Unknown,Transcriptomics,Genomics,Proteomics

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Expression data from Caenorhabditis elegans exposed to single-walled carbon nanotubes

ABSTRACT: The potential environmental risk of single-walled carbon nanotubes (SWCNTs) is evaluated using Caenorhabditis elegans (C. elegans) as an ecotoxicological animal model. Highly soluble amide-modified SWCNTs (a-SWCNTs) are used in the present study so that the dose-response impact of SWCNTs could be studied. mechanisms. a-SWCNTs are efficiently taken up by worms during feeding and cause significant toxicity in worms, including retarded growth, shortened lifespan and defective embryogenesis. Genome-wide gene expression analysis is performed to investigate the toxic molecular We examined the effect of different concentrations of a-SWCNTs (0, 100, 250 and 500 Î¼g/mL) on the growth of C. elegans. We measured the body length of worms reaching the L4 stage after a-SWCNT exposure for 48 hr at 22Â°C. Compared to the untreated worms, we found that the average length of worms exposed to a-SWCNTs (500 Î¼g mL-1) was significantly shorter than the untreated groups. In addition, the dose of a-SWCNTs also caused retarded growth, reduced lifespan and defective embryogenesis in worms. Genome-wide gene expression analysis using an Affymetrix GeneChip was performed to further investigate the molecular basis of these defects.

ORGANISM(S): Caenorhabditis elegans

SUBMITTER: Cheng Chou

PROVIDER: E-GEOD-41486 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Animal mRNAs are regulated by hundreds of RNA binding proteins (RBPs). The identification of RBP targets is crucial for understanding their function. A recent method, PAR-CLIP, uses photoreactive nucleosides to crosslink RBPs to target RNAs in cells prior to immunoprecipitation. Here, we establish iPAR-CLIP (in vivo PAR-CLIP) to determine, at nucleotide resolution, transcriptome-wide target sites of GLD-1, a conserved, germline-specific translational repressor in C. elegans. We identified 439 reproducible targets and demonstrate an excellent dynamic range of target detection by iPAR-CLIP. Upon GLD-1 knock-down, protein but not mRNA expression of the 439 targets was specifically and highly significantly upregulated, demonstrating functionality. Finally, we discovered strongly conserved GLD-1 binding sites nearby the start codon of target genes. We propose that GLD-1 interacts with the translation machinery nearby the start codon, a so far unknown mode of gene regulation in eukaryotes. Arrested L1 worms were grown in liquid culture supplemented with 2mM 4SU or 6SG. 250,000 worms were sufficient for one iPAR-CLIP experiment. Living adult worms were transferred to NGM plates and crosslinked on ice using a Stratalinker (Stratagene) with customized 365nm UV-lamps (energy setting: 2J/cm2). Worms were lysed on ice by douncing in NP40 lysis buffer (50 mM HEPES-K pH 7.5, 150 mM KCl, 2 mM EDTA, 0.5% (v/v) NP-40, 0.5 mM DTT, protease inhibitor cocktail (Roche)). Cleared lysates were treated with RNase T1 (Fermentas) (final concentration 1U/?l) for 15 min at 22ÂºC. GLD-1::GFP::FLAG fusion proteins were immunoprecipitated for 1h at 4ÂºC using anti-FLAG antibody (Sigma, F3165) coupled to Protein G magnetic beads (Invitrogen). For one iPAR-CLIP experiment (1ml cleared lysate obtained from 250,000 worms), 300Âµl beads and 150Âµg antibody were used. Immunoprecipitates were treated with RNase T1 (100U/?l) for exactly 12 min at 22 ÂºC. Subsequently, PAR-CLIP was carried out as described previously (Hafner et al, 2010). cDNA libraries were sequenced on a Genome Analyzer II (Illumina).

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Expression data from Caenorhabditis elegans exposed to single-walled carbon nanotubes

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