Oxidative stress influences positive strand RNA virus genome synthesis and capping

Gullberg, Rebekah C.; Steel, J. Jordan; Moon, Stephanie L.; Soltani, Elnaz; Geiss, Brian J.

doi:10.1016/j.virol.2014.10.037

Cited by 94 publications

(78 citation statements)

References 56 publications

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“…The increased expression of the ISE6 proteins observed on infection with LGTV may reflect a generalized cellular response or metabolic processes and products used by the virus [4]. Flaviviruses are thought to exploit the cellular stress response to aid replication [51] and oxidative stress is thought to aid the replication of positive-strand RNA viruses [52]. Further, it has been proposed that the balance between antioxidant responses maintains an anti-apoptotic environment during flavivirus infection of the cell [53, 54], presumably facilitating virus replication and transmission.…”

Section: Discussionmentioning

confidence: 99%

RNAi reveals proteins for metabolism and protein processing associated with Langat virus infection in Ixodes scapularis (black-legged tick) ISE6 cells

et al. 2017

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BackgroundTick-borne flaviviruses (TBFs) cause thousands of human cases of encephalitis worldwide each year, with some TBF infections progressing to hemorrhagic fever. TBFs are of medical and veterinary importance and strategies to reduce flavivirus transmission by the tick vector may have significant application. Analyses of the proteome of ISE6 cells derived from the black legged tick, Ixodes scapularis infected with the TBF, Langat virus (LGTV), have provided insights into proteins and cellular processes involved with LGTV infection.MethodsRNA interference (RNAi)-induced knockdown of transcripts was used to investigate the role of ten tick proteins in the LGTV infection cycle in ISE6 cells. LGTV-infected cells were separately transfected with dsRNA corresponding to each gene of interest and the effect on LGTV genome replication and release of infectious virus was assessed by RT-qPCR and plaque assays, respectively.ResultsRNAi-induced knockdown of transcripts for two enzymes that likely function in amino acid, carbohydrate, lipid, terpenoid/polykeytide and vitamin metabolism, and a transcript for one protein of unknown function were associated with decreased replication of the LGTV genome and release of infectious virus from cells. The knockdown of transcripts for five enzymes predicted to function in metabolism, a protein likely associated with folding, sorting and degradation, and a protein of unknown function was associated with a decrease only in the amount of infectious LGTV released from cells.ConclusionsThese data suggest tick proteins potentially associated with metabolism and protein processing may be involved in LGTV infection of ISE6 cells. Our study provides information to begin to elucidate the function of these proteins and identify targets for the development of new interventions aimed at controlling the transmission of TBFs.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1944-0) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

RNAi reveals proteins for metabolism and protein processing associated with Langat virus infection in Ixodes scapularis (black-legged tick) ISE6 cells

et al. 2017

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“…In the case of flaviviruses, it was shown that the guanylyltransferase activity of NS5 encoded polymerase was enhanced by oxidative conditions. 61 It remains to be determined if there are specific aspects of mitochondrial dysfunction that may be controlled by antioxidant treatment, which may have important implications in protecting terminally differentiated neurons in the infected host. Our follow up hypothesis was that mitochondrial fission may contribute to the ensuing apoptosis in infected cells.…”

Section: Discussionmentioning

confidence: 99%

Altered mitochondrial dynamics as a consequence of Venezuelan Equine encephalitis virus infection

et al. 2017

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Mitochondria are sentinel organelles that are impacted by various forms of cellular stress, including viral infections. While signaling events associated with mitochondria, including those activated by pathogen associated molecular patterns (PAMPs), are widely studied, alterations in mitochondrial distribution and changes in mitochondrial dynamics are also beginning to be associated with cellular insult. Cells of neuronal origin have been demonstrated to display remarkable alterations in several instances, including neurodegenerative disorders. Venezuelan Equine Encephalitis Virus (VEEV) is a New World alphavirus that infects neuronal cells and contributes to an encephalitic phenotype. We demonstrate that upon infection by the vaccine strain of VEEV (TC-83), astrocytoma cells experience a robust drop in mitochondrial activity, which corresponds with an increased accumulation of reactive oxygen species (ROS) in an infection-dependent manner. Infection status also corresponds with a prominent perinuclear accumulation of mitochondria. Cellular enzymatic machinery, including PINK1 and Parkin, appears to be enriched in mitochondrial fractions as compared with uninfected cells, which is indicative of mitochondrial damage. Dynamin related protein 1 (Drp1), a protein that is associated with mitochondrial fission, demonstrated a modest enrichment in mitochondrial fractions of infected cells. Treatment with an inhibitor of mitochondrial fission, Mdivi-1, led to a decrease in caspase cleavage, suggesting that mitochondrial fission was likely to contribute to apoptosis of infected cells. Finally, our data demonstrate that mitophagy ensues in infected cells. In combination, our data suggest that VEEV infection results in significant changes in the mitochondrial landscape that may influence pathological outcomes in the infected cell.

