The exon junction complex core factor eIF4A3 is a key regulator of HPV16 gene expression.
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ABSTRACT:
SUBMITTER: Meznad K
PROVIDER: S-EPMC8026852 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Regulation of dendritically localized mRNAs offers an important means by which neurons can sculpt precise signals at synapses. Arc is one such dendritically localized mRNA, and it has been shown to contain two exon-junction complexes (EJCs) within its 3'UTR. The EJC has been postulated to regulate cytoplasmic Arc mRNA availability through translation-dependent decay and thus contribute to synaptic plasticity. Core proteins of the EJC include eIF4A3, an RNA helicase, and Magoh, which stabilizes the interaction of eIF4A3 with target mRNAs. Arc mRNA expression is activity-regulated in numerous brain regions, including the dorsal striatum and hippocampus. Therefore in this study, the in vivo expression of these core EJC components was investigated in adult Sprague-Dawley rats to determine whether there are also behaviorally regulated changes in their expression. In the present work, there was no change in the expression of Magoh mRNA following spatial exploration, a paradigm previously reported to robustly and reliably upregulate Arc mRNA expression. Interestingly, however, there were increases in eIF4A3 mRNA levels in the dorsal striatum and hippocampus following spatial exploration, similar to previous reports for Arc mRNA. Furthermore, there were activity-dependent changes in eIF4A3 protein distribution and expression within the striatum following spatial exploration. Importantly, eIF4A3 protein colocalized with Arc mRNA in vivo. Like Arc mRNA expression, eIF4A3 mRNA expression in the dorsomedial striatum, but not dorsolateral striatum or hippocampus, significantly correlated with behavioral performance on a striatally-mediated, response-reversal learning task. This study provides direct evidence that a core EJC component, eIF4A3, shows activity-dependent changes in both mRNA and protein expression in the adult mammalian brain. These findings thus further implicate eIF4A3 as a key mediator of Arc mRNA availability underlying learning and memory processes in vivo.
| S-EPMC3490057 | biostudies-literature
Project description:Pre-mRNA splicing not only removes introns and joins exons to generate spliced mRNA but also results in remodeling of the spliced messenger ribonucleoprotein, influencing various downstream events. This remodeling includes the loading of an exon-exon junction complex (EJC). It is unclear how the spliceosome recruits the EJC onto the mRNA and whether EJC formation or EJC components are required for pre-mRNA splicing. Here we immunodepleted the EJC core component eIF4A3 from HeLa cell nuclear extract and found that eIF4A3 is dispensable for pre-mRNA splicing in vitro. However, eIF4A3 is required for the splicing-dependent loading of the Y14/Magoh heterodimer onto mRNA, and this activity of human eIF4A3 is also present in the Drosophila ortholog. Surprisingly, the loading of six other EJC components was not affected by eIF4A3 depletion, suggesting that their binding to mRNA involves different or redundant pathways. Finally, we found that the assembly of the EJC onto mRNA occurs at the late stages of the splicing reaction and requires the second-step splicing and mRNA-release factor HRH1/hPrp22. The EJC-dependent and -independent recruitment of RNA-binding proteins onto mRNA suggests a role for the EJC in messenger ribonucleoprotein remodeling involving interactions with other proteins already bound to the pre-mRNA, which has implications for nonsense-mediated mRNA decay and other mRNA transactions.
| S-EPMC1913901 | biostudies-other
Project description:The exon junction complex (EJC) is loaded onto mRNAs as a consequence of splicing and regulates multiple posttranscriptional events. MLN51, Magoh, Y14, and eIF4A3 form a highly stable EJC core, but where this tetrameric complex is assembled in the cell remains unclear. Here we show that EJC factors are enriched in domains that we term perispeckles and are visible as doughnuts around nuclear speckles. Fluorescence resonance energy transfer analyses and EJC assembly mutants show that perispeckles do not store free subunits, but instead are enriched for assembled cores. At the ultrastructural level, perispeckles are distinct from interchromatin granule clusters that may function as storage sites for splicing factors and intermingle with perichromatin fibrils, where nascent RNAs and active RNA Pol II are present. These results support a model in which perispeckles are major assembly sites for the tetrameric EJC core. This subnuclear territory thus represents an intermediate region important for mRNA maturation, between transcription sites and splicing factor reservoirs and assembly sites.
