Investigating the role of SNU66 in 5' splice site maintenance
Ontology highlight
ABSTRACT:
ORGANISM(S): Caenorhabditis elegans
PROVIDER: GSE252787 | GEO | 2024/02/20
REPOSITORIES: GEO
Similar Datasets
Project description:The carboxy-terminus of the spliceosomal protein PRPF8, which regulates the RNA helicase Brr2, is a hotspot for mutations causing retinitis pigmentosa-type 13, with unclear role in human splicing and tissue-specificity mechanism. We used patient induced pluripotent stem cells-derived cells, carrying the heterozygous PRPF8 c.6926A>C (p.H2309P) mutation to demonstrate retinal-specific endophenotypes comprising photoreceptor loss, apical-basal polarity and ciliary defects. Comprehensive molecular, transcriptomic, and proteomic analyses revealed a role of the PRPF8/Brr2 regulation in 5’-splice site (5’SS) selection by spliceosomes, for which disruption impaired alternative splicing and weak/suboptimal 5’SS selection, and enhanced cryptic splicing, predominantly in ciliary and retinal-specific transcripts. Altered splicing efficiency, nuclear speckles organisation, and PRPF8 interaction with U6 snRNA, caused accumulation of active spliceosomes and poly(A)+ mRNAs in unique splicing clusters located at the nuclear periphery of photoreceptors. Collectively these elucidate the role of PRPF8/Brr2 regulatory mechanisms in splicing and the molecular basis of retinal disease, informing therapeutic approaches.
2024-03-01 | PXD043645 | Pride
Project description:The carboxy-terminus of the spliceosomal protein PRPF8, which regulates the RNA helicase Brr2, is a hotspot for mutations causing retinitis pigmentosa-type 13, with unclear role in human splicing and tissue-specificity mechanism. We used patient induced pluripotent stem cells-derived cells, carrying the heterozygous PRPF8 c.6926A>C (p.H2309P) mutation to demonstrate retinal-specific endophenotypes comprising photoreceptor loss, apical-basal polarity and ciliary defects. Comprehensive molecular, transcriptomic, and proteomic analyses revealed a role of the PRPF8/Brr2 regulation in 5’-splice site (5’SS) selection by spliceosomes, for which disruption impaired alternative splicing and weak/suboptimal 5’SS selection, and enhanced cryptic splicing, predominantly in ciliary and retinal-specific transcripts. Altered splicing efficiency, nuclear speckles organisation, and PRPF8 interaction with U6 snRNA, caused accumulation of active spliceosomes and poly(A)+ mRNAs in unique splicing clusters located at the nuclear periphery of photoreceptors. Collectively these elucidate the role of PRPF8/Brr2 regulatory mechanisms in splicing and the molecular basis of retinal disease, informing therapeutic approaches.
2024-02-16 | GSE236702 | GEO
Project description:The spliceosome undergoes extensive rearrangements as it assembles in multiple steps onto the precursor messenger RNA. In the earliest assembly step, U1snRNA identifies the 5' splice site through base-pairing interactions. However, U1snRNA leaves the spliceosome relatively early in the assembly process. The 5' splice site identity is subsequently maintained through interactions with U6snRNA, protein factor PRP8, and other components of the spliceosome during the complex assembly and rearrangements that build the catalytic site. Using a forward genetic screen in C. elegans, we have identified splicing suppressors of a locomotion defect caused by a 5'ss mutation. Here we report three new extragenic suppressor alleles from this screen, two in PRP8 and one in SNRNP200/Brr2. mRNASeq studies of these suppressor strains indicates that there are specific native targets with alternative 5' and alternative 3' splicing events affected by these suppressors, especially for the suppressor PRP8 D 1549N (position D1556 in humans). The strong suppressor at the unstructured N-terminus of SNRP200, N18K, indicates a potential regulatory role for this region. By examining distinct changes in the splicing of native genes, and by mapping these conserved suppressor residues onto cryoEM structural models of assembling human spliceosomes, we conclude that there are multiple interactions in the spliceosome that are required to ensure that the initial 5'ss identified by U1snRNA early in spliceosome assembly is the one that gets loaded into the catalytic core.The spliceosome undergoes extensive rearrangements as it assembles in multiple steps onto the precursor messenger RNA. In the earliest assembly step, U1snRNA identifies the 5' splice site through base-pairing interactions. However, U1snRNA leaves the spliceosome relatively early in the assembly process. The 5' splice site identity is subsequently maintained through interactions with U6snRNA, protein factor PRP8, and other components of the spliceosome during the complex assembly and rearrangements that build the catalytic site. Using a forward genetic screen in C. elegans, we have identified splicing suppressors of a locomotion defect caused by a 5'ss mutation. Here we report three new extragenic suppressor alleles from this screen, two in PRP8 and one in SNRNP200/Brr2. mRNASeq studies of these suppressor strains indicates that there are specific native targets with alternative 5' and alternative 3' splicing events affected by these suppressors, especially for the suppressor PRP8 D 1549N (position D1556 in humans). The strong suppressor at the unstructured N-terminus of SNRP200, N18K, indicates a potential regulatory role for this region. By examining distinct changes in the splicing of native genes, and by mapping these conserved suppressor residues onto cryoEM structural models of assembling human spliceosomes, we conclude that there are multiple interactions in the spliceosome that are required to ensure that the initial 5'ss identified by U1snRNA early in spliceosome assembly is the one that gets loaded into the catalytic core.
