ChemRAP uncovers specific mRNA translation regulation via RNA 5'-Pme
Ontology highlight
ABSTRACT:
SUBMITTER: Dr. Hélène Ipas
PROVIDER: S-SCDT-10_1038-S44319-024-00059-Z | biostudies-other |
REPOSITORIES: biostudies-other
Similar Datasets
Project description:5'-end modifications play key roles in determining RNA fates. Phospho-methylation is a noncanonical cap occurring on either 5'-PPP or 5'-P ends. We used ChemRAP, in which affinity purification of cellular proteins with chemically synthesized modified RNAs is coupled to quantitative proteomics, to identify 5'-Pme "readers". We show that 5'-Pme is directly recognized by EPRS, the central subunit of the multisynthetase complex (MSC), through its linker domain, which has previously been involved in key noncanonical EPRS and MSC functions. We further determine that the 5'-Pme writer BCDIN3D regulates the binding of EPRS to specific mRNAs, either at coding regions rich in MSC codons, or around start codons. In the case of LRPPRC (leucine-rich pentatricopeptide repeat containing), a nuclear-encoded mitochondrial protein associated with the French Canadian Leigh syndrome, BCDIN3D deficiency abolishes binding of EPRS around its mRNA start codon, increases its translation but ultimately results in LRPPRC mislocalization. Overall, our results suggest that BCDIN3D may regulate the translation of specific mRNA via RNA-5'-Pme.
| S-EPMC10933402 | biostudies-literature
Project description:Regulation of gene expression at the translational level allows rapid adaptation of cellular proteomes to quickly changing environmental conditions and is thus central for prokaryotic organisms. Small non-coding RNAs (sRNAs) have been reported to effectively orchestrate translation control in bacteria and archaea mainly by targeting mRNAs by partial base complementarity. Here we report an unprecedented mechanism how sRNAs are capable of modulating protein biosynthesis in the halophilic archaeon Haloferax volcanii. By analyzing the ribosome-associated ncRNAs (rancRNAs) under different stress conditions we identified an intergenic sRNA, termed rancRNA_s194, that is primarily expressed during exponential growth under all tested conditions. By interaction with the ribosome rancRNA_s194 inhibits peptide bond formation and protein synthesis in vitro but appears to target a specific mRNA in vivo. The respective knock-out strain shows a reduced lag phase in media containing xylose as sole carbon source and outcompetes the wildtype cells under these conditions. Mass spectrometry, polysome profiling and mRNA binding competition experiments suggest that rancRNA_s194 prevents the cstA mRNA from being efficiently translated by H. volcanii ribosomes. These findings enlarge the regulatory repertoire of archaeal sRNAs in modulating post-transcriptional gene expression.
| S-EPMC6104027 | biostudies-literature
Project description:Down regulation of mRNA translation is an important problem in various bio-medical domains ranging from developing effective medicines for tumors and for viral diseases to developing attenuated virus strains that can be used for vaccination. Here, we study the problem of down regulation of mRNA translation using a mathematical model called the ribosome flow model (RFM). In the RFM, the mRNA molecule is modeled as a chain of n sites. The flow of ribosomes between consecutive sites is regulated by n + 1 transition rates. Given a set of feasible transition rates, that models the outcome of all possible mutations, we consider the problem of maximally down regulating protein production by altering the rates within this set of feasible rates. Under certain conditions on the feasible set, we show that an optimal solution can be determined efficiently. We also rigorously analyze two special cases of the down regulation optimization problem. Our results suggest that one must focus on the position along the mRNA molecule where the transition rate has the strongest effect on the protein production rate. However, this rate is not necessarily the slowest transition rate along the mRNA molecule. We discuss some of the biological implications of these results.