| S-EPMC3338441 | biostudies-literature
Project description:Using TSG101 pre-mRNA, we previously discovered cancer-specific re-splicing of mature mRNA that generates aberrant transcripts/proteins. The fact that mRNA is aberrantly re-spliced in various cancer cells implies there must be an important mechanism to prevent deleterious re-splicing on the spliced mRNA in normal cells. We thus postulated that mRNA re-splicing is controlled by specific repressors, and we searched for repressor candidates by siRNA-based screening for mRNA re-splicing activity. We found that knock-down of EIF4A3, which is a core component of the exon junction complex (EJC), significantly promoted mRNA re-splicing. Remarkably, we could recapitulate cancer-specific mRNA re-splicing in normal cells by knock-down of any of the core EJC proteins, EIF4A3, MAGOH, or RBM8A (Y14), implicating the EJC core as the repressor of mRNA re-splicing often observed in cancer cells. We propose that the EJC core is a critical mRNA quality control factor to prevent over-splicing of mature mRNA.
| S-EPMC8234774 | biostudies-literature
Project description:Key messageMolecular and functional characterization of four gene families of the Physalis exon junction complex (EJC) core improved our understanding of the evolution and function of EJC core genes in plants. The exon junction complex (EJC) plays significant roles in posttranscriptional regulation of genes in eukaryotes. However, its developmental roles in plants are poorly known. We characterized four EJC core genes from Physalis floridana that were named PFMAGO, PFY14, PFeIF4AIII and PFBTZ. They shared a similar phylogenetic topology and were expressed in all examined organs. PFMAGO, PFY14 and PFeIF4AIII were localized in both the nucleus and cytoplasm while PFBTZ was mainly localized in the cytoplasm. No protein homodimerization was observed, but they could form heterodimers excluding the PFY14-PFBTZ heterodimerization. Virus-induced gene silencing (VIGS) of PFMAGO or PFY14 aborted pollen development and resulted in low plant survival due to a leaf-blight-like phenotype in the shoot apex. Carpel functionality was also impaired in the PFY14 knockdowns, whereas pollen maturation was uniquely affected in PFBTZ-VIGS plants. Once PFeIF4AIII was strongly downregulated, plant survival was reduced via a decomposing root collar after flowering and Chinese lantern morphology was distorted. The expression of Physalis orthologous genes in the DYT1-TDF1-AMS-bHLH91 regulatory cascade that is associated with pollen maturation was significantly downregulated in PFMAGO-, PFY14- and PFBTZ-VIGS flowers. Intron-retention in the transcripts of P. floridana dysfunctional tapetum1 (PFDYT1) occurred in these mutated flowers. Additionally, the expression level of WRKY genes in defense-related pathways in the shoot apex of PFMAGO- or PFY14-VIGS plants and in the root collar of PFeIF4AIII-VIGS plants was significantly downregulated. Taken together, the Physalis EJC core genes play multiple roles including a conserved role in male fertility and newly discovered roles in Chinese lantern development, carpel functionality and defense-related processes. These data increase our understanding of the evolution and functions of EJC core genes in plants.
| S-EPMC6280879 | biostudies-literature
Project description:Using the TSG101 pre-mRNA, we previously discovered cancer-specific re-splicing of mature mRNA that generates aberrant transcripts/proteins. The fact that mRNA is aberrantly re-spliced in various cancer cells implies there must be an important mechanism to prevent deleterious re-splicing on the spliced mRNA in normal cells. We thus postulated that the mRNA re-splicing is controlled by specific repressors and we searched for repressor candidates by siRNA-based screening for mRNA re-splicing activity. We found that knock-down of EIF4A3, which is a core component of the exon junction complex (EJC), significantly promoted mRNA re-splicing. Remarkably, we could recapitulate cancer-specific mRNA re-splicing in normal cells by knock-down of any of the core EJC proteins, EIF4A3, MAGOH or RBM8A (Y14), implicating the EJC core as the repressor of mRNA re-splicing often observed in cancer cells. We propose that the EJC core is a critical mRNA quality control factor to prevent over-splicing of mature mRNA.
2021-06-15 | GSE178102 | GEO
Project description:In metazoan cells, spliced mRNAs are marked by the exon junction complex (EJC), a multi-protein complex that serves as a key regulator of post-transcriptional mRNA metabolism. Deposition of EJCs on mRNA is intimately linked to the splicing process. The spliceosomal protein CWC22 directly binds the core EJC-protein eIF4A3, guides it to the spliceosome and initiates EJC assembly. In addition, CWC22 is involved in the splicing process itself, but the molecular details of its dual function remain elusive. Here we analyze the mechanisms, by which CWC22 co-regulates pre-mRNA splicing and EJC assembly. We show that the core of CWC22 is sufficient to mediate both pre-mRNA splicing and EJC assembly. Nonetheless, both processes can be functionally uncoupled with an eIF4A3-binding deficient mutant of CWC22, which impedes EJC assembly. A C-terminal domain of CWC22 strongly enhances its spliceosomal interaction and likely regulates its function. High-throughput RNA-sequencing identifies global defects of pre-mRNA splicing and downregulation of diverse gene expression pathways in CWC22-depleted cells. We propose a model, in which CWC22 represents an integral component of the spliceosome and orchestrates pre-mRNA splicing and eIF4A3 binding to achieve global assembly of exon junction complexes.
