Identification of Interactions between Sindbis Virus Capsid Protein and Cytoplasmic vRNA as Novel Virulence Determinants
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ABSTRACT:
PROVIDER: PRJNA389816 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5664292.fastq.gz | Fastqsanger.gz | |||
| SRR5664293.fastq.gz | Fastqsanger.gz | |||
| SRR5664294.fastq.gz | Fastqsanger.gz | |||
| SRR5664295.fastq.gz | Fastqsanger.gz | |||
| SRR5664296.fastq.gz | Fastqsanger.gz |
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Project description:Alphaviruses are arthropod-borne viruses that represent a significant threat to public health at a global level. While the formation of alphaviral nucleocapsid cores, consisting of cargo nucleic acid and the viral capsid protein, is an essential molecular process of infection, the precise interactions between the two partners are ill-defined. A CLIP-seq approach was used to screen for candidate sites of interaction between the viral Capsid protein and genomic RNA of Sindbis virus (SINV), a model alphavirus. The data presented in this report indicates that the SINV capsid protein binds to specific viral RNA sequences in the cytoplasm of infected cells, but its interaction with genomic RNA in mature extracellular viral particles is largely non-specific in terms of nucleotide sequence. Mutational analyses of the cytoplasmic viral RNA-capsid interaction sites revealed a functional role for capsid binding early in infection. Interaction site mutants exhibited decreased viral growth kinetics; however, this defect was not a function of decreased particle production. Rather mutation of the cytoplasmic capsid-RNA interaction sites negatively affected the functional capacity of the incoming viral genomic RNAs leading to decreased infectivity. Furthermore, cytoplasmic capsid interaction site mutants are attenuated in a murine model of neurotropic alphavirus infection. Collectively, the findings of this study indicate that the identified cytoplasmic interactions of the viral capsid protein and genomic RNA, while not essential for particle formation, are necessary for genomic RNA function early during infection. This previously unappreciated role of capsid protein during the alphaviral replication cycle also constitutes a novel virulence determinant.
2017-06-19 | GSE99879 | GEO
Project description:Alphaviruses are arthropod-borne viruses that represent a significant threat to public health at a global level. While the formation of alphaviral nucleocapsid cores, consisting of cargo nucleic acid and the viral capsid protein, is an essential molecular process of infection, the precise interactions between the two partners are ill-defined. A CLIP-seq approach was used to screen for candidate sites of interaction between the viral Capsid protein and genomic RNA of Sindbis virus (SINV), a model alphavirus. The data presented in this report indicates that the SINV capsid protein binds to specific viral RNA sequences in the cytoplasm of infected cells, but its interaction with genomic RNA in mature extracellular viral particles is largely non-specific in terms of nucleotide sequence. Mutational analyses of the cytoplasmic viral RNA-capsid interaction sites revealed a functional role for capsid binding early in infection. Interaction site mutants exhibited decreased viral growth kinetics; however, this defect was not a function of decreased particle production. Rather mutation of the cytoplasmic capsid-RNA interaction sites negatively affected the functional capacity of the incoming viral genomic RNAs leading to decreased infectivity. Furthermore, cytoplasmic capsid interaction site mutants are attenuated in a murine model of neurotropic alphavirus infection. Collectively, the findings of this study indicate that the identified cytoplasmic interactions of the viral capsid protein and genomic RNA, while not essential for particle formation, are necessary for genomic RNA function early during infection. This previously unappreciated role of capsid protein during the alphaviral replication cycle also constitutes a novel virulence determinant.
