Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Identification of genes whose expressions are affected by hedgehog RNAi in spider embryos

ABSTRACT: 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.

ORGANISM(S): Parasteatoda tepidariorum

SUBMITTER: Hiroki Oda

PROVIDER: E-GEOD-26468 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Travelling and splitting of a wave of hedgehog expression involved in spider-head segmentation.

Kanayama Masaki M Akiyama-Oda Yasuko Y Nishimura Osamu O Tarui Hiroshi H Agata Kiyokazu K Oda Hiroki H

Nature communications 20111011

During development segmentation is a process that generates a spatial periodic pattern. Peak splitting of waves of gene expression is a mathematically predicted, simple strategy accounting for this type of process, but it has not been well characterized biologically. Here we show temporally repeated splitting of gene expression into stripes that is associated with head axis growth in the spider Achaearanea embryo. Preceding segmentation, a wave of hedgehog homologue gene expression is observed t ...[more]

PMID: 21988916

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Project description:Linker histones are involved in the formation of higher-order chromatin structure. Although linker histones have been implicated in the regulation of specific genes, it still remains unclear what their principal binding determinants are and how their repressive function in vitro can be reconciled with presumed broad binding in vivo. We generated a full genome, high resolution binding map of linker histone H1 in Drosophila Kc cells, using DamID. H1 binds at similar levels across much of the genome, both in classical euchromatin and heterochromatin. Strikingly, there are pronounced dips of low H1 occupancy around transcription start sites of active genes and at many distant cis-regulatory sites. H1 dips are not due to lack of nucleosomes. Rather, all regions with low binding of H1 show enrichment of the histone variant H3.3 which itself has been linked to high nucleosome turnover. Upon knockdown of H3.3, we find that H1 levels increase at sites previously not covered with H1 with a concomitant increase in nucleosome repeat length. These changes are independent of transcriptional changes. Our results show that the H3.3 protein counteracts association of H1 at genomic sites with high rates of histone turnover. This antagonism provides a mechanism to keep diverse genomic sites in an open chromatin conformation. For this study, we generated DamID profiles of histone H1 and RpII18 in Drosophila Kc167 cells. Additionally, we generated H1 profiles in cells treated with RNAi against white, H3.3B, or H3.3A and H3.3B. Nucleosome positioning profiles were generated in untreated cells and cells treated with RNAi against white, H3.3B, or H3.3A and H3.3B. Profiles of expression changes were generated for H3.3B RNAi and H3.3A and H3.3B RNAi. DamID experiments for H1 and RpII18 were performed in Drosophila cell cultures. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing. Formaldehyde-assisted isolation of regulatotry elements (FAIRE) was performed in Drosophila Kc167 cells. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing over non-crosslinked genomic DNA. Nucleosome positioning profiles were made by hybridizing mononucleosomal DNA over MNase digested purified genomic DNA on 380k NimbleGen arrays with 10 bp probe spacing. Expression profiles were made as H3.3 RNAi over white RNAi cohybridizations on spotted INDAC long oligo arrays. Every experiment was done in duplicate in the reverse dye orientation.

Dataset Information

Identification of genes whose expressions are affected by hedgehog RNAi in spider embryos

Publications

Travelling and splitting of a wave of hedgehog expression involved in spider-head segmentation.

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