Project description:We identified a lineage-specific GATA-like gene named fuchi nashi (fuchi) as a gene required for establishing a germ disc in the early embryo of the common house spider Parasteatoda tepidariorum. To investigate possible involvement of the early fuchi activity in regulation of chromatin accessibility, we used an assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq). Whole embryos at stage 3 that were untreated (wild-type) or treated for fuchi or hedgehog (hh) parental RNAi (pRNAi), as well as wild-type whole embryos at stages 1 and 2, were analyzed by ATAC-seq. We found that chromatin accesibilities were specifically altered by fuchi pRNAi at many regions of the genome, whereas chromatin accessibilities were little affected by Pt-hh pRNAi, which served as negative control.
Project description:Hedgehog signaling plays pivotal roles in formation of the embryonic anterior-posterior axis in the spider Parasteatoda tepidariorum. In this work, using the combination of RNA-sequencing and parental RNA interference (pRNAi), we genome-widely identified genes that transcriptionally responded to altered states of Hedgehog signaling in spider embryos, which were caused by Pt-hedgehog (Pt-hh) pRNAi and Pt-patched (Pt-ptc) pRNAi. The identified genes were classified into four classes (Classes I to IV) based on the positive/negative responses to the Pt-hh pRNAi and Pt-ptc pRNAi states. The Class II genes were negatively regulated by Hh signaling (positive response to Pt-hh pRNAi and negative response to Pt-ptc pRNAi). Among the Class II genes, we identified Pt-msx1 as a key segmentation gene in the spider embryo. Moreover, we conducted a transcriptomic analysis of Pt-msx1 pRNAi embryos and identified additional genes whose expression showed patterns associated with segmentation.
Project description:The common house spider Parasteatoda tepidariorum is a chelicerate model organism for studying developmental mechanisms and their evolution in arthropods. In contrast to the well-studied model insect, Drosophila melanogaster, embryos of the spider undergo patterning in a cellular environment from early stages (at least after the number of the nuclei increase to 16). Use of spider embryos provide new opportunities to understand the evolution of developmental mechanisms underlying arthropod body plans. This analysis aims to generate genome-scale, developmental profiles of gene expression in embryos of the spider P. tepidariorum, which facilitate a wide range of studies using this spider species.
Project description:We identified a GATA-like gene named fuchi nashi (fuchi) as a gene required for establishing a germ disc in the early embryo of the spider Parasteatoda tepidariorum. fuchi transcript is initially expressed in cells at both polar regions of the embryo and later in endodermal and extraembryonic cells. To genome-widely identify genes whose expressions are regulated by fuchi, we conducted RNA-sequencing of whole embryos at three different early stages (stages 2 and 3 and early stage 5) that were untreated (control) or treated for fuchi parental RNA interference (pRNAi).
Project description:The early embryo of the spider Parasteatoda tepidariorum initially shows spherical symmetry in morphology. Following a symmetry breaking event, it forms a radially symmetric germ disc in the hemisphere of the egg. In this work, we conducted RNA-sequencing of cells isolated from small different regions of radially symmetric forming/formed germ discs, aming to identify candidate genes whose transcripts are locally expressed in early spider embryos.
Project description:In the spider Achaearanea tepidariorum, Hedgehog (Hh) signaling plays a key role in the formation of the two major embryonic axes. Analyses of expression patterns of the spider hh homolog, At-hh, suggested that Hh signaling might be involved in the subsequent segmentation process also. In this microarray experiment, we attempted to identify candidate genes whose expressions are regulated by Hh signaling during early phases of spider segmentation. The objective of this experiment was a screen for genes whose expressions might be affected by parental RNA interference (pRNAi) against At-hh in spider embryos. Oligonucleotide probes were designed based on accumulated A. tepidariorum EST sequences and embedded in custom microarrays (12K x2). Total RNA was extracted from late stage 5 embryos derived from a mated female before and after injection of dsRNA targeted for At-hh. Gene expression levels were compared between the untreated (normal) and the At-hh pRNAi-treated samples. Probes for At-hh and a homolog of Drosophila orthodenticle, At-otd, served as positive controls to validate this experiment.
Project description:In a cellular field of the early spider embryo, Hedgehog signaling operates to specify a “fuzzy” French-flag-like pattern along the primary axis. We applied single-cell and single-nucleus RNA sequencing to the early spider embryo. We confirmed that these techniques successfully detected three cell population corresponding to germ layers and some known cell types. We showed that the data had sufficient information for reconstruction of a correct global polarity of the presumptive ectoderm.