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“…AdoHcy thus plays a crucial role in the interplay between the nsP1 functions, probably not by competition with AdoMet for the MTase inhibition but rather by inducing conformational changes favoring the GT reaction. During the preparation of the manuscript, Gullberg and coworkers showed that AdoMet and consequently AdoHcy are not necessary to produce the m 7 GMP-nsP1 product (58). However, the study was performed in the presence of GTP-ATTO 680, a GTP analog whose modification in position 8 could possibly affect the chemical properties of GTP and favor the guanylylation reaction.…”

Section: Discussionmentioning

confidence: 99%

mRNA Capping by Venezuelan Equine Encephalitis Virus nsP1: Functional Characterization and Implications for Antiviral Research

Guillén

Rabah

et al. 2015

J Virol

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Alphaviruses are known to possess a unique viral mRNA capping mechanism involving the viral nonstructural protein nsP1. This enzyme harbors methyltransferase (MTase) and nsP1 guanylylation (GT) activities catalyzing the transfer of the methyl group from S-adenosylmethionine (AdoMet) to the N7 position of a GTP molecule followed by the formation of an m 7 GMPnsP1 adduct. Subsequent transfer of m 7 GMP onto the 5= end of the viral mRNA has not been demonstrated in vitro yet. Here we report the biochemical characterization of Venezuelan equine encephalitis virus (VEEV) nsP1. We have developed enzymatic assays uncoupling the different reactions steps catalyzed by nsP1. The MTase and GT reaction activities were followed using a nonhydrolyzable GTP (GIDP) substrate and an original Western blot assay using anti-m 3 G/m 7 G-cap monoclonal antibody, respectively. The GT reaction is stimulated by S-adenosyl-L-homocysteine (Ado-Hcy), the product of the preceding MTase reaction, and metallic ions. The covalent linking between nsP1 and m 7 GMP involves a phosphamide bond between the nucleotide and a histidine residue. Final guanylyltransfer onto RNA was observed for the first time with an alphavirus nsP1 using a 5=-diphosphate RNA oligonucleotide whose sequence corresponds to the 5= end of the viral genome. Alanine scanning mutagenesis of residues H37, H45, D63, E118, Y285, D354, R365, N369, and N375 revealed their respective roles in MT and GT reactions. Finally, the inhibitory effects of sinefungin, aurintricarboxylic acid (ATA), and ribavirin triphosphate on MTase and capping reactions were investigated, providing possible avenues for antiviral research. IMPORTANCEEmergence or reemergence of alphaviruses represents a serious health concern, and the elucidation of their replication mechanisms is a prerequisite for the development of specific inhibitors targeting viral enzymes. In particular, alphaviruses are able, through an original reaction sequence, to add to their mRNA a cap required for their protection against cellular nucleases and initiation of viral proteins translation. In this study, the capping of a 5= diphosphate synthetic RNA mimicking the 5= end of an alphavirus mRNA was observed in vitro for the first time. The different steps for this capping are performed by the nonstructural protein 1 (nsP1). Reference compounds known to target the viral capping inhibited nsP1 enzymatic functions, highlighting the value of this enzyme in antiviral development. E mergence or reemergence of alphaviruses represents a serious health concern, as exemplified by the worldwide epidemics of Chikungunya virus during the last 10 years (1). The Alphavirus genus comprises 31 species that together with the genus Rubivirus forms the Togaviridae family. Alphaviruses have been classified on the basis of their geographical distribution. Alphaviruses circulating in the Old World (OW) most commonly cause febrile illness and painful arthralgias or polyarthralgias, particularly in the small joints (2). In contrast, New World (NW) alph...

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Oxidative stress influences positive strand RNA virus genome synthesis and capping

Cited by 94 publications

References 56 publications

RNAi reveals proteins for metabolism and protein processing associated with Langat virus infection in Ixodes scapularis (black-legged tick) ISE6 cells

RNAi reveals proteins for metabolism and protein processing associated with Langat virus infection in Ixodes scapularis (black-legged tick) ISE6 cells

Altered mitochondrial dynamics as a consequence of Venezuelan Equine encephalitis virus infection

mRNA Capping by Venezuelan Equine Encephalitis Virus nsP1: Functional Characterization and Implications for Antiviral Research

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