Target-dependent RNA polymerase as universal platform for gene expression control in response to intracellular molecules
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ABSTRACT:
SUBMITTER: Komatsu S
PROVIDER: S-EPMC10656481 | biostudies-literature | 2023 Jan
REPOSITORIES: biostudies-literature
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Project description:BackgroundEukaryotes use RNA interference and RNA-based epigenetic regulation to control transposon activity. In the standard pathways of RNA-based transcriptional and post-transcriptional silencing the protein complex RNA-dependent RNA polymerase (RdRP) plays a crucial role. However, alternative pathways that bypass RdRP have recently been described. Hence two important questions are: is RdRP truly a necessary component for transposon control, and are the alternative RNA-based strategies also capable of controlling transposable elements?ResultsWe have studied the interplay between host RNAi pathways and transposons using mathematical models. We show that the canonical RdRP-based model controls transposons tightly, mainly via the feedback of cytoplasmic small RNA amplification. Next, we consider two variants lacking RdRP and instead employing antisense transcription of transposons. We show that transposon activity is also controlled by the alternative pathways, although cytoplasmic small RNA amplification is absent. Instead, control occurs in the nucleus, through a feedback in the epigenetic regulation.ConclusionsConcluding, our models show that the control of transposon activity can be achieved by alternative pathways that lack RdRP and act through different feedback mechanisms. Thus, although RdRP activity is ubiquitous in eukaryotes, it need not be a general requirement for transposon control.
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Project description:PurposeEbola virus (EBOV) is such kind of virus which is responsible for 23,825 cases and 9675 deaths worldwide only in 2014 and with an average diseases fatality rate between 25 % and 90 %. Although, medical technology has tried to handle the problems, there is no Food and Drug Administration (FDA)-approved therapeutics or vaccines available for the prevention, post exposure, or treatment of Ebola virus disease (EVD).MethodsIn the present study, we used the immunoinformatics approach to design a potential epitope-based vaccine against the RNA-dependent RNA polymerase-L of EBOV. BioEdit v7.2.3 sequence alignment editor, Jalview v2 and CLC Sequence Viewer v7.0.2 were used for the initial sequence analysis for securing the conservancy from the sequences. Later the Immune Epitope Database and Analysis Resource (IEDB-AR) was used for the identification of T-cell and B-cellepitopes associated with type I and II major histocompatibility complex molecules analysis. Finally, the population coverage analysis was employed.ResultsThe core epitope "FRYEFTAPF" was found to be the most potential one, with 100 % conservancy among all the strains of EBOV. It also interacted with both type I and II major histocompatibility complex molecules and is considered as nonallergenic in nature. Finally, with impressive cumulative population coverage of 99.87 % for the both MHC-I and MHC-II class throughout the world population was found for the proposed epitope.ConclusionTo end, the projected peptide gave us a solid stand to propose for vaccine consideration and that might be experimented for its potency in eliciting immunity through humoral and cell mediated immune responses in vitro and in vivo.
| S-EPMC4529428 | biostudies-literature
Project description:COVID-19 respiratory disease caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has rapidly become a global health issue since it emerged in December 2019. While great global efforts are underway to develop vaccines and to discover or repurpose therapeutic agents for this disease, as of this writing only the nucleoside drug remdesivir has been approved under Emergency Use Authorization to treat COVID-19. The RNA-dependent RNA polymerase (RdRP), a viral enzyme for viral RNA replication in host cells, is one of the most intriguing and promising drug targets for SARS-CoV-2 drug development. Because RdRP is a viral enzyme with no host cell homologs, selective SARS-CoV-2 RdRP inhibitors can be developed that have improved potency and fewer off-target effects against human host proteins and thus are safer and more effective therapeutics for treating COVID-19. This review focuses on biochemical enzyme and cell-based assays for RdRPs that could be used in high-throughput screening to discover new and repurposed drugs against SARS-CoV-2.
| S-EPMC7684788 | biostudies-literature
Project description:RNA-dependent RNA polymerase (RdRp), also called nsp12, is considered a promising but challenging drug target for inhibiting replication and hence, the growth of various RNA-viruses. In this report, a computational study is performed to offer insights on the binding of Remdesivir and Galidesivir with SARS-CoV2 RdRp with natural substrate, ATP, as the control. It was observed that Remdesivir and Galidesivir exhibited similar binding energies for their best docked poses, -6.6 kcal/mole and -6.2 kcal/mole, respectively. ATP also displayed comparative and strong binding free energy of -6.3 kcal/mole in the catalytic site of RdRp. However, their binding locations within the active site are distinct. Further, the interaction of catalytic site residues (Asp760, Asp761, and Asp618) with Remdesivir and Galidesivir is comprehensively examined. Conformational changes of RdRp and bound molecules are demonstrated using 100 ns explicit solvent simulation of the protein-ligand complex. Simulation suggests that Galidesivir binds at the non-catalytic location and its binding strength is relatively weaker than ATP and Remdesivir. Remdesivir also binds at the catalytic site and showed high potency to inhibit the function of RdRp. Binding of co-factor units nsp7 and nsp8 with RdRp (nsp12) complexed with Remdesivir and Galidesivir was also examined. MMPBSA binding energy for all three complexes has been computed across the 100 ns simulation trajectory. Overall, this study suggests, Remdesivir has anti-RdRp activity via binding at a catalytic site. In contrast, Galidesivir may not have direct anti-RdRp activity but it can induce a conformational change in the RNA polymerase.
