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RNA Seq analysis of sorted Schmidtea mediterranea X1 and Xins cells to identify factors specific to neoblasts

ABSTRACT: Adult stem cells are tissue-specific cells with the capacity to self-renew and differentiate to continually replace cells lost to normal physiological turnover or injury. Neoblasts, the planarian stem cells, are widely distributed throughout the body mesenchyme, driving constitutive renewal of tissues during homeostasis and endowing planarians with the remarkable capacity to regenerate wholly from tiny tissue fragments. Neoblasts are the only dividing cells in planarians and are believed to be collectively comprised of both a heterogeneous population of pluripotent cells with broad differentiation potential and also lineage-committed progenitor cells that give rise to specific tissues. Recent technology has allowed one to isolate stem cells so we used a well-established method to isolate planarian stem cells by Hoechst blue staining and flow cytometry. To understand the molecular mechanisms underlying neoblast differentiation, we performed an RNA-Seq analysis of X1 and Xins cells looking at the differentially expressed genes between the two populations. Examine gene expression profiles of adult flatwormâ??s X1 and Xins cell types. The experiment was performed in quadruplicate yielding a total of 8 samples.

ORGANISM(S): Schmidtea mediterranea

SUBMITTER: Alex Chitsazan

PROVIDER: E-GEOD-73027 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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RNA Seq analysis of sorted Schmidtea mediterranea X1 and Xins cells to identify factors specific to neoblasts

Project description:Adult stem cells are tissue-specific cells with the capacity to self-renew and differentiate to continually replace cells lost to normal physiological turnover or injury. Neoblasts, the planarian stem cells, are widely distributed throughout the body mesenchyme, driving constitutive renewal of tissues during homeostasis and endowing planarians with the remarkable capacity to regenerate wholly from tiny tissue fragments. Neoblasts are the only dividing cells in planarians and are believed to be collectively comprised of both a heterogeneous population of pluripotent cells with broad differentiation potential and also lineage-committed progenitor cells that give rise to specific tissues. Recent technology has allowed one to isolate stem cells so we used a well-established method to isolate planarian stem cells by Hoechst blue staining and flow cytometry. To understand the molecular mechanisms underlying neoblast differentiation, we performed an RNA-Seq analysis of X1 and Xins cells looking at the differentially expressed genes between the two populations.

2015-12-17 | GSE73027 | GEO

Transcriptome analysis reveals strain-specific and conserved stemness genes in Schmidtea mediterranea

Project description:The transcriptome of the planarian Schmidtea mediterranea is not well characterized. We have used RNA-Seq to characterize the transcriptome in both sexual and asexual strains of S. mediterranea from both untreated and irradiated animals. Moreover, we have performed RNA-Seq on RNA purified from FACS sorted neoblasts and differentiated cells. Together these studies expand our understanding of the planarian transcriptome and have identified strain-specific, neoblast-specific, and conserved transcripts. RNA-Seq was performed on RNA isolated from untreated and irradiated S. mediterranea animals from both the sexual and asexual strains, on FACS purified X1 neoblasts, X2 neoblasts, and Xins differentiated cells. One of the raw data files for GSM847465 is missing. The fasta file is provided at http://genome.vcell.uchc.edu/GenomeData02/Graveley_Lab_Public_Data/Planarian/S.mediterranea_SexNIRmRNA3.fa.gz

2012-04-24 | E-GEOD-34326 | biostudies-arrayexpress

H3K36me3 is significantly more enriched at Set1-affected loci than at MLL1/2-affected loci in planarian stem cells

Project description:Histone H3 lysine 4 trimethylation (H3K4me3) is known to correlate with both active and poised genomic loci, yet many questions remain regarding its functional roles in vivo. We identify functional genomic targets of two H3K4 methyltransferases, Set1 and MLL1/2, in both the stem cells (X1) and differentiated tissue (Xins) of the planarian flatworm Schmidtea mediterranea. We show that, despite their common substrate, these enzymes target distinct genomic loci in vivo, which are distinguishable by the footprint each enzyme leaves on the chromatin template, i.e., the breadth of the H3K4me3 peak. Whereas Set1 targets are broadly associated with the maintenance of the stem cell population, MLL1/2 targets are specifically enriched for genes involved in ciliogenesis. These data not only confirm that chromatin regulation is fundamental to planarian stem cell function, but also provide evidence for post-embryonic functional specificity of H3K4me3 methyltransferases in vivo. To test this, we looked at the H3K4me3 chromatin profile in WW, X1 and Xins cell types upon knockdown of set1, mll1/2, as well as WT. Examine binding of H3K36me3 in WT in Planarian. The experiment performed yielded a total of 2 samples. These samples validate the main ChIP samples.

2015-12-17 | E-GEOD-74419 | biostudies-arrayexpress

Single-cell analysis reveals prominent classes within the planarian stem cell compartment

Project description:Single cell qPCR analysis identified two prominent expression signatures within the planarian stem cells, the neoblasts. RNAi against zinc finger protein zfp-1 results in elimination of one of the two classes. Worms were treated with RNAi by feeding. X1 or X2 cells were isolated by FACS, or blastemas were dissected. mRNA libraries were sequenced.

