RNA to rule them all: Critical steps in Lassa virus ribonucleoparticle assembly and recruitment
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q-Tof Ultima, Q Exactive HF
ORGANISM(S): Lassa Mammarenavirus
SUBMITTER: 
Lennart Sänger
LAB HEAD: Charlotte Uetrecht
PROVIDER: PXD040186 | Pride | 2023-12-18
REPOSITORIES: Pride
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| HDX_peptide_pool.xlsx | Xlsx | |||
| mass_table.xlsx | Xlsx | |||
| raw_data_spectra.zip | Other |
Items per page: 5 1 - 3 of 3 |
Similar Datasets
Project description:At the plasma membrane of mammalian cells, major histocompatibility complex class I molecules (MHC-I) present antigenic peptides to cytotoxic T cells. Following the loss of the peptide and the light chain beta-2 microglobulin (beta2m), the resulting free heavy chains (FHCs) can associate into homotypic complexes in the plasma membrane. Here, we investigate the stoichiometry and dynamics of MHC-I FHCs assemblies by combining a micropattern assay with fluorescence recovery after photobleaching (FRAP) and with single molecule co-tracking. We identify non-covalent MHC-I FHC dimers mediated by the alpha-3 domain as the prevalent species at the plasma membrane, leading a moderate decrease in the diffusion coefficient. MHC-I FHC dimers show increased tendency to cluster into higher order oligomers as concluded from an increased immobile fraction with higher single molecule colocalization. In vitro studies with isolated proteins in conjunction with molecular docking and dynamics simulations suggest that in the complexes, the alpha-3 domain of one FHC binds to another FHC in a manner similar to the beta2m light chain.
2022-05-11 | PXD033485 | Pride
Project description:Heldt2012 - Influenza Virus Replication
The model describes the life cycle of influenza A virus in a mammalian cell including the following steps: attachment of parental virions to the cell membrane, receptor-mediated endocytosis, fusion of the virus envelope with the endosomal membrane, nuclear import of vRNPs, viral transcription and replication, translation of the structural viral proteins, nuclear export of progeny vRNPs and budding of new virions. It also explicitly accounts for the stabilization of cRNA by viral polymerases and NP and the inhibition of vRNP activity by M1 protein binding. In short, the model focuses on the molecular mechanism that controls viral transcription and replication.
This model is described in the article:
Modeling the intracellular dynamics of influenza virus replication to understand the control of viral RNA synthesis.
Heldt FS, Frensing T, Reichl U.
J Virol.
Abstract:
Influenza viruses transcribe and replicate their negative-sense RNA genome inside the nucleus of host cells via three viral RNA species. In the course of an infection, these RNAs show distinct dynamics, suggesting that differential regulation takes place. To investigate this regulation in a systematic way, we developed a mathematical model of influenza virus infection at the level of a single mammalian cell. It accounts for key steps of the viral life cycle, from virus entry to progeny virion release, while focusing in particular on the molecular mechanisms that control viral transcription and replication. We therefore explicitly consider the nuclear export of viral genome copies (vRNPs) and a recent hypothesis proposing that replicative intermediates (cRNA) are stabilized by the viral polymerase complex and the nucleoprotein (NP). Together, both mechanisms allow the model to capture a variety of published data sets at an unprecedented level of detail. Our findings provide theoretical support for an early regulation of replication by cRNA stabilization. However, they also suggest that the matrix protein 1 (M1) controls viral RNA levels in the late phase of infection as part of its role during the nuclear export of viral genome copies. Moreover, simulations show an accumulation of viral proteins and RNA toward the end of infection, indicating that transport processes or budding limits virion release. Thus, our mathematical model provides an ideal platform for a systematic and quantitative evaluation of influenza virus replication and its complex regulation.
With the current parameter set, the model reproduces an infection at a multiplicity of infection (MOI) of 10. Figure 2A of the paper is reproduced here, with parameters kDegRnp and kSynP changed to zeros.
Initial conditions and parameter changes that were used to obtain specific figures in the article can be found in Table A2.
The model has the correct value for kAttLo as 4.55e-04. The value of this parameter mentioned as 4.55e-02 in Table 1 of the paper is incorrect. This is checked with the author.
This model is hosted on BioModels Database
and identified
by: MODEL1307270000
.
To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource
for published quantitative kinetic models
.
To the extent possible under law, all copyright and related or
neighbouring rights to this encoded model have been dedicated to the public
domain worldwide. Please refer to CC0 Public Domain
Dedication
for more information.
