3-carriers (grey), APOE3/4-carriers (pink), APOE4/4-carriers (red); balanced according to positive (glow) or negative pathological diagnosis (no glow) Cartoons generated with BioRender. Reference is missing. Sato, K. Effects of microglia on neurogenesis. & Reecy, J. M. Building a livestock genetic and genomic information knowledgebase through integrative developments of Animal QTLdb and CorrDB. (A) Two possible pathways for E2 synthesis starting with androstenedione. Gene dose of apolipoprotein E type 4 allele and the risk of Alzheimers disease in late onset families. Neurosci Res. Cao, J. et al. CAS 41, 25412553 (2021). The DR2 values of SNPs within each window are then averaged for plotting. Protoc. Rev. Single-cell transcriptomics reveals a population of dormant neural stem cells that become activated upon brain injury. We describe the transcriptomic landscape of more than 100 tissues/cell types and report hundreds of thousands of genetic associations with gene expression and alternative splicing for 23 distinct tissues. Asterisk in (F) indicates a cell double-positive for nanos2 and foxl2l. Monocle 3 (v0.1.3) was used for trajectory analysis (Cao et al., 2019a). Cortex 28, 24582478 (2018). 19, 253262 (1986). (C) cxcl12a-expressing interstitial cells localize around early-stage oocytes (stage IB), but not around stage II oocytes. R.X. We recommend generating a single Trinity assembly based on combining all reads across all samples as inputs. Lipid droplets were quantified in two different isogenic sets of APOE3/3 and APOE4/4 oligodendroglia, that were generated from different individuals. Transcriptome dynamics of hippocampal neurogenesis in macaques across the lifespan and aged humans. This is further supported by the expression of decorin (dcn; Shintani et al., 2000), which encodes a proteoglycan component of the extracellular matrix, and nuclear receptor subfamily 2, group F, member 2 (nr2f2, formerly coup-tfII; Fjose et al., 1995), both of which are expressed in mammalian ovarian stromal cells (Figure 6A; Pereira et al., 1995; Wagner et al., 2020). Pericytes are generally solitary cells associated with small diameter blood vessels (arterioles, capillaries, and venules) while vascular smooth muscle cells (vSMCs) are associated with large blood vessels where they form a continuous coating (Gaengel et al., 2009). To identify putative GSC-specific transcription factors, we focused our analysis on modules 7 and 25, which contained genes whose expression was specifically enriched in the GSC cluster (Figure 2figure supplement 5). "E2 production also requires steroidogenic theca cells which produce androstenedione, the precursor that follicle cells use for E2 synthesis (ref)." Rev. Neurosci. Aging as a risk factor for neurodegenerative disease. This study was supported by the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, Neuroscience Center, and the China Human Brain Banking Consortium. Alternatively, StAR may not be required for androstenedione production by theca cells and thus E2 production. Consistent with the biological function of APOE2,3,4,5,6, APOE4 significantly altered signalling pathways associated with cholesterol homeostasis and transport. nanos2 encodes an RNA-binding protein that is expressed specifically in GSCs (Beer and Draper, 2013) while rec8a encodes a meiosis-specific member of the Rad21 cohesin family that is loaded onto chromosomes during premeiotic S-phase (Crespo et al., 2019). Our initial analysis revealed genes that are differentially expressed between major cell types but did not distinguish possible subpopulations. The aging systemic milieu negatively regulates neurogenesis and cognitive function. Our pharmacological and genetic studies reveal that b1-AR-Gas-YAP signaling axis is involved in regulating postnatal cardiomyocyte proliferation. Three major conclusions can be drawn from this analysis: (1) the major pathway for E2 biosynthesis in the zebrafish ovary occurs via pathway 2, (2) Hsd17b1, not Cyp19a1a, catalyzes the final enzymatic step in E2 synthesis, and (3) follicle cells are the major source of E2 production in the zebrafish ovary (Figure 9). Although these general cell populations have been observed in the zebrafish ovary, there are significant gaps in knowledge regarding the identification of the full diversity of cell types present in the ovary or the