| S-EPMC5507600 | biostudies-literature
Project description:Microarray analysis comparing cells that are resistant to Sindbis virus-induced cell death (clones 9, 43) versus cells that are sensitive to Sindbis virus-induced cell death (WT293) Keywords = Sindbis alphavirus functinal phenotype Keywords: repeat sample
2004-05-01 | GSE1264 | GEO
Project description:Arthropod-borne viruses, such as the members of genus Alphavirus, are a significant concern to global public health. As obligate intracellular pathogens, RNA viruses must interact with the host cell machinery to establish, and complete, their viral lifecycles. Despite considerable efforts to define the host/pathogen interactions essential for alphaviral replication, an unbiased and inclusive assessment of alphaviral RNA:protein interactions has not been undertaken. Moreover, the biological and molecular importance of these interactions, in the full context of their molecular function as RNA-binding proteins, has not been fully realized. The data presented here introduces a robust viral RNA:protein discovery method to elucidate the Sindbis virus (SINV) RNA:Protein host interface. Cross-Link Assisted mRNP Purification (CLAMP) assessment reveals an extensive array of host/pathogen interactions centered on the viral RNAs (vRNAs). After prioritization of the host proteins associated with the vRNAs, we identified the site of Protein:vRNA interaction via a CLIP-seq approach and assessed the consequences of the RNA:protein binding event of hnRNP K, hnRNP I, and hnRNP M in regards to viral infection. Herein we demonstrate that mutation of the prioritized hnRNP:vRNA interaction sites effectively disrupted the hnRNP:vRNA interaction. Correlating with disrupted hnRNP:vRNA binding, SINV growth kinetics were reduced relative to wild type parental viral infections in a vertebrate and invertebrate tissue culture models of infection. The molecular mechanism leading to reduced viral growth kinetics was found to be reduced vRNA accumulation and dysregulated structural gene expression. Collectively, this study further defines the scope and importance of the alphavirus host/pathogen vRNA:protein interactions.
2018-01-02 | GSE103693 | GEO
Project description:The re-emerged Zika virus (ZIKV) can cause severe neurological complications termed congenital Zika syndrome (CZS) in infants born to ZIKV-infected mothers. However, increasing evidence shows the ancestral African lineage ZIKV displays more virulent phenotype than the contemporary Asian ZIKV in multiple cell and animal models. However, the viral determinants and underlying mechanism of this lineage specific virulence phenotype remain largely unknown. In the present study, phylogenetic analysis and sequence alignment identified a panel of lineage specific amino acid substitutions between African and Asian ZIKVs. Then, these amino acid substitutions were introduced into an infectious clone of Asian ZIKV by standard reverse genetic technology. Further characterization in vitro demonstrated that the recovered mutant virus with a lysine to arginine substitution at position 101 of capsid (C) protein (termed K101R) produced larger plaques in BHK-21 cells, and replicated more efficiently and induced more severe cytopathic effects (CPE) in multiple cells including human neuronal precursor cells (NPCs). More importantly, the K101R mutant virus replicated more efficiently in mouse brain tissues, and induced stronger inflammatory responses, leading to higher mortality than the wild type (WT) virus in neonatal mice. Despite no effect on RNA binding affinity of recombinant C protein to viral RNA, structural modeling and biochemical assays showed that the K101R substitution increased the viral protease cleavage efficiency of full C protein. Taken together, our study identified a single amino acid substitution K101R in the capsid protein responsible for the well-characterized virulence enhancement phenotypes, highlighting the importance of viral determinants for the co-circulating of two lineages of ZIKV.
2023-09-01 | GSE226495 | GEO
Project description:Microarray analysis comparing cells that are resistant to Sindbis virus-induced cell death (clones 9, 43) versus cells that are sensitive to Sindbis virus-induced cell death (WT293)
2007-08-03 | E-GEOD-1264 | biostudies-arrayexpress
Project description:Sindbis virus structural polyprotein -1 PRF deep mutational scan
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Project description:Brief assessment of VEEV TC-83 Capsid:RNA interactions.
2020-05-25 | GSE132125 | GEO
Project description:Viral vectors are attractive tools to express genes in neurons. Transduction of neurons with a recombinant, replication-deficient Sindbis viral vector is a method of choice for studying the effects of short-term protein overexpression on neuronal function. However, to which extent Sindbis by itself may affect neurons is not fully understood. We assessed effects of neuronal transduction with a Sindbis viral vector on the transcriptome and proteome in organotypic hippocampal slice cultures, and analyzed the electrophysiological properties of individual CA1 neurons, at 24h and 72h after viral vector injection. Whereas Sindbis caused substantial gene expression alterations, changes at the protein level were less pronounced. Alterations in transcriptome and proteome were predominantly limited to proteins involved in mediating anti-viral innate immune responses. Sindbis transduction did not affect the electrophysiological properties of individual neurons: the membrane potential, excitability and synaptic currents were similar between transduced and nontransduced CA1 neurons up to 72h after Sindbis injection. We conclude that Sindbis viral vectors are suitable for studying short-term effects of a protein of interest on electrophysiological properties of neurons, but not for studies on the regulation of gene expression.
2019-08-26 | PXD013634 | Pride