2014-09-30 | E-GEOD-58057 | biostudies-arrayexpress

454 Life Sciences/Roche and Illumina sequencing of small RNAs from planarian samples

Project description:Freshwater planarian flatworms possess uncanny regenerative capacities mediated by abundant and collectively totipotent adult stem cells. Key functions of these cells during regeneration and tissue homeostasis have been shown to depend on PIWI, a molecule required for piRNA expression in planarians. Nevertheless, the full complement of piRNAs, and microRNAs (miRNAs) in this organism has yet to be defined. Here, we report on the large scale cloning and sequencing of small RNAs from the planarian Schmidtea mediterranea, yielding altogether millions of sequenced unique small RNAs. We show that piRNAs are in part organized in genomic clusters and that they share characteristic features with mammalian and fly piRNAs. We further identify 61 novel miRNA genes and thus double the number of known planarian miRNAs. Sequencing, as well as quantitative PCR of small RNAs uncovered ten miRNAs enriched in planarian stem cells. These miRNAs are down-regulated in animals in which stem cells have been abrogated by irradiation, and thus constitute miRNAs likely associated with specific stem cell functions. Altogether, we present the first comprehensive small RNA analysis in animals belonging to the third animal superphylum, the Lophotrochozoa, and single out a number of miRNAs that may function in regeneration. Several of these miRNAs are deeply conserved in animals. Small RNAs were sequenced from FACS-sorted neoblast stem cells, untreated planarians and irradiated planarians, depleted of neoblasts

2009-06-30 | E-GEOD-16159 | biostudies-arrayexpress

Set1 and MLL1/2 target distinct sets of functionally different genomic loci in vivo: An in depth ChIP-seq analysis of H3K4me3 targets in vivo and upon Set1 and mll1 knockdown

Project description:Histone H3 lysine 4 trimethylation (H3K4me3) is known to correlate with both active and poised genomic loci, yet many questions remain regarding its functional roles in vivo. We identify functional genomic targets of two H3K4 methyltransferases, Set1 and MLL1/2, in both the stem cells (X1) and differentiated tissue (Xins) of the planarian flatworm Schmidtea mediterranea. We show that, despite their common substrate, these enzymes target distinct genomic loci in vivo, which are distinguishable by the footprint each enzyme leaves on the chromatin template, i.e., the breadth of the H3K4me3 peak. Whereas Set1 targets are broadly associated with the maintenance of the stem cell population, MLL1/2 targets are specifically enriched for genes involved in ciliogenesis. These data not only confirm that chromatin regulation is fundamental to planarian stem cell function, but also provide evidence for post-embryonic functional specificity of H3K4me3 methyltransferases in vivo. To test this, we looked at the H3K4me3 chromatin profile in WW, X1 and Xins cell types upon knockdown of set1, mll1/2, as well as WT. Examine differential binding of H3K4me3 in different experimental conditions. The experiment performed yielded a total of 58 samples.

2015-12-17 | E-GEOD-74153 | biostudies-arrayexpress

Conserved functional antagonism between CELF and MBNL proteins regulates stem cell-specific alternative splicing and regeneration in planarians (dataset 2)

Project description:Here, we analyze the RNA-binding preferences of the planarian smed-mbnl-like-2, smed-mbnl-1 (isoform X1), smed-bruli, smed-mbnl-1 (isoform Xins), and smed-mbnl-like-1 protein using RNAcompete.

2016-08-01 | GSE75554 | GEO

Set1 and MLL1/2 target distinct sets of functionally different genomic loci in vivo: An in depth ChIP-seq analysis of H3K4me3 targets in vivo and upon Set1 and mll1 knockdown

2015-12-17 | GSE74153 | GEO

H3K36me3 is significantly more enriched at Set1-affected loci than at MLL1/2-affected loci in planarian stem cells

2015-12-17 | GSE74419 | GEO

Canonical and early lineage-specific stem cell types identified in planarian SirNeoblasts

Project description:The pluripotent stem cells in planarians, a model for tissue and cellular regeneration, have yet to be definitively identified. We recently developed a method to enrich piwi-1+ cells in Schmidtea mediterranea, by staining cells with SiR-DNA and Cell Tracker Green, named SirNeoblasts that permits their propagation and subsequent functional study in vivo. Since traditional enrichment for planarian neoblasts by Hoechst 33342 staining, to generate X1 cells, blocks the cell cycle, inducing cytotoxicity, this method represents a substantive technological advance for functional investigation of cell fate and regeneration in basic and applied research questions. However, the similarities in heterogeneity of cell subtypes between SirNeoblasts and X1 remain unknown. In this work, we performed single cell RNA sequencing for SirNeoblasts for comparison with differential expression patterns in a publicly available X1 single cell RNA sequencing data. We found first that all of the lineage-specific progenitor cells in X1 were present in comparable proportions in SirNeoblasts. In addition, SirNeoblasts contain an early muscle progenitor unreported in X1. Analysis of new markers for putative pluripotent stem cells identified here, with subsequent sub-clustering analysis revealed earlier lineages of epidermal, muscular, intestinal, and pharyngeal progenitors than have been observed in X1. Using the ski-3 and GCM markers, we also identified a pluripotent cell population at higher resolution than that provided by tgs-1. The use of SirNeoblast will enable broad experimental advances in regeneration and cell fate specification, given the possibility for propagation and transplantation of recombinant and mutagenized pluripotent stem cells not previously afforded to this rapid and versatile model system.

2021-03-23 | GSE158706 | GEO

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