Structural recognition of the mRNA 3’ UTR by PUF-8 restricts the lifespan of C. elegans
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ABSTRACT:
PROVIDER: PRJNA693156 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR13481282_1.fastq.gz | Fastqsanger.gz | |||
| SRR13481282_2.fastq.gz | Fastqsanger.gz | |||
| SRR13481283_1.fastq.gz | Fastqsanger.gz | |||
| SRR13481283_2.fastq.gz | Fastqsanger.gz | |||
| SRR13481284_1.fastq.gz | Fastqsanger.gz |
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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 molecular mechanisms of aging are unsolved 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 with the 3' untranslated regions (3' UTR) in the target mRNAs. Recently, PUF-8 was reported to alter mitochondrial dynamics and mitophagy by regulating MFF-1, a mitochondrial fission factor, and subsequently regulated 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. Those mutations reduced the fertility and extended the lifespan of C. elegans. Deep sequencing of total mRNAs from wild-type and puf-8 mutant worms as well as in vivo RNA Crosslinking and Immunoprecipitation (CLIP) experiments identified six PUF-8 regulated genes, which contain at least one PUF-binding element (PBE) at the 3' UTR. 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 mutant animals. We conclude that PUF-8 regulate the lifespan of C. elegans may not only via MFF but also via modulating pqm-1-related pathways.
| S-EPMC8464079 | biostudies-literature
Project description:RNA-coimmunopurifications with TAP-tagged Puf proteins from Saccharomyces cereviseae. Untagged strain (BY4741) served as a control. Cells were grown to midlog phase and harvested by centrifugation. TAP-tagged Puf proteins were affinity purified from cell-free extracts with IgG sepharose and eluted with TEV protease. RNA was isolated from extract (=input)and from purified protein samples by phenol-chloroform extraction. RNA samples were reverse transcribed using a mixture of oligo-dT and random nonamer oligos in the presence of amino-allyl dUTP/ dNTP mixture. cDNAs were fluorescently labeled and hybridized on yeast DNA microarrays over night at 65 degrees. For a detailed procedure see http://microarray-pubs.stanford.edu/yeast_puf and also Gerber AP et al. PLoS Biology, 2004. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set
2006-01-04 | GSE3968 | GEO
Project description:Gene expression profiling was performed to compare RNA abundances in mycelium of Puf knockout strains compared to that in mycelium of wild-type N. crassa
2015-03-16 | GSE50995 | GEO
Project description:RNA-coimmunopurifications with TAP-tagged Puf proteins from Saccharomyces cereviseae. Untagged strain (BY4741) served as a control. Cells were grown to midlog phase and harvested by centrifugation. TAP-tagged Puf proteins were affinity purified from cell-free extracts with IgG sepharose and eluted with TEV protease. RNA was isolated from extract (=input)and from purified protein samples by phenol-chloroform extraction. RNA samples were reverse transcribed using a mixture of oligo-dT and random nonamer oligos in the presence of amino-allyl dUTP/ dNTP mixture. cDNAs were fluorescently labeled and hybridized on yeast DNA microarrays over night at 65 degrees. For a detailed procedure see http://microarray-pubs.stanford.edu/yeast_puf and also Gerber AP et al. PLoS Biology, 2004. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Computed
2006-01-03 | E-GEOD-3968 | biostudies-arrayexpress
Project description:RNA-coimmunopurifications with TAP-tagged Puf proteins from Saccharomyces cereviseae. Untagged strain (BY4741) served as a control. Cells were grown to midlog phase and harvested by centrifugation. TAP-tagged Puf proteins were affinity purified from cell-free extracts with IgG sepharose and eluted with TEV protease. RNA was isolated from extract (=input)and from purified protein samples by phenol-chloroform extraction. RNA samples were reverse transcribed using a mixture of oligo-dT and random nonamer oligos in the presence of amino-allyl dUTP/ dNTP mixture. cDNAs were fluorescently labeled and hybridized on yeast DNA microarrays over night at 65 degrees. For a detailed procedure see http://microarray-pubs.stanford.edu/yeast_puf and also Gerber AP et al. PLoS Biology, 2004.
2005-11-01 | E-SMDB-2622 | biostudies-arrayexpress
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:Self-recognition of an inducible host lncRNA by RIG-I feedback restricts innate immune response
| PRJNA416003 | 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