2024-09-02 | BIOMD0000000463 | BioModels
Project description:Negative-sense RNA viruses (NSVs) rely on prepackaged viral RNA-dependent RNA polymerases (RdRp) to replicate and transcribe their viral genomes. Their replication machinery consists of an RdRp bound to viral RNA which is wound around a nucleoprotein (NP) scaffold, forming a viral ribonucleoprotein complex. NSV NP is known to regulate transcription and replication of genomic RNA, however its role in maintaining and protecting the viral genetic material is unknown. Here, we exploited host microRNA expression to target NP of influenza A virus and Sendai virus to ascertain how this would impact genomic levels and the host response to infection. We find that in addition to inducing a drastic decrease in genome replication, the antiviral host response in the absence of NP is dramatically enhanced. Additionally, our data shows that insufficient levels of NP prevent the replication machinery of these NSVs to process full-length genomes, resulting in aberrant replication products which form pathogen-associated molecular patterns in the process. These dynamics facilitate immune recognition by cellular pattern recognition receptors leading to a strong host antiviral response. Moreover, we observe that the consequences of limiting NP levels are universal amongst NSVs including Ebola virus, Lassa virus and Measles virus. Overall, these results provide new insights into viral genome replication of negative-sense RNA viruses and highlight novel avenues towards developing effective antiviral strategies, adjuvants, and/or live-attenuated vaccines.
2021-02-02 | GSE165340 | GEO
Project description:An essential element of adaptive immunity is the selective binding of peptide antigens by major histocompatibility complex (MHC) class I proteins and their presentation to cytotoxic T lymphocytes on the cell surface. Using native mass spectrometry, we here analyze the binding of peptides to an empty disulfide-stabilized HLA-A*02:01 molecule. This novel approach allows us to examine the binding properties of diverse peptides. The unique stability of our MHC class I even enables us to determine the binding affinity of complexes, which are suboptimally loaded with truncated or charge-reduced peptides. Notably, a unique erucamide adduct decouples affinity analysis from peptide identity alleviating issues usually attributed to clustering. We discovered that two anchor positions at the binding surface between MHC and peptide can be stabilized independently and further analyze the contribution of other peptidic amino acids on the binding. We propose this as an alternative, likely universally applicable method to artificial prediction tools to estimate the binding strength of peptides to MHC class I complexes quickly and efficiently. This newly described MHC class I-peptide binding affinity quantitation represents a much needed orthogonal, confirmatory approach to existing computational affinity predictions and has the potential to eliminate binding affinity biases and thus accelerate drug discovery in infectious diseases autoimmunity, vaccine design, and cancer immunotherapy.
2022-05-31 | PXD027725 | Pride
Project description:The exploration of scale-down models to imitate the influence of large scale bioreactor inhomogeneities on cellular metabolism is a topic with increasing relevance. While gradients of substrates, pH, or dissolved oxygen are often investigated, oscillating CO2/HCO3- levels, a typical scenario in large industrial bioreactors, is rarely addressed. Hereby, we investigate the metabolic and transcriptional response in Corynebacterium glutamicum wild type as well as the impact on L-lysine production in a model strain exposed to pCO2 gradients of (75-315) mbar. A novel three-compartment cascade bioreactor system was developed and characterized that offers high flexibility for installing gradients and residence times to mimic industrial-relevant conditions, and provides the potential of accurate carbon balancing. The phenomenological analysis of cascade fermentations imposed to the pCO2 gradients at industry-relevant residence times of about 3.55 min did not significantly impair the process performance, with growth and product formation being similar to control conditions. However, transcriptional analysis disclosed up to 66 differentially expressed genes already after 3.55min under stimulus exposure, with the overall change in gene expression directly correlateable to the pCO2 gradient intensity and the residence time of the cells.
2014-05-15 | E-MTAB-2585 | biostudies-arrayexpress
Project description:Both inhalation of environmental (nano)particles (NP) and persistent herpesvirus-infection have been implicated to contribute to the development of chronic lung disease. We hypothesized that the combination of NP exposure and virus infection leads to a different outcome than each factor alone. To test this hypothesis, we applied different carbonaceous NP in the model system of murine gammaherpesvirus 68 (MHV-68), which serves as a small animal model for gammaherpesvirus pathogenesis in vitro and in vivo. NP exposure of latently infected cells induced lytic virus production, while virus growth after de novo infection was not affected. To investigate the effect of pulmonary NP exposure in vivo, latently infected mice were instilled with carbonaceous NP or left untreated. After 24 hours, lung tissue was harvested for analysis. Immunohistochemistry demonstrated an increase in lytic viral proteins after exposure to NP. Simultaneously, gene expression analysis revealed a signature with considerable parallels to the one seen during acute infection. Analysis of the metabolome demonstrated an enrichment of phospholipids in latently infected mice after short-time exposure to NP, matching profoundly with the pattern observed during acute virus infection. Furthermore, the effect of NP exposure on virus reactivation could also be observed in human cells latently infected with Epstein-Barr-Virus (EBV). Our results indicate that the combination of NP exposure and persistent herpesvirus infection restores a molecular signature found in acute virus infection, boosts production of lytic viral proteins, and induces an inflammatory response in the lung – a pattern which might finally result in tissue damage and fibrotic alterations.
