Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Pervasive changes of mRNA splicing in upf1 deficient zebrafish identify rpl10a as a regulator of T cell development

ABSTRACT: The transcriptome of eukaryotic cells is constantly monitored for errors to avoid the production of undesired protein variants. The evolutionarily conserved nonsense-mediated mRNA decay (NMD) pathway degrades aberrant mRNAs, but also functions in the regulation of transcript abundance in response to changed physiological states. As a result of its essential functions, only few animal models of impaired NMD activity have been developed. Here, we describe a zebrafish mutant of upf1, encoding the central component of the NMD machinery. Fish homozygous for the upf1t20450 allele (Y163X) survive until day 10 after fertilization, presenting with impaired T cell development as one of the most conspicuous features of the mutant phenotype. Analysis of differentially expressed genes identified dysregulation of the pre-mRNA splicing pathway, accompanied by perturbed auto-regulation of canonical splicing activators (SRSF) and repressors (HNRNP). In upf1-deficient mutants, NMD-susceptible transcripts of ribosomal proteins that are known for their role as non-canonical splicing regulators were greatly increased, most notably, rpl10a. When the levels of NMD-susceptible rpl10a transcripts were artifically increased in zebrafish morphants, T cell development was significantly impaired, suggesting that perturbed autoregulation of rpl10a splicing contributes to failing T cell development in upf1 deficiency. Our results identify an extra-ribosomal tissue-specific function to rpl10a in the immune system, and thus exemplify the advantages of the zebrafish model to study the effects of upf1-deficiency in the context of a vertebrate organism.

ORGANISM(S): Danio rerio

PROVIDER: GSE136669 | GEO | 2020/07/13

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

Pervasive changes of mRNA splicing in upf1 deficient zebrafish identify rpl10a as a regulator of T cell development

Project description:Pervasive changes of mRNA splicing in upf1 deficient zebrafish identify rpl10a as a regulator of T cell development

| PRJNA563011 | ENA

Nonsense-mediated decay of alternative precursor mRNA splicing variants is a major determinant of the eukaryotic steady state transcriptome

Project description:Nonsense-mediated mRNA decay (NMD) is a eukaryotic surveillance pathway that recognizes aberrant transcripts arising from mutations and transcriptional mistakes. Moreover, differential transcript processing such as alternative precursor mRNA splicing (AS) can generate NMD substrates, however, the extent of coupled AS and NMD remained unclear. To investigate NMD targeting of AS variants, we performed transcriptome-wide splicing studies using Arabidopsis thaliana single and double mutants in the NMD factor homologues UPF1 and UPF3, as well as samples treated with the translation inhibitor cycloheximide (CHX). Our analyses revealed that at least 17.5% of all multi-exon, protein-coding genes produce splicing variants of all major types that are targeted by NMD, with a significant fraction originating from the splicing of cryptic introns. Furthermore, accumulation of many intron-retained mRNAs in the mutants, but not in response to CHX suggests the action of distinct routes of NMD with variable impact of translation. Importantly, 92.3% of the NMD-responsive mRNAs exhibit classical NMD-eliciting features, supporting their authenticity as direct targets. NMD-dependent AS variants are linked to diverse biological functions, including the poison exon-mediated regulation of signaling and posttranslational protein modification components. In addition to mRNAs, numerous non-coding RNAs as well as newly identified transcripts derived from intergenic regions were shown to be NMD responsive. In summary, our comprehensive analysis of AS-coupled NMD provides strong evidence for a major function of this pathway in shaping the transcriptome, having fundamental implications in gene regulation and quality control of transcript processing steps in higher eukaryotes. Comparison of gene expression and alternative splicing patterns in control and nonsense-mediated decay (NMD)-impaired Arabidopsis seedlings

2013-10-01 | E-GEOD-41432 | biostudies-arrayexpress

Nonsense Mediated mRNA decay mutes the splicing defects of spliceosome component mutations

Project description:The role of many splicing factors in pre-mRNA splicing and the involvement of these factors in the processing of specific transcripts have often been defined through the analysis of loss of function mutants in vivo. Here we show that inactivating the nonsense mediated mRNA decay (NMD) results in an enhancement of splicing phenotypes associated with several splicing factors mutations. Tiling microarrays showed that inactivation of the NMD factor Upf1p in the prp17Î and prp18Î mutant strains reveals a larger spectrum of splicing defects than what is observed in the single mutants, including new transcripts previously shown unaffected by Prp17p or Prp18p inactivation. In addition, inactivation of Upf1 in the prp22-1 mutant enhances the unspliced precursor accumulation phenotype of several specific transcripts and partially suppresses growth defects associated with the prp17Î or prp22-1 mutations. These results support the idea that RNA surveillance by NMD mutes some of the effects of splicing factors mutations and show that the roles of splicing factors and their transcripts specificity cannot be fully understood in vivo unless RNA degradation systems that degrade unspliced precursors are also inactivated. Three samples from the Prp17delta, Prp18delta, Prp17deltaUpf1delta and Prp18Upf1delta mutants were grown indepedently and analyzed by tiling arrays to understand the role of the NMD component Upf1 in minimizing the accumulation of unspliced RNAs generated by the Prp17delta and Prp18delta splicing mutations.

