Abstract:Messenger RNA surveillance, the selective and rapid degradation of mRNAs containing premature stop codons, occurs in all eukaryotes tested. The biological role of this decay pathway, however, is not well understood. To identify natural substrates of mRNA surveillance, we used a cDNA-based representational difference analysis to identify mRNAs whose abundance increases in Caenorhabditis elegans smg(−) mutants, which are deficient for mRNA surveillance. Alternatively spliced mRNAs of genes encoding ribosomal pro… Show more
“…However, we failed to obtain stable transgenic lines in the wild-type, n5119 or n5132 background using an rbm-5a cDNA transgene, probably due to the toxicity of the transgene. To overcome this problem, we used the ubiquitous eft-3 promoter [45] to drive an rbm-5a cDNA::GFP fusion transgene and obtained stable transgenic lines. The RBM-5A::GFP fusion protein was exclusively localized in the nuclei of the numerous visible cells (Figure 4(e,f)), suggesting that RBM-5 is a nuclear protein.10.1080/15476286.2018.1526540-F0004Figure 4.Fluorescent pictures of animals expressing a Prbm-5::GFP transcriptional fusion transgene and a Peft-3::rbm-5a cDNA::GFP translational fusion transgene.(a) A low resolution picture of Prbm-5::GFP transgenic animals showing GFP expression in the anterior, posterior and ventral nerve cord (VNC).…”
Section: Resultsmentioning
confidence: 99%
“…To construct the Peft-3::rbm-5a cDNA::GFP plasmid, a PCR-amplified eft-3 promoter (a 597-bp (−16 to −612 base) fragment upstream of the eft-3 start codon) [45] was subcloned to pPD95_79 using Sbf I/ Xma I sites. The rbm-5a cDNA was amplified and subcloned to the pPD95_79- Peft-3 backbone using Xma I/ Kpn I sites.…”
A key step in pre-mRNA splicing is the recognition of 3ʹ splicing sites by the U2AF large and small subunits, a process regulated by numerous trans-acting splicing factors. How these trans-acting factors interact with U2AF in vivo is unclear. From a screen for suppressors of the temperature-sensitive (ts) lethality of the C. elegans U2AF large subunit gene uaf-1(n4588) mutants, we identified mutations in the RNA binding motif gene rbm-5, a homolog of the tumor suppressor gene RBM5. rbm-5 mutations can suppress uaf-1(n4588) ts-lethality by loss of function and neuronal expression of rbm-5 was sufficient to rescue the suppression. Transcriptome analyses indicate that uaf-1(n4588) affected the expression of numerous genes and rbm-5 mutations can partially reverse the abnormal gene expression to levels similar to that of wild type. Though rbm-5 mutations did not obviously affect alternative splicing per se, they can suppress or enhance, in a gene-specific manner, the altered splicing of genes in uaf-1(n4588) mutants. Specifically, the recognition of a weak 3ʹ splice site was more susceptible to the effect of rbm-5. Our findings provide novel in vivo evidence that RBM-5 can modulate UAF-1-dependent RNA splicing and suggest that RBM5 might interact with U2AF large subunit to affect tumor formation.
