Project description:RNA interference (RNAi) is a phylogenetically widespread gene silencing process triggered by doublestranded RNA (dsRNA). In plants and C. elegans, two distinct populations of small RNAs have been proposed to participate in RNAi : "Primary siRNAs" (derived from Dicer nuclease-mediated cleavage of the original trigger) and "Secondary siRNAs" (additional small RNAs whose synthesis requires an RNA-directed RNA polymerase [RdRP]). Analyzing small RNAs associated with ongoing RNAi in C. elegans, we found secondary siRNAs to comprise the vast majority. The bulk of secondary siRNAs exhibited structure and sequence indicative of a biosynthetic mode where each molecule derives from an independent de novo initiation by RdRP. Analysis of endogenous small RNAs indicated that a fraction derive from a biosynthetic mechanism that is similar to that of secondary siRNAs formed during RNAi, suggesting that small antisense transcripts derived from cellular mRNAs by RdRP activity may have key roles in cellular regulation. Keywords: C. elegans small RNA sequences from wild type animals fed on sel-1 dsRNA producing bacteria C. elegans small RNA sequences from wild type animals fed on sel-1 dsRNA producing bacteria
Project description:RNA interference (RNAi) is a phylogenetically widespread gene silencing process triggered by doublestranded RNA (dsRNA). In plants and C. elegans, two distinct populations of small RNAs have been proposed to participate in RNAi : "Primary siRNAs" (derived from Dicer nuclease-mediated cleavage of the original trigger) and "Secondary siRNAs" (additional small RNAs whose synthesis requires an RNA-directed RNA polymerase [RdRP]). Analyzing small RNAs associated with ongoing RNAi in C. elegans, we found secondary siRNAs to comprise the vast majority. The bulk of secondary siRNAs exhibited structure and sequence indicative of a biosynthetic mode where each molecule derives from an independent de novo initiation by RdRP. Analysis of endogenous small RNAs indicated that a fraction derive from a biosynthetic mechanism that is similar to that of secondary siRNAs formed during RNAi, suggesting that small antisense transcripts derived from cellular mRNAs by RdRP activity may have key roles in cellular regulation. Keywords: C. elegans small RNA sequences from wild type animals fed on sel-1 dsRNA producing bacteria
Project description:Indole is an intercellular and interkingdom signaling molecule, which is widespread in diverse ecological niches. Caenorhabditis elegans is a bacterivorous nematode living in soil and compost environments and a useful model host for the study of host-microbe interactions. While various bacteria and some plants produce a large quantity of extracellular indole, little is known about the effects of indole, its derivatives, and indole-producing bacteria on behaviors in C. elegans and animals. Here, we show that C. elegans senses and moves toward indole and indole-producing bacteria, such as Escherichia coli, Shigella boydii, Providencia stuartii, and Klebsiella oxytoca, while avoids non-indole producing pathogenic bacteria. It was also found that indole-producing bacteria and non-indole-producing bacteria exert divergent effects on egg-laying behavior of C. elegans via indole. In addition, various indole derivatives also modulate chemotaxis, egg-laying behavior, and survival of C. elegans. In contrast, indole at a high concentration to kill C. elegans that has the ability to detoxify indole via oxidation and glucosylation, indicating predator-prey interactions via a double-edged molecule indole. Transcriptional analysis showed that indole markedly up-regulated gene expression of cytochrome P450 family, UDP-glucuronosyltransferase, glutathione S-transferase, which explained well the modification of indole in C. elegans, while down-regulated expression of collagen genes and F-box genes. Our findings suggest that indole and its derivatives are important interkingdom signaling molecules in bacteria-nematode interactions.
Project description:Expression data from Caenorhabditis elegans let-418(RNAi), mep-1(RNAi) and gfp(RNAi) L1 larvae. The C. elegans genome encodes two homologs of the human protein Mi-2, namely LET-418 and CHD-3. LET-418 plays an essential role during development; its depletion leads to a pleiotropic and lethal phenotype that includes larval arrest, an everted vulva and sterility. Without maternal contribution, let-418 mutants stop their development at the L1 larval stage (von Zelewsky et al., 2000). We further characterized this arrest and showed that it is very similar to the L1 diapause induced by starvation; both germline and somatic cells remain in a quiescent state in let-418 L1 arrested larvae, indicating that LET-418 activity is required to bypass the L1 arrest in presence of food. The let-418 L1 larvae express ectopically the P granule component PGL-1 in somatic cells (Unhavaithaya et al., 2002). Interestingly, the phenotype of mep-1 mutants is remarkably similar to that of let-418: RNAi targeting mep-1 also induced an L1 arrest phenotype; furthermore, MEP-1 and LET-418 have been shown to physically interact (Unhavaithaya et al., 2002 and M. Passannante). The null allele mep-1(q660) is temperature sensitive and shows a more severe phenotype at higher temperatures. At 20°C, about 10% of mep-1 homozygotes derived from heterozygous mothers arrest as young larvae, whereas the remaining 90% develop into sterile adults (Belfiore et al., 2002). Later in development, the somatic gonad is affected in mep-1(q660) mutants. This results in an abnormal and disorganized gonad, a phenotype also observed in let-418(s1617) mutants. Both let-418 and mep-1 mutants produce a very limited number of oocytes and have pseudovulvae derived from P8.p (Belfiore et al., 2002; von Zelewsky et al., 2000 and C. Wicky, personal communication). Preliminary quantitative real-time PCR revealed that the expression of genes coding for P granule components was deregulated in both mep-1(RNAi) and let-418(RNAi) L1 larvae (data not shown). To further investigate this issue, we performed a complete gene expression analysis. Given the fact that mep-1(q660) mutants are sterile, we used RNA interference to generate mep-1 depleted worms. Bacteria expressing gfp dsRNA (pPE128.110 in HT115) were used as reference, since RNA interference may induce gene expression changes by itself. C. elegans L1 larvae treated with RNA interference were selected for RNA extraction and hybridization on Affymetrix microarrays. Synchronized wild type L4 animals were grown at 25° on bacteria expressing either gfp, let-418 or mep-1 dsRNA. Eggs were collected by bleaching gravid adults and allowed to hatch in the absence of food at 25°C. Newly hatched L1 larvae were fed on bacteria expressing the different dsRNA for three hours to recover from starvation. Three replicates per RNAi.
Project description:Transcriptional profiling of Caenorhabditis elegans comparing control E. coli OP50-fed C. elegans with L. sphaericus-fed C. elegans
Project description:Transcriptional profiling of Caenorhabditis elegans comparing control E. coli OP50-fed C. elegans with L. sphaericus-fed C. elegans Two-condition experiment, E. coli OP50-fed C. elegans vs. L. sphaericus-fed C. elegans