2010-05-16 | E-GEOD-18288 | biostudies-arrayexpress

A post-translational regulatory switch on UPF1 controls targeted mRNA degradation.

Project description:Nonsense-mediated mRNA decay (NMD) controls the quality of eukaryotic gene expression and also degrades physiologic mRNAs. How NMD targets are identified is incompletely understood. A central NMD factor is the ATP-dependent RNA helicase UPF1. Neither the distance in space between the termination codon and the poly(A) tail nor the binding of steady-state, largely hypophosphorylated UPF1 is a discriminating marker of cellular NMD targets, unlike for PTC-containing reporter mRNAs when compared to their PTC-free counterparts. Here, we map phosphorylated UPF1 (p-UPF1) binding sites using transcriptome-wide footprinting or DNA oligonucleotide-directed mRNA cleavage to report that p-UPF1 provides the first reliable cellular NMD-target marker. p-UPF1 is enriched on NMD target 3'UTRs along with SMG5 and SMG7 but not SMG1 or SMG6. Immunoprecipitations of UPF1 variants deficient in various aspects of the NMD process in parallel with FRET experiments reveal that ATPase/helicase-deficient UPF1 manifests high levels of RNA binding and disregulated hyperphosphorylation, whereas wild-type UPF1 releases from nonspecific RNA interactions in an ATP hydrolysis-dependent mechanism until an NMD target is identified. 3'UTR-associated UPF1 undergoes regulated phosphorylation on NMD targets, providing a binding platform for mRNA degradative activities. p-UPF1 binding to NMD target 3'UTRs is stabilized by SMG5 and SMG7. Our results help to explain why steady-state UPF1 binding is not a marker for cellular NMD substrates and how this binding is transformed to induce mRNA decay.

2014-08-15 | GSE60045 | GEO

A post-translational regulatory switch on UPF1 controls targeted mRNA degradation.

2014-08-15 | E-GEOD-60045 | biostudies-arrayexpress

The origins and consequences of UPF1 variants in pancreatic adenosquamous carcinoma

Project description:Pancreatic adenosquamous carcinoma (PASC) is an aggressive cancer whose mutational origins are poorly understood. An early study reported high-frequency somatic mutations affecting UPF1, a core nonsense-mediated mRNA decay (NMD) factor, in PASC, but subsequent studies did not observe these lesions. The corresponding controversy about whether UPF1 mutations are important contributors to PASC has been exacerbated by a paucity of functional studies. Here, we modeled two UPF1 mutations to find no significant effects on pancreatic cancer growth, acquisition of adenosquamous features, UPF1 splicing, UPF1 protein levels, or NMD efficiency. We subsequently discovered that ~40% of UPF1 mutations reportedly present in PASCs are identical to standing genetic variants in the human population, suggesting that they may be non-pathogenic inherited variants rather than pathogenic mutations. Our data suggest that UPF1 is not a common functional driver of PASC and motivate further attempts to identify unique genetic features defining these malignancies.

2020-12-19 | GSE163517 | GEO

RNA-seq profiling of the Tg(myo6b:GFP-2A-rpl10a-3xHA) zebrafish hair cell IP vs. whole fish IN

Project description:We have developed and tested the efficiency of the Tg(myo6b:GFP-2A-rpl10a-3xHA) zebrafish to specifically enrich for and evaluate the translatome of inner ear and lateral line sensory hair cells (IP) compared to the whole fish transcriptome (IN). We show through RNA-seq that HA-tagged ribosome immunoprecipitation significantly enriches for RNA transcripts of known zebrafish hair cell expressed transcripts.

2018-04-24 | GSE102861 | GEO

Target discrimination in nonsense-mediated mRNA decay requires Upf1 ATPase activity (CLIP-Seq)

Project description:RNA quality control pathways serve to get rid of faulty RNAs and therefore must be able to discriminate these RNAs from those that are normal. Here we present evidence that the ATPase cycle of SF1 Helicase Upf1 is required for mRNA discrimination during Nonsense-Mediated Decay (NMD). Mutations affecting the Upf1 ATPase cycle disrupt the mRNA selectivity of Upf1, leading to indiscriminate accumulation of NMD complexes on both NMD target and non-target mRNAs. In addition, two modulators of NMD - translation and termination codon-proximal poly(A) binding protein - depend on Upf1 ATPase to limit Upf1-non-target association. Preferential ATPase-dependent dissociation of Upf1 from non-target mRNAs in vitro suggests that selective release of Upf1 contributes to the ATPase-dependence of Upf1 target discrimination. Given the prevalence of helicases in RNA regulation, ATP hydrolysis may be an underappreciated, yet widely employed, activity in target RNA discrimination. CLIP and RIP-seq against Wild Type and Mutant Upf1 in HEK293-T cell lines

2015-07-06 | E-GEOD-69584 | biostudies-arrayexpress

Target discrimination in nonsense-mediated mRNA decay requires Upf1 ATPase activity (RIP-Seq)

2015-07-06 | E-GEOD-69585 | biostudies-arrayexpress