“…However, we failed to obtain stable transgenic lines in the wild-type, n5119 or n5132 background using an rbm-5a cDNA transgene, probably due to the toxicity of the transgene. To overcome this problem, we used the ubiquitous eft-3 promoter [45] to drive an rbm-5a cDNA::GFP fusion transgene and obtained stable transgenic lines. The RBM-5A::GFP fusion protein was exclusively localized in the nuclei of the numerous visible cells (Figure 4(e,f)), suggesting that RBM-5 is a nuclear protein.10.1080/15476286.2018.1526540-F0004Figure 4.Fluorescent pictures of animals expressing a Prbm-5::GFP transcriptional fusion transgene and a Peft-3::rbm-5a cDNA::GFP translational fusion transgene.(a) A low resolution picture of Prbm-5::GFP transgenic animals showing GFP expression in the anterior, posterior and ventral nerve cord (VNC).…”
Section: Resultsmentioning
confidence: 99%
“…To construct the Peft-3::rbm-5a cDNA::GFP plasmid, a PCR-amplified eft-3 promoter (a 597-bp (−16 to −612 base) fragment upstream of the eft-3 start codon) [45] was subcloned to pPD95_79 using Sbf I/ Xma I sites. The rbm-5a cDNA was amplified and subcloned to the pPD95_79- Peft-3 backbone using Xma I/ Kpn I sites.…”
A key step in pre-mRNA splicing is the recognition of 3ʹ splicing sites by the U2AF large and small subunits, a process regulated by numerous trans-acting splicing factors. How these trans-acting factors interact with U2AF in vivo is unclear. From a screen for suppressors of the temperature-sensitive (ts) lethality of the C. elegans U2AF large subunit gene uaf-1(n4588) mutants, we identified mutations in the RNA binding motif gene rbm-5, a homolog of the tumor suppressor gene RBM5. rbm-5 mutations can suppress uaf-1(n4588) ts-lethality by loss of function and neuronal expression of rbm-5 was sufficient to rescue the suppression. Transcriptome analyses indicate that uaf-1(n4588) affected the expression of numerous genes and rbm-5 mutations can partially reverse the abnormal gene expression to levels similar to that of wild type. Though rbm-5 mutations did not obviously affect alternative splicing per se, they can suppress or enhance, in a gene-specific manner, the altered splicing of genes in uaf-1(n4588) mutants. Specifically, the recognition of a weak 3ʹ splice site was more susceptible to the effect of rbm-5. Our findings provide novel in vivo evidence that RBM-5 can modulate UAF-1-dependent RNA splicing and suggest that RBM5 might interact with U2AF large subunit to affect tumor formation.
“…That is, as an RNA binding protein increases in abundance, it increasingly binds its own pre-mRNA and facilitates production of a 3UI-containing form subject to NMD, thereby maintaining protein homeostasis [44,67,68]. Proteins undergoing this type of regulation include ribosomal proteins [42,[69][70][71], core spliceosomal proteins [66,72] and alternative splicing regulators such as hnRNP and SR proteins [45,67]. In fact, every one of the 11 human SR proteins has a 3UI isoform and regulates its own production by AS-NMD [67].…”
Section: Nmd Inhibition Points To Functional 3ui-containing Transcriptsmentioning
Although introns in 5 0 -and 3 0 -untranslated regions (UTRs) are found in many protein coding genes, rarely are they considered distinctive entities with specific functions. Indeed, mammalian transcripts with 3 0 -UTR introns are often assumed nonfunctional because they are subject to elimination by nonsense-mediated decay (NMD). Nonetheless, recent findings indicate that 5 0 -and 3 0 -UTR intron status is of significant functional consequence for the regulation of mammalian genes. Therefore these features should be ignored no longer.
“…The ratio of productive to unproductive alternative splicing of rpl-12 is affected by levels of RPL-12 protein, indicating that unproductive splicing of rpl-12 is under feedback control. 64 More recently, NMD-target isoforms of the human ribosomal protein genes rpL3 and rpL12 were identified and the unproductive splicing of rpL3 was shown to be autoregulated by rpL3 protein in a negative feedback loop. 99 RUST thus seems to play a similar role in the regulation of ribosomal proteins in species from yeast to human.…”
M ost human genes exhibit alternative splicing, but not all alternatively spliced transcripts produce functional proteins. Computational and experimental results indicate that a substantial fraction of alternative splicing events in humans result in mRNA isoforms that harbor a premature termination codon (PTC). These transcripts are predicted to be degraded by the nonsense-mediated mRNA decay (NMD) pathway. One explanation for the abundance of PTC-containing isoforms is that they represent splicing errors that are identified and degraded by the NMD pathway. Another potential explanation for this startling observation is that cells may link alternative splicing and NMD to regulate the abundance of mRNA transcripts. This mechanism, which we call "Regulated Unproductive Splicing and Translation" (RUST), has been experimentally shown to regulate expression of a wide variety of genes in many organisms from yeast to human. It is frequently employed for autoregulation of proteins that affect the splicing process itself. Thus, alternative splicing and NMD act together to play an important role in regulating gene expression.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
