Project description:Background: Freshwater planarians are well known for their regenerative abilities. Less well known is how planarians maintain spatial patterning in long-lived adult animals or how they re-pattern tissues during regeneration. HOX genes are good candidates to regulate planarian spatial patterning, yet the full complement or genomic clustering of planarian HOX genes has not yet been described, primarily because only a few have been detectable by in situ hybridization, and none have given morphological phenotypes when knocked down by RNAi. Results: Because the planarian Schmidtea mediterranea (S. med) is unsegmented, appendage-less, and morphologically simple, it has been proposed that it may have a simplified HOX gene complement. Here we argue against this hypothesis and show that S. med has a total of 13 HOX genes, which represent homologs to all major axial categories, and can be detected by whole-mount in situ hybridization using a highly-sensitive method. In addition, we show that planarian HOX genes do not cluster in the genome, yet 5/13 have retained aspects of axially-restricted expression. Finally, we confirm HOX gene axial expression by RNA-deep-sequencing 6 anterior-to-posterior “zones” of the animal, which we provide as a dataset to the community to discover other axially-restricted transcripts. Conclusions: Freshwater planarians have an unappreciated HOX gene complexity, with all major axial categories represented. However, we conclude based on adult expression patterns that planarians have a derived body plan and their asexual lifestyle may have allowed for large changes in HOX expression from the last common ancestor between arthropods, flatworms, and vertebrates. Using our in situ method and axial zone RNAseq data, it should be possible to further understand the pathways that pattern the anterior-posterior axis of adult planarians. Overall design: 10 animals were immobilized on peltier coolers and carefully amputated into 6 Anterior-Posterior zones, as illustrated in Figure 2 of the manuscript. RNA was extracted from each zone, and deep sequenced to a depth >52 million reads.
Project description:The freshwater planarian Schmidtea mediterranea is well known by its amazing regeneration capabilities thanks to the presence of adult stem cells, the neoblasts, the only proliferative cells and responsible for the differentiation in all the cell types of the organism. This study involves the creation of separated transcript libraries from both, isolated neoblasts and differentiated cells, as well as their posterior sequencing and quantification through Digital Gene Expression (DGE) for the characterization of the neoblast transcriptome. Three DGE libraries were produced from FACS isolated cell populations X1 (proliferating stem cells, S/G2/M), X2 (a mix of proliferating stem cells and stem cell progeny, G0/G1) and Xin (differentiated cells, G0/G1). Cells were isolated from a pool of 32 regenerating animals two days after being cut off pre- and post-pharingeally to trigger regeneration and neoblast proliferation.
Project description:The transcriptome of a cell dictates its unique cell-type biology. We used single-cell RNA sequencing to determine the transcriptomes for essentially every cell type of a complete animal: the regenerative planarian Schmidtea mediterranea. Planarians contain a diverse array of cell types, possess lineage progenitors for differentiated cells (including pluripotent stem cells), and constitutively express positional information, making them ideal for this undertaking. We generated data for 66,783 cells, defining transcriptomes for known and many previously unknown planarian cell types and for putative transition states between stem and differentiated cells. We also uncovered regionally expressed genes in muscle, which harbors positional information. Identifying the transcriptomes for potentially all cell types for many organisms should be readily attainable and is a powerful new approach to metazoan biology. Overall design: Single cell transcriptomes for the planarian Schmidtea mediterranea were generated using Drop-seq. Please note that each processed data file was generated from multiple samples, as indicated in the corresposponding sample description field.
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
Project description:Identification of differentially expressed genes in intestinal phagocytes, compared to non-intestinal cells in Schmidtea mediterranea. Total RNA was isolated from either intestinal phagocytes or non-intestinal cells (control). Four independent biological replicates were conducted with two dye swaps. Labeled targets were hybridized to two oligonucleotide arrays on which 11,521 planarian transcripts were represented.