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Project description: Not available
| S-EPMC2194811 | biostudies-literature
Project description:Eukaryotic nuclei contain three different types of RNA polymerases (RNAPs), each consisting of 12-18 different subunits. The evolutionarily highly conserved RNAP subunit RPB5 is shared by all three enzymes and therefore represents a key structural/functional component of all eukaryotic RNAPs. Here we present the crystal structure of the RPB5 subunit from Saccharomyces cerevisiae. The bipartite structure includes a eukaryote-specific N-terminal domain and a C-terminal domain resembling the archaeal RNAP subunit H. RPB5 has been implicated in direct protein-protein contacts with transcription factor IIB, one of the components of the RNAP(II) basal transcriptional machinery, and gene-specific activator proteins, such as the hepatitis B virus transactivator protein X. The experimentally mapped regions of RPB5 involved in these interactions correspond to distinct and surface-exposed alpha-helical structures.
| S-EPMC18598 | biostudies-literature
Project description:Here, we report on the discovery in Caenorhabditis nematodes of multiple vertically transmitted RNAs coding for putative RNA-dependent RNA polymerases. Their sequences share similarity to distinct RNA viruses, including bunyaviruses, narnaviruses, and sobemoviruses. The sequences are present exclusively as RNA and are not found in DNA form. The RNAs persist in progeny after bleach treatment of adult animals, indicating vertical transmission of the RNAs. We tested one of the infected strains for transmission to an uninfected strain and found that mating of infected animals with uninfected animals resulted in infected progeny. By in situ hybridization, we detected several of these RNAs in the cytoplasm of the male and female germline of the nematode host. The Caenorhabditis hosts were found defective in degrading exogenous double-stranded RNAs, which may explain retention of viral-like RNAs. Strikingly, one strain, QG551, harbored three distinct virus-like RNA elements. Specific patterns of small RNAs complementary to the different viral-like RNAs were observed, suggesting that the different RNAs are differentially recognized by the RNA interference (RNAi) machinery. While vertical transmission of viruses in the family Narnaviridae, which are known as capsidless viruses, has been described in fungi, these observations provide evidence that multicellular animal cells harbor similar viruses.
| S-EPMC6900638 | biostudies-literature
Project description:The coronavirus disease (COVID-19), a worldwide pandemic, is caused by the severe acute respiratory syndrome-corona virus-2 (SARS-CoV-2). At this moment in time, there are no specific therapeutics available to combat COVID-19. Drug repurposing and identification of naturally available bioactive molecules to target SARS-CoV-2 are among the key strategies to tackle the notorious virus. The enzyme RNA-dependent RNA polymerase (RdRp) performs a pivotal role in replicating the virus. RdRp is a prime target for Remdesivir and other nucleotides analog-based antiviral drugs. In this study, we showed three bioactive molecules from tea (epicatechin-3,5-di-O-gallate, epigallocatechin-3,5-di-O-gallate, and epigallocatechin-3,4-di-O-gallate) that showed better interaction with critical residues present at the catalytic center and the NTP entry channel of RdRp than antiviral drugs Remdesivir and Favipiravir. Our computational approach to identify these molecules included molecular docking studies, followed by robust molecular dynamics simulations. All the three molecules are readily available in tea and could be made accessible along with other medications to treat COVID-19 patients. However, these results require validation by further in vitro and in vivo studies.
| S-EPMC8200525 | biostudies-literature
Project description:Introduction: The current SARS-CoV-2 pandemic urgently demands for both prevention and treatment strategies. RNA-dependent RNA-polymerase (RdRp), which has no counterpart in human cells, is an excellent target for drug development. Given the time-consuming process of drug development, repurposing drugs approved for other indications or at least successfully tested in terms of safety and tolerability, is an attractive strategy to rapidly provide an effective medication for severe COVID-19 cases.Areas covered: The currently available data and upcominSg studies on RdRp which can be repurposed to halt SARS-CoV-2 replication, are reviewed.Expert opinion: Drug repurposing and design of novel compounds are proceeding in parallel to provide a quick response and new specific drugs, respectively. Notably, the proofreading SARS-CoV-2 exonuclease activity could limit the potential for drugs designed as immediate chain terminators and favor the development of compounds acting through delayed termination. While vaccination is awaited to curb the SARS-CoV-2 epidemic, even partially effective drugs from repurposing strategies can be of help to treat severe cases of disease. Considering the high conservation of RdRp among coronaviruses, an improved knowledge of its activity in vitro can provide useful information for drug development or drug repurposing to combat SARS-CoV-2 as well as future pandemics.
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Project description:Intracellular delivery of mRNA, DNA, and other large macromolecules into cells plays an essential role in an array of biological research and clinical therapies. However, current methods yield a wide variation in the amount of material delivered, as well as limitations on the cell types and cargoes possible. Here, we demonstrate quantitatively controlled delivery into a range of primary cells and cell lines with a tight dosage distribution using a nanostraw-electroporation system (NES). In NES, cells are cultured onto track-etched membranes with protruding nanostraws that connect to the fluidic environment beneath the membrane. The tight cell-nanostraw interface focuses applied electric fields to the cell membrane, enabling low-voltage and nondamaging local poration of the cell membrane. Concurrently, the field electrophoretically injects biomolecular cargoes through the nanostraws and into the cell at the same location. We show that the amount of material delivered is precisely controlled by the applied voltage, delivery duration, and reagent concentration. NES is highly effective even for primary cell types or different cell densities, is largely cargo agnostic, and can simultaneously deliver specific ratios of different molecules. Using a simple cell culture well format, the NES delivers into >100,000 cells within 20 s with >95% cell viability, enabling facile, dosage-controlled intracellular delivery for a wide variety of biological applications.
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