2016-12-19 | GSE79501 | GEO
Project description:Background: Negative-pressure wound therapy has become widely available in modern chronic wound cares. Accelerated keratinocyte (HaCaT cell) movements and decreased E-cadherin expressions induced by the applied negative pressure gradient of 125 mmHg (NP) have been reported in previous studies. However, the molecular mechanism for E-cadherin regulations under NP remains unexplored. We highlighted the signal transduction involved in NP-induced E-cadherin regulation for keratinocytes in the study. Results: Microarray results showed that catenin 1 (CTNND1) gene, the gene encoding the p120-catenin (p120ctn) in cell-cell junctions, was significantly (p = 0.0005) upregulated in HaCaT cells under NP for 12 hours in comparison with those at ambient pressure (AP). Cell fractionations and immunoblotting data showed that NP increased p120ctn phosphorylation, and resulted in p120ctn translocation from the plasma membrane to the cytoplasm. Fluorescent stains suggested that NP diminished the co-localization of p120ctn and E-cadherin on the plasma membrane. Similar phenomenon was also observed in the cells with overexpressed p120ctn at NP. Interestingly, overexpression of dN- p120ctn, a deletion mutant of p120ctn without the N-terminal phosphorylation sites, restored the adherens junctions (AJ) at NP. Knockdown of p120ctn with lentiviral small hairpin RNA not only diminished E-cadherin expressions but also accelerated cell movement at AP. Conclusions: Phosphorylation of p120ctn is endowed to respond rapidly to NP and contributes to the AJ disassembly. This NP-induced E-cadherin downregulation can possibly accelerate wound-healing process. Conclusions: Phosphorylation of p120ctn is endowed to respond rapidly to NP and contributes to the AJ disassembly. This NP-induced E-cadherin downregulation can possibly accelerate wound-healing process. HaCaT cells under negative pressure (NP) gradient of 125 mmHg for 12 hours in comparison with those at ambient pressure (AP) were tested for RNA extraction and hybridization on Affymetrix microarrays. The experiments have been biological replicated three times. The differential expression gene between NP and AP were selected and validated with qPCR method.
2015-12-31 | E-GEOD-70512 | biostudies-arrayexpress
Project description:Clathrin, forming the triskelion network, orchestrates highly regulated cellular processes facilitating cargo internalization and trafficking in eukaryotes, with its N-terminal domain (NTD) pivotal for adaptor protein (AP) interactions. The NTD contains up to four AP-binding sites, and their roles in preferential occupancy by APs have not been addressed. Here we present Native MS data showing competition between peptides from Adaptor proteins Ent1 and Ent2.
2024-11-05 | PXD052864 | Pride
Project description:Recent technical advances have resulted in a significant increase in our understanding of the RNA-binding protein (RBP) repertoire present within eukaryotic cells. A particular focus has been on the RBPs that interact with cellular polyadenylated mRNAs. However, recent studies utilising the same technologies have begun to tease apart the RBP interactome of viral mRNAs, most notably SARS-CoV-2, demonstrating clear similarities but also differences between the RBP profiles of viral and cellular mRNAs. Herein, for the first time we comprehensively mapped the RBPs that associate with the NP mRNA of an influenza A virus. Moreover, we provide evidence that the viral polymerase is essential for the recruitment of RPBs to viral mRNAs through direct polymerase-RBP interactions during transcription. We show that loss of TDP-43, that associates with the viral mRNA, results in a decrease in viral mRNA availability within infected cells, with an impact on the accumulation of viral genomic RNA and the yield of infectious viral particles. Overall, we uncover an important role for TDP-43 in the influenza A virus replication cycle via a direct interaction with viral mRNAs and we demonstrate an essential role of the viral polymerase in orchestrating the assembly of viral mRNPs.
2024-04-16 | PXD049316 | Pride
Project description:Recent technical advances have resulted in a significant increase in our understanding of the RNA-binding protein (RBP) repertoire present within eukaryotic cells. A particular focus has been on the RBPs that interact with cellular polyadenylated mRNAs. However, recent studies utilising the same technologies have begun to tease apart the RBP interactome of viral mRNAs, most notably SARS-CoV-2, demonstrating clear similarities but also differences between the RBP profiles of viral and cellular mRNAs. Herein, for the first time we comprehensively mapped the RBPs that associate with the NP mRNA of an influenza A virus. Moreover, we provide evidence that the viral polymerase is essential for the recruitment of RPBs to viral mRNAs through direct polymerase-RBP interactions during transcription. We show that loss of TDP-43, that associates with the viral mRNA, results in a decrease in viral mRNA availability within infected cells, with an impact on the accumulation of viral genomic RNA and the yield of infectious viral particles. Overall, we uncover an important role for TDP-43 in the influenza A virus replication cycle via a direct interaction with viral mRNAs and we demonstrate an essential role of the viral polymerase in orchestrating the assembly of viral mRNPs.
2024-04-16 | PXD040783 | Pride