| S-EPMC5264618 | biostudies-literature
Project description:Passage through mitosis is driven by precisely-timed changes in transcriptional regulation and protein degradation. However, the importance of translational regulation during mitosis remains poorly understood. Here, using ribosome profiling, we find both a global translational repression and identified ~200 mRNAs that undergo specific translational regulation at mitotic entry. In contrast, few changes in mRNA abundance are observed, indicating that regulation of translation is the primary mechanism of modulating protein expression during mitosis. Interestingly, 91% of the mRNAs that undergo gene-specific regulation in mitosis are translationally repressed, rather than activated. One of the most pronounced translationally-repressed genes is Emi1, an inhibitor of the anaphase promoting complex (APC) which is degraded during mitosis. We show that full APC activation requires translational repression of Emi1 in addition to its degradation. These results identify gene-specific translational repression as a means of controlling the mitotic proteome, which may complement post-translational mechanisms for inactivating protein function.
| S-EPMC4548207 | biostudies-literature
Project description:In eukaryotic cells, protein synthesis is compartmentalized; mRNAs encoding secretory/membrane proteins are translated on endoplasmic reticulum (ER)-bound ribosomes, whereas mRNAs encoding cytosolic proteins are translated on free ribosomes. mRNA partitioning between the two compartments occurs via positive selection: free ribosomes engaged in the translation of signal sequence-encoding mRNAs are trafficked from the cytosol to the ER. After translation termination, ER-bound ribosomes are thought to dissociate, thereby completing a cycle of mRNA partitioning. At present, the physiological basis for termination-coupled ribosome release is unknown. To gain insight into this process, we examined ribosome and mRNA partitioning during the unfolded protein response, key elements of which include suppression of the initiation stage of protein synthesis and polyribosome breakdown. We report that unfolded protein response (UPR)-elicited polyribosome breakdown resulted in the continued association, rather than release, of ER-bound ribosomes. Under these conditions, mRNA translation in the cytosol was suppressed, whereas mRNA translation on the ER was sustained. Furthermore, mRNAs encoding key soluble stress proteins (XBP-1 and ATF-4) were translated primarily on ER-bound ribosomes. These studies demonstrate that ribosome release from the ER is termination independent and identify new and unexpected roles for the ER compartment in the translational response to induction of the unfolded protein response.
| S-EPMC1289424 | biostudies-literature
Project description:DDX3 subfamily DEAD-box RNA helicases are essential developmental regulators of RNA metabolism in eukaryotes. belle, the single DDX3 ortholog in Drosophila, is required for fly viability, fertility, and germline stem cell maintenance. Belle is involved both in translational activation and repression of target mRNAs in different tissues; however, direct targets of Belle in the testes are essentially unknown. Here we showed that belle RNAi knockdown in testis cyst cells caused a disruption of adhesion between germ and cyst cells and generation of tumor-like clusters of stem-like germ cells. Ectopic expression of β-integrin in cyst cells rescued early stages of spermatogenesis in belle knockdown testes, indicating that integrin adhesion complexes are required for the interaction between somatic and germ cells in a cyst. To address Belle functions in spermatogenesis in detail we performed cross-linking immunoprecipitation and sequencing (CLIP-seq) analysis and identified multiple mRNAs that interacted with Belle in the testes. The set of Belle targets includes transcripts of proteins that are essential for preventing the tumor-like clusters of germ cells and for sustaining spermatogenesis. By our hypothesis, failures in the translation of a number of mRNA targets additively contribute to developmental defects observed in the testes with belle knockdowns both in cyst cells and in the germline.
| S-EPMC7140462 | biostudies-literature
Project description:mRNA translation is the most energy consuming process in the cell. In addition, it plays a pivotal role in the control of gene expression and is therefore tightly regulated. In response to various extracellular stimuli and intracellular cues, signaling pathways induce quantitative and qualitative changes in mRNA translation by modulating the phosphorylation status and thus the activity of components of the translational machinery. In this work we focus on the phosphoinositide 3-kinase (PI3K)/AKT and the mitogen-activated protein kinase (MAPK) pathways, as they are strongly implicated in the regulation of translation in homeostasis, whereas their malfunction has been linked to aberrant translation in human diseases, including cancer.