| S-EPMC4482076 | biostudies-literature
Project description:The exon junction complex (EJC) is an RNA binding complex comprised of the core components Magoh, Rbm8a, and Eif4a3. Human mutations in EJC components cause neurodevelopmental pathologies. Further, mice heterozygous for either Magoh or Rbm8a exhibit aberrant neurogenesis and microcephaly. Yet despite the requirement of these genes for neurodevelopment, the pathogenic mechanisms linking EJC dysfunction to microcephaly remain poorly understood. Here we employ mouse genetics, transcriptomic and proteomic analyses to demonstrate that haploinsufficiency for each of the 3 core EJC components causes microcephaly via converging regulation of p53 signaling. Using a new conditional allele, we first show that Eif4a3 haploinsufficiency phenocopies aberrant neurogenesis and microcephaly of Magoh and Rbm8a mutant mice. Transcriptomic and proteomic analyses of embryonic brains at the onset of neurogenesis identifies common pathways altered in each of the 3 EJC mutants, including ribosome, proteasome, and p53 signaling components. We further demonstrate all 3 mutants exhibit defective splicing of RNA regulatory proteins, implying an EJC dependent RNA regulatory network that fine-tunes gene expression. Finally, we show that genetic ablation of one downstream pathway, p53, significantly rescues microcephaly of all 3 EJC mutants. This implicates p53 activation as a major node of neurodevelopmental pathogenesis following EJC impairment. Altogether our study reveals new mechanisms to help explain how EJC mutations influence neurogenesis and underlie neurodevelopmental disease.
| S-EPMC5019403 | biostudies-literature
Project description:The formation of the nervous system requires a balance between proliferation, differentiation, and migration of neural progenitors (NPs). Mutations in genes regulating development impede neurogenesis and lead to neuropsychiatric diseases, including autism spectrum disorders (ASDs) and schizophrenia (SZ). Recently, mutations in nonsense-mediated mRNA decay genes have been associated with ASDs, intellectual disability (ID), and SZ. Here, we examine the function of a gene in the exon junction complex, Rbm8a, in the cortical development. When Rbm8a is selectively knocked out in neural stem cells, the resulting mice exhibit microcephaly, early postnatal lethality, and altered distribution of excitatory neurons in the neocortex. Moreover, Rbm8a haploinsufficiency in the central nervous system decreases cell proliferation in the ganglionic eminences. Parvalbumin+ and neuropeptide Y+ interneurons in the cortex are significantly reduced, and distribution of interneurons are altered. Consistently, neurons in the cortex of conditional knockout (cKO) mice show a significant decrease in GABA frequency. Transcriptomic analysis revealed differentially expressed genes enriched in telencephalon development and mitosis. To further investigate the role of Rbm8a in interneuron differentiation, conditional KO of Rbm8a in NKX2.1 interneuron progenitor cells reduces progenitor proliferation and alters interneuron distributions. Taken together, these data reveal a critical role of Rbm8a in interneuron development, and establish that perturbation of this gene leads to profound cortical deficits.
| S-EPMC7644723 | biostudies-literature
Project description:The exon junction complex (EJC) is deposited on mRNAs as a consequence of splicing and influences postsplicing mRNA metabolism. The Mago-Y14 heterodimer is a core component of the EJC. Recently, the protein PYM has been identified as an interacting partner of Mago-Y14. Here we show that PYM is a cytoplasmic RNA-binding protein that is excluded from the nucleus by Crm1. PYM interacts directly with Mago-Y14 by means of its N-terminal domain. The crystal structure of the Drosophila ternary complex at 1.9 A resolution reveals that PYM binds Mago and Y14 simultaneously, capping their heterodimerization interface at conserved surface residues. Formation of this ternary complex is also observed with the human proteins. Mago residues involved in the interaction with PYM have been implicated in nonsense-mediated mRNA decay (NMD). Consistently, human PYM is active in NMD tethering assays. Together, these data suggest a role for PYM in NMD.
| S-EPMC1299003 | biostudies-literature