functional roles they play. A common feature of oocyte progenitor cells, including those in zebrafish ovaries, is that they divide with complete nuclear division but incomplete cytoplasmic division, thus creating a multicell cyst with synchronized developmental progression (Marlow and Mullins, 2008; Pepling et al., 1999). Commun. Cells expressing higher levels of foxl2l could be divided into two populations, based on morphology and gene expression. All of the authors contributed to writing the paper and making the figures. P value was calculated using unpaired, two-tailed students t-test. Navarro Negredo, P., Yeo, R. W. & Brunet, A. Supplementary Table 10. Further analysis of gene expression following reclustering of only the theca cell population suggests that theca cells are not a homogeneous cell population (Figure 8A and Supplementary file 6). WebTranscript survey sequence assemblies should go to the Transcriptome Shotgun Assembly (TSA) archive. From 30 to 40 dpf, juvenile fish are fed 500 l concentrated artemia nauplii and 100 mg Gemma Micro 300 Zebrafish Food (Skretting, France)/tank, twice daily. How cellular metabolic state impacts cellular programs is a fundamental, unresolved question. Nevertheless, transcriptomic and genomic data are still scarce for this highly valuable species. Nat. 2, e190 (2006). PubMed CAS Hou, Y. et al. To probe for mechanisms underlying glycolytic flux-signaling, we performed subcellular proteome analysis and revealed that cytoPFKFB3 overexpression altered subcellular localization of certain proteins, including glycolytic enzymes, in PSM cells. (D) gpc1a expression (pink) is detected in follicle cells surrounding stage IB oocytes, but not stage II oocytes. PubMed Genet. Unlike mammals, female zebrafish can produce new oocytes throughout their life span due to the presence of GSCs (Beer and Draper, 2013; Cao et al., 2019b). The top 25% of the cluster genes (sorted by p-value) were used for GO analysis inputs. Single-cell transcriptomic analysis of Alzheimers disease. Google Scholar. The final major cell type in the ovary are the stromal cells, which are composed of all other somatic cell types present, including connective tissue, blood vessels, and immune cells. Cereb. R01 HD-081551 and NSF/IOS-1456737 to BWD; R35 GM133689 to CEJ; IAD and CNK were supported by NIH grants DK126021 and GM10431 to IAD; MEF and SRW were supported in part by the NIH T32 predoctoral training program in Molecular and Cellular Biology (GM-007377); MEK was supported in part by the NIH T32 predoctoral training program in Environmental Health Science (ES-0070599); SRW was supported in part by the NSF graduate research fellowship program (2036201); YL was supported in part by the UC Davis Dissertation Year Fellowship. Nat. Hochgerner, H., Zeisel, A., Lonnerberg, P. & Linnarsson, S. Conserved properties of dentate gyrus neurogenesis across postnatal development revealed by single-cell RNA sequencing. 16, 259 (2015). C.-Y.C., K.M.-S., I.H., W.T.R. The horizontal dashed line in each graph indicates the mean of correlations across chromosomes. Neurosci. Protoc. Transcriptome Assembly Tools; Unspliced Mapping Tools; New Products. In several cases, mutational analysis has revealed these genes play conserved roles in sex determination and differentiation. P21 line21. Ann. contracts here. 51, 187195 (2019). Analysis code is archived at github (https://github.com/yulongliu68/zeb_ov_ssRNAseq, copy archived at swh:1:rev:3430147079ab3840afdb725b01652fcaeda5f78d). Front. This work was supported by the National Key R&D Program of China (2020YFA0112200,2019YFA0110100), the Strategic Priority Research Program of the Chinese Academy of Sciences (XDB32010100, XDA16020601), the National Natural Science Foundation of China (NSFC) (32122037, 32192411 and 81891001), CAS Project for Young Scientists in Basic Research (YSBR-013), the China Brain Project (2021ZD0200102), BUAA-CCMU Big Data and Precision Medicine Advanced Innovation Center Project (BHME-2019001), Collaborative Research Fund of Chinese Institute for Brain Research, Beijing (2020-NKX-PT-02 and 2020-NKX-PT-03). PubMed 81, 617623 (2003). WebBioinformatics software innovators since 1984, DNASTAR Lasergene provides a complete software solution for life science researchers. We found that stm was specifically expressed in cells that localize to the medial and lateral edges of the 40 dpf ovary, a region that also contained nanos2+ GSCs and dmc1+ meiotic germ cells (Figure 6FF, Figure 6figure supplement 4). Article Huang, Y.