2021-11-23 | GSE189437 | GEO
Project description:Despite the major progress made into spliceosome mechanisms through CryoEM, a major obstacle of this approach is its inability to map the positioning of helicases on the RNA substrate. Here we present a method ‘psiCLIP’ which probes protein-RNA interactions in specific spliceosomal states. The method is based on iCLIP, but is different to previous iCLIP studies in that it is performed on step-specific spliceosomes that are prepared from in vitro splicing extracts, similarly to those prepared for CryoEM. We applied psiCLIP to SmB (C complex), Prp16 (C complex) and Prp22 (C* and P complexes), using pre-mRNA substrates based on UBC4 and ACT1. We also used ATPase deficient dominant negative (dn) mutants of Prp16 and Prp22 to determine the contribution of ATPase activity to binding patterns.
2020-03-15 | E-MTAB-8895 | biostudies-arrayexpress
Project description:Prp2 (U2AF65) is an essential splicing factor, that recognizes the poly-Py track near the 3' SS. We have previously observed that when Prp2 is inactivated using a prp2 temperature-sensitive allele, while most of the pre-mRNA is not processed, some are normally spliced. This observation even more noticeable during meiosis, since pre-mRNA regulation is executed at different levels (splicing, polyadenylation and stability). Using RNAseq, we have determined genome-wide the splicing efficiency of fission yeast cells as they progress into synchronous meiosis, in the presence or absence of functional Prp2. To our surprise, the main characteristic that defines if any given intron requires Prp2 to be spliced depends on the last nucleotides of the preceding exon, next to the 5’ SS in the nucleotide that anneal with U1 RNA. Thus, by simple genetic manipulation, we can modulate the dependency of any given intron on Prp2. We propose that the role of Prp2 consist in the stabilization of a loop between the 5’SS and the 3’ SS, which also depends on the annealing strength between U1 and the 5’ SS.
2021-10-01 | GSE97203 | GEO
Project description:RNA transcriptome sequencing analysis was performed in SNU-668 Erastin-resistant cells and SNU-668 parental cells, SNU-484 RSL3-resistant cells and SNU-484 parental cells
2024-04-04 | GSE262114 | GEO
Project description:Transcriptional profiling of C. elegans NHR-49, NHR-66 and NHR-80
2012-03-15 | GSE34856 | GEO
Project description:Sphingolipids are required for diverse biological functions and are degraded by specific catabolic enzymes. However, the mechanisms that regulate sphingolipid catabolism are not known. Here we characterize a transcriptional axis that regulates sphingolipid breakdown to control resistance against bacterial infection. From an RNAi screen for transcriptional regulators of pathogen resistance in the nematode C. elegans, we identified the nuclear hormone receptor nhr-66, a ligand-gated transcription factor homologous to human hepatocyte nuclear factor 4. Tandem chromatin immunoprecipitation-sequencing (ChIP-seq) and RNA sequencing (RNA-seq) experiments revealed that NHR-66 is a transcriptional repressor, which directly targets sphingolipid catabolism genes. Transcriptional de-repression of two sphingolipid catabolic enzymes in nhr-66 loss-of-function mutants drives the breakdown of sphingolipids, which enhances host susceptibility to infection with the bacterial pathogen Pseudomonas aeruginosa. These data define transcriptional control of sphingolipid catabolism in the regulation of cellular sphingolipids, a process that is necessary for pathogen resistance.
2023-09-08 | GSE240425 | GEO
Project description:Sphingolipids are required for diverse biological functions and are degraded by specific catabolic enzymes. However, the mechanisms that regulate sphingolipid catabolism are not known. Here we characterize a transcriptional axis that regulates sphingolipid breakdown to control resistance against bacterial infection. From an RNAi screen for transcriptional regulators of pathogen resistance in the nematode C. elegans, we identified the nuclear hormone receptor nhr-66, a ligand-gated transcription factor homologous to human hepatocyte nuclear factor 4. Tandem chromatin immunoprecipitation-sequencing (ChIP-seq) and RNA sequencing (RNA-seq) experiments revealed that NHR-66 is a transcriptional repressor, which directly targets sphingolipid catabolism genes. Transcriptional de-repression of two sphingolipid catabolic enzymes in nhr-66 loss-of-function mutants drives the breakdown of sphingolipids, which enhances host susceptibility to infection with the bacterial pathogen Pseudomonas aeruginosa. These data define transcriptional control of sphingolipid catabolism in the regulation of cellular sphingolipids, a process that is necessary for pathogen resistance.
2023-09-08 | GSE240426 | GEO
Project description:Transcriptional profiling of C. elegans NHR-49, NHR-66 and NHR-80 Independent data sets were generated for each mutant vs wildtpe comparison: NHR-49 (N=2), NHR-66 (N=3), NHR-80 (N=4)
2012-03-15 | E-GEOD-34856 | biostudies-arrayexpress