| S-EPMC3536343 | biostudies-literature
Project description:Optogenetic tools have revolutionized the study of receptor-mediated processes, but such tools are lacking for RNA-controlled systems. In particular, light-activated regulatory RNAs are needed for spatiotemporal control of gene expression. To fill this gap, we used in vitro selection to isolate a novel riboswitch that selectively binds the trans isoform of a stiff-stilbene (amino-tSS)-a rapidly and reversibly photoisomerizing small molecule. Structural probing revealed that the RNA binds amino-tSS about 100-times stronger than the cis photoisoform (amino-cSS). In vitro and in vivo functional analysis showed that the riboswitch, termed Werewolf-1 (Were-1), inhibits translation of a downstream open reading frame when bound to amino-tSS. Photoisomerization of the ligand with a sub-millisecond pulse of light induced the protein expression. In contrast, amino-cSS supported protein expression, which was inhibited upon photoisomerization to amino-tSS. Reversible photoregulation of gene expression using a genetically encoded RNA will likely facilitate high-resolution spatiotemporal analysis of complex RNA processes.
| S-EPMC7051177 | biostudies-literature
Project description:The regulation of translation initiation factor 2 (eIF2) is important for erythroid survival and differentiation. Lack of iron, a critical component of heme and hemoglobin, activates Heme Regulated Inhibitor (HRI). This results in phosphorylation of eIF2 and reduced eIF2 availability, which inhibits protein synthesis. Translation of specific transcripts such as Atf4, however, is enhanced. Upstream open reading frames (uORFs) are key to this regulation. The aim of this study is to investigate how tunicamycin treatment, that induces eIF2 phosphorylation, affects mRNA translation in erythroblasts. Ribosome profiling combined with RNA sequencing was used to determine translation initiation sites and ribosome density on individual transcripts. Treatment of erythroblasts with Tunicamycin (Tm) increased phosphorylation of eIF2 2-fold. At a false discovery rate of 1%, ribosome density was increased for 147 transcripts, among which transcriptional regulators such as Atf4, Tis7/Ifrd1, Pnrc2, Gtf2h, Mbd3, JunB and Kmt2e. Translation of 337 transcripts decreased more than average, among which Dym and Csde1. Ribosome profiling following Harringtonine treatment uncovered novel translation initiation sites and uORFs. Surprisingly, translated uORFs did not predict the sensitivity of transcripts to altered ribosome recruitment in presence or absence of Tm. The regulation of transcription and translation factors in reponse to eIF2 phosphorylation may explain the large overall response to iron deficiency in erythroblasts.
| S-EPMC5892948 | biostudies-literature
Project description:tRNA gene copy number is a primary determinant of tRNA abundance and therefore the rate at which each tRNA delivers amino acids to the ribosome during translation. Low-abundance tRNAs decode rare codons slowly, but it is unclear which genes might be subject to tRNA-mediated regulation of expression. Here, those mRNA targets were identified via global simulation of translation. In-silico mRNA translation rates were compared for each mRNA in both wild-type and a [Formula: see text] sup70-65 mutant, which exhibits a pseudohyphal growth phenotype and a 75% slower CAG codon translation rate. Of 4900 CAG-containing mRNAs, 300 showed significantly reduced in silico translation rates in a simulated tRNA mutant. Quantitative immunoassay confirmed that the reduced translation rates of sensitive mRNAs were [Formula: see text] concentration-dependent. Translation simulations showed that reduced [Formula: see text] concentrations triggered ribosome queues, which dissipated at reduced translation initiation rates. To validate this prediction experimentally, constitutive gcn2 kinase mutants were used to reduce in vivo translation initiation rates. This repaired the relative translational rate defect of target mRNAs in the sup70-65 background, and ameliorated sup70-65 pseudohyphal growth phenotypes. We thus validate global simulation of translation as a new tool to identify mRNA targets of tRNA-specific gene regulation.
| S-EPMC5100601 | biostudies-literature