Project description:This dataset contains several sponge-derived bacteria, along with data from the sponges to enable the assignation of the biosynthetic source of detected metabolite features. Media controls and solvent blanks are also included. Please see metadata for the bacterial source sponge.
Project description:Pseudovibrio a-Proteobacteria have been repeatedly isolated from marine sponges and proposed to be beneficial to the host. Bacterial motility is known to contribute to host colonization. We have previously identified pseudovibriamides A and B, produced in culture by Pseudovibrio brasiliensis Ab134, and shown that pseudovibriamide A promotes flagellar motility. Pseudovibriamides are encoded in a hybrid nonribosomal peptide synthetase-polyketide synthase gene cluster that also includes several accessory genes. Pseudovibriamide A is a linear heptapeptide and pseudovibriamide B is a nonadepsipeptide derived from pseudovibriamide A. Here we define the borders of the pseudovibriamides gene cluster, assign function to biosynthetic genes using reverse genetics and test the hypothesis that pseudovibriamides impact motility by modulating gene transcription. RNA-seq transcriptomic analyses of strains having different compositions of pseudovibriamides suggested that both pseudovibriamides A and B affect genes potentially involved in motility, and that a compensatory mechanism is at play in mutants that produce only pseudovibriamide A, resulting in comparable swarming motility as the wild type. The data gathered suggest that pseudovibriamides A and B have opposite roles in modulating a subset of genes, with pseudovibriamide B having a primary effect in gene activation, and pseudovibriamide A on inhibition. Finally, we observed many differentially expressed genes (up to 29% of the total gene number) indicating that pseudovibriamides have a global effect on transcription that goes beyond motility.
Project description:In order to compare sponge and eumetazoan (higher animal) body plans, we identified and studied expression of a broad range of eumetazoan developmental regulatory genes in Sycon ciliatum (Calcispongiae). In this species, embryonic development is semi-synchronous within a population, synchronous within individuals, and oocytes and embryos occupy a significant fraction of the volume of the sponges during the reproductive period. RNASeq libraries representing non-reproductive (somatic) tissue slices along the body axis, as well as oocytes, embryos and free swimming larvae were generated from material obtained by sampling throughout the life cycle.
Project description:MicroRNA (miRNA) sponges containing miRNA complementary binding sites constitute a potentially useful strategy for miRNA-inhibition therapeutics in cancer patients. Recently, naturally occurring circular RNAs (circRNAs) have been revealed to function as efficient microRNA sponges. We hypothesized that synthetic circRNA sponges targeting oncomiRs could be constructed and used to achieve potentially therapeutic microRNA loss of function. In this study, linear RNA molecules containing five miR-21 binding sites were transcribed in vitro. After dephosphorylation by calf intestinal phosphatase and phosphorylation by T4 polynucleotide kinase, circRNA sponges were circularized using 5’-3’ end ligation by T4 RNA ligase 1. Synthetic circular sponge stability was assayed in the presence of RNase R or fetal bovine serum. Luciferase reporter and cell proliferation assays were performed to assess competitive inhibition of miR-21 activity by circRNA sponges in NCI-N87 gastric cancer cells. Tandem Mass Tag (TMT) labeling proteomics analysis and Western blotting were performed to delineate effects of circRNA sponges on miR-21 downstream targeted proteins. Our experiments revealed that artificial circRNA sponges can be synthesized using enzymatic ligation. These synthetic circRNA sponges are more resistant than their linear RNA counterparts to nuclease degradation in vitro. They effectively suppress the activity of miR-21 on its downstream protein targets, including the important cancer protein DAXX. Finally, they also inhibit gastric cancer cell proliferation. Our results suggest that synthetic circRNA sponges represent a rapid, effective, convenient strategy to achieve loss of miRNA function in vitro, with potential future therapeutic application in vivo.