Caenorhabditis elegans
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PROVIDER: PRJNA306416 | ENA |
REPOSITORIES: ENA
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Project description:RIP-chip analysis of the C. elegans PUF protein FBF
| PRJNA121991 | ENA
Project description:We determined the genomic landscape of FBF-1 and FBF-2 binding in germline stem cells using iCLIP, a method that allows identification of protein-RNA interactions at high resolution. We first developed reagents to explore the genomic binding landscapes of full-length FBF-1 and FBF-2 in vivo and then used our iCLIP data to test the precision of several commonly used methods for CLIP peak calling. Based on this iCLIP data, we discovered that FBF-1 and FBF-2 have similar global protein-RNA interaction profiles and that they both target conserved cell cycle regulators and lincRNAs. We found that FBF-1 and FBF-2 regulate their targets through canonical as well as unexpected motif sequences. We elucidated the first in vivo crosslink site analysis for a PUF protein from which we precisely determined FBF-1 and FBF-2 binding sites. Taken together, our data provide an updated model of PUF binding in stem cells. Our study also provides new insight on the control of gene expression in stem cells by RNA binding proteins.
2016-05-11 | GSE76136 | GEO
Project description:The molecular mechanisms of aging are unsolved and fascinating fundamental biological questions. Caenorhabditis elegans is an ideal model organism for investigating aging. PUF-8, a PUF (Pumilio and FBF) protein in C. elegans, is crucial for germline development through binding to the 3’ untranslated regions (3’ UTR) in the mRNA of target genes. Recently, PUF-8 was reported to alter mitochondrial dynamics and mitophagy by regulating MFF, a mitochondrial fission factor, and subsequently regulate longevity. Here, we determined the crystal structure of the PUF domain of PUF-8 with an RNA substrate. Mutagenesis experiments were performed to alter PUF-8 recognition of its target mRNAs. We generate these mutations in C. elegans, those mutations reduced the fertility and extended the lifespan. We deep sequenced total mRNAs from wild-type and puf-8 mutant worms and conducted in vitro RNA pull-down experiments. Six PUF-8 regulated genes were identified, in which their mRNA 3’ UTRs contain at least one PUF-binding element (PBE). One of the six genes, pqm-1, is crucial for lipid storage and aging process. Knockdown of pqm-1 could revert the lifespan extension of puf-8(-) animals. Therefore, PUF-8 may regulate the lifespan of C. elegans via modulating pqm-1-related pathways
2021-05-01 | GSE165061 | GEO
Project description:The PUF family of RNA binding proteins has a conserved role in maintaining stem cell self-renewal. FBF is a C. elegans PUF that is required to maintain germline stem cells (GSCs). To understand how FBF controls GSCs, we sought to identify is target mRNAs. Briefly, we immunoprecipitated FBF-mRNA complexes from worm extracts and then used microarrays to identify the FBF-associated mRNAs. To focus on germline targets of FBF, we used a FBF-GFP transgene under the control of a germline promoter and we used an anti-GFP antibody to purify FBF-GFP from worm extracts. In parallel, we also processed a strain expressing TUBULIN-GFP in the germline to control for mRNAs that non-specifically co-purify with GFP. We found that FBF associates with >1,000 unique mRNAs and likely controls a broad network of key cellular and developmental regulators.
2010-01-27 | GSE19922 | GEO
Project description:PUF RNA-binding proteins control stem cells in diverse species, including mammalian, arthropod, and nematode, in addition to other biological functions. The C. elegans PUF protein FBF serves as a paradigm for metazoan PUFs. FBF is essential for the maintenance of germline stem cells but also regulates the hermpahrodite sperm/oocyte cell fate switch and is critical for the process of spermatogenesis. We have attempted to “disentangle” the different roles of FBF by comparing its targets in spermatogenic and oogenic germlines. To this end, we used FBF iCLIP to learn its binding profile in an adult hermaphrodite germline that is sexually transformed and makes only sperm due to a temperature-sensitive sex-determination mutant. As a control, we analyzed FBF iCLIP data from oogenic germlines at the same temperature. Using a modified peak calling algorithm, we identified FBF binding sites in oogenic animals at 20°C, oogenic animals at 25°C, and spermatogenic animals at 25°C. Oogenic FBF targets were similar at 20°C and 25°C. By contrast, FBF mRNA targets in spermatogenetic animals had a distinct profile, revealing sperm-specific targets that are likely critical for the FBF role in spermatogenesis. Most importantly, we found FBF bound to mRNAs regardless of germline gender. In particular, a group of 22 mRNAs clustered as bound with high frequency in a gender- and temperature-independent manner. These 22 mRNAsencode RNA-binding proteins and stem cell regulators and may be crucial for the FBF role in in stem cell maintenance.
2018-03-30 | GSE83695 | GEO
Project description:Regulation of germline gene expression in C. elegans germline stem cells.
| PRJNA288637 | ENA
Project description:DCC binding and function (Expression Analysis)
| PRJNA114437 | ENA
Project description:DAF-12 is involved in development, dauer formation, and aging in Caenorhabditis elegans. To understand how DAF-12 is involved in these biologic processes, we performed chIP-chip using a DAF-12:TAP transgene and an anti-DAF-12 antibody. We identified 1175 genomic binding sites which were located within 5 kb of 3179 genes. These genes include known DAF-12 target genes as well as new genes involved in developmental timing, microRNA function, and stress resistance. The supplemental BED file contains all 1175 genomic binding sites described in our work.
2011-04-05 | GSE28350 | GEO
Project description:The PUF family of RNA binding proteins has a conserved role in maintaining stem cell self-renewal. FBF is a C. elegans PUF that is required to maintain germline stem cells (GSCs). To understand how FBF controls GSCs, we sought to identify is target mRNAs. Briefly, we immunoprecipitated FBF-mRNA complexes from worm extracts and then used microarrays to identify the FBF-associated mRNAs. To focus on germline targets of FBF, we used a FBF-GFP transgene under the control of a germline promoter and we used an anti-GFP antibody to purify FBF-GFP from worm extracts. In parallel, we also processed a strain expressing TUBULIN-GFP in the germline to control for mRNAs that non-specifically co-purify with GFP. We found that FBF associates with >1,000 unique mRNAs and likely controls a broad network of key cellular and developmental regulators. Experiment Overall Design: Worm extracts were prepared from synchronized adult C. elegans (24 h after L4 stage) expressing either a rescuing FBF-1-GFP or TUB-GFP transgene under the control of a germline promoter (pie-1). An immoblized anti-GFP antibody was then used to purify the GFP fusion proteins from extracts. RNA was then extracted from the pellets and analyzed on Affymetrix microarrays. Four biological replicates were performed, each consisting of a FBF-GFP and a TUB-GFP sample processed in parallel.
2010-02-21 | E-GEOD-19922 | biostudies-arrayexpress
Project description:An RNA binding polymer specifies nematode sperm fate
| PRJNA307605 | ENA