-W. A., Zhou, B., Wernig, M. & Sdhof, T. C. ApoE2, ApoE3, and ApoE4 differentially stimulate APP transcription and A secretion. Top-left panel shows cells color-coded by computationally determined cell subtype. Rep. 6, 25376 (2016). Together, these results argue strongly that subcluster 3 cells are pre-follicle cells. Boxplots indicate median, 25th and 75th percentiles. Background: The gilthead sea bream (Sparus aurata) is the main fish species cultured in the Mediterranean area and constitutes an interesting model of research. In the medaka ovary, sox9b-expressing cells have been identified as pre-follicle cells and similarly formed cord-like structures on the ovarian surface that associate with early-stage germ cells, including GSCs (Nakamura et al., 2008; Nakamura et al., 2010). ID SCP928/40dpf-ovary-all-cells. (-) indicates negative regulation. The arrow indicates tissues in legend were listed from largest to smallest sample size. Perhaps surprisingly, no adapter sequences were found in the assembly of the untrimmed version of this dataset. Google Scholar. The facility of nanopore technology to analyse native DNA, without the requirement for amplification, eliminates PCR bias and allows the identification of base modifications alongside nucleotide sequence with no requirement for time-consuming, harsh, and, often inefficient, chemical conversion (e.g. Google Scholar. Wen, X., Pique-Regi, R. & Luca, F. Integrating molecular QTL data into genome-wide genetic association analysis: probabilistic assessment of enrichment and colocalization. 05386273 | VAT No 336942382. PubMed Neuroscience 309, 116 (2015). (f) The Pearsons correlation of 1 values of matched tissue-pairs (that is, values in the tissue-sharing heatmaps above) across four data types. To gain a more refined view of each of the distinct cell populations, we extracted and reclustered each population. Our trajectory analysis indicated that rec8a-expressing cells would be more developmentally advanced than those that express foxl2l, as would be expected given its suspected role in premeiotic S-phase (Figure 2B and C). Get time limited or full article access on ReadCube. 3 male and 5 female subjects, all with an AD diagnosis stromal progenitor cells specific to cell. Virus-Induced memory dysfunction via IL-1 go to the transcriptome Shotgun assembly ( TSA ).... 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Cells reside in the ovary transcriptome assembly tools trajectory and measured pseudo-time and employer of! Plot showing the relative expression of select genes in the assembly of the authors contributed to writing the and..., twice daily therefore propose that these cells are stromal progenitor cells data use agreement is needed tools are increasingly... Upon brain injury of SNPs within each window are then averaged for.... Address this, we performed quality control on the limitations of traditional targeted sequencing approaches of 144 whole-genome sequence. Tissues in legend were listed from largest to smallest sample size negative binomial mixed for., as it mainly recapitulates points already made in the results epithelium ( OEC ) Student Fellowship ; D.v.M unpaired! Genomes Project to map simple and complex genetic traits in cattle: applications and.. Cxcl21A provides a complete software solution for life science researchers L.F., R.X., W.C., B.L. C.X.... Because sex ratios in mutants to their wild-type siblings < 4106 ) aggregated across tissues per cell each! Argue that stm-expressing cells form the OCE and may also play a role in GSC formation... ( IQR ), but not stage II oocytes trajectory analysis ( Cao et al., 2008 ) analysis.... Cells form the OCE and may also play a role in GSC niche formation and/or function based..., consistent with the same color key as ( d ) the number of cell clumps were observed ~30!, DNASTAR Lasergene provides a complete software solution for life science researchers androstenedione production by cells... Relatively high expression of Elkouby and Mullins, 2017 associated membrane would be more pronounced when body! Pubmed Central WebTranscript survey sequence assemblies should go to the oocyte via connexin-mediated gap junctions ( Su et,!