The Experts below are selected from a list of 204 Experts worldwide ranked by ideXlab platform
Varpu Marjomäki - One of the best experts on this subject based on the ideXlab platform.
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Human Enterovirus Group B Viruses Rely on Vimentin Dynamics for Efficient Processing of Viral Nonstructural Proteins.
Journal of Virology, 2020Co-Authors: Paula Turkki, Mira Laajala, Malin Flodström-tullberg, Varpu MarjomäkiAbstract:We report that several viruses from the human enterovirus group B cause massive vimentin rearrangements during lytic infection. Comprehensive studies suggested that Viral protein synthesis was triggering the vimentin rearrangements. Blocking the host cell vimentin dynamics with beta, beta'-iminodipropionitrile (IDPN) did not significantly affect the production of progeny viruses and only moderately lowered the synthesis of structural Proteins such as VP1. In contrast, the synthesis of the Nonstructural Proteins 2A, 3C, and 3D was drastically lowered. This led to attenuation of the cleavage of the host cell substrates PABP and G3BP1 and reduced caspase activation, leading to prolonged cell survival. Furthermore, the localization of the Proteins differed in the infected cells. Capsid protein VP1 was found diffusely around the cytoplasm, whereas 2A and 3D followed vimentin distribution. Based on protein blotting, smaller amounts of Nonstructural Proteins did not result from proteasomal degradation but from lower synthesis without intact vimentin cage structure. In contrast, inhibition of Hsp90 chaperone activity, which regulates P1 maturation, lowered the amount of VP1 but had less effect on 2A. The results suggest that the vimentin dynamics regulate Viral Nonstructural protein synthesis while having less effect on structural protein synthesis or overall infection efficiency. The results presented here shed new light on differential fate of structural and Nonstructural Proteins of enteroviruses, having consequences on host cell survival.IMPORTANCE A virus needs the host cell in order to replicate and produce new progeny viruses. For this, the virus takes over the host cell and modifies it to become a factory for Viral Proteins. Irrespective of the specific virus family, these Proteins can be divided into structural and Nonstructural Proteins. Structural Proteins are the building blocks for the new progeny virions, whereas the Nonstructural Proteins orchestrate the takeover of the host cell and its functions. Here, we have shown a mechanism that viruses exploit in order to regulate the host cell. We show that Viral protein synthesis induces vimentin cages, which promote production of specific Viral Proteins that eventually control apoptosis and host cell death. This study specifies vimentin as the key regulator of these events and indicates that Viral Proteins have different fates in the cells depending on their association with vimentin cages.
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human enterovirus group b viruses rely on vimentin dynamics for efficient processing of Viral Nonstructural Proteins
Journal of Virology, 2019Co-Authors: Paula Turkki, Mira Laajala, Malin Flodstromtullberg, Varpu MarjomäkiAbstract:: We report that several viruses from the human enterovirus group B cause massive vimentin rearrangements during lytic infection. Comprehensive studies suggested that Viral protein synthesis was triggering the vimentin rearrangements. Blocking the host cell vimentin dynamics with IDPN did not significantly affect the production of progeny viruses and only moderately lowered the synthesis of structural Proteins such as VP1. In contrast, the synthesis of the non-structural Proteins 2A, 3C, and 3D was drastically lowered. This led to attenuation of the cleavage of the host cell substrates PABP and G3BP1 and reduced caspase activation, thus leading to prolonged cell survival. Furthermore, the localization of the Proteins differed in the infected cells. Capsid protein VP1 was found diffusely around the cytoplasm, whereas 2A and 3D followed vimentin distribution. Based on protein blotting, lower amounts of non-structural Proteins did not result from proteasomal degradation, but from lower synthesis without intact vimentin cage structure. In contrast, inhibition of Hsp90 chaperone activity, which regulates P1 maturation, lowered the amount of VP1, but had less effect on 2A. The results suggest that, the vimentin dynamics regulate Viral non-structural protein synthesis while having no effect on structural protein synthesis or overall infection efficiency. The results presented here shed new light on differential fate of structural and non-structural Proteins of enteroviruses, having consequences on host cell survival.Importance A virus needs the host cell in order to replicate and produce new progeny viruses. For this, the virus takes over the host cell and modifies it to become a factory for Viral Proteins. Irrespective of the specific virus family, these Proteins can be divided into structural and non-structural Proteins. Structural Proteins are the building blocks for the new progeny virions, whereas the non-structural Proteins orchestrate the take-over of the host cell and its functions. Here we have shown a mechanism that viruses exploit in order to regulate the host cell. We show that Viral protein synthesis induces vimentin cages, which promote production of specific Viral Proteins that eventually control apoptosis and the host cell death. This study specifies vimentin as the key regulator of these events and indicates that Viral Proteins have different fates in the cells depending on their association with vimentin cages.
Ilya Frolov - One of the best experts on this subject based on the ideXlab platform.
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noncytopathic replication of venezuelan equine encephalitis virus and eastern equine encephalitis virus replicons in mammalian cells
Journal of Virology, 2005Co-Authors: Olga Petrakova, Slobodan Paessler, Evgeniya Volkova, Rodion Gorchakov, Richard M Kinney, Ilya FrolovAbstract:Venezuelan equine encephalitis (VEE) and eastern equine encephalitis (EEE) viruses are important, naturally emerging zoonotic viruses. They are significant human and equine pathogens which still pose a serious public health threat. Both VEE and EEE cause chronic infection in mosquitoes and persistent or chronic infection in mosquito-derived cell lines. In contrast, vertebrate hosts infected with either virus develop an acute infection with high-titer viremia and encephalitis, followed by host death or virus clearance by the immune system. Accordingly, EEE and VEE infection in vertebrate cell lines is highly cytopathic. To further understand the pathogenesis of alphaviruses on molecular and cellular levels, we designed EEE- and VEE-based replicons and investigated their replication and their ability to generate cytopathic effect (CPE) and to interfere with other Viral infections. VEE and EEE replicons appeared to be less cytopathic than Sindbis virus-based constructs that we designed in our previous research and readily established persistent replication in BHK-21 cells. VEE replicons required additional mutations in the 5′ untranslated region and nsP2 or nsP3 genes to further reduce cytopathicity and to become capable of persisting in cells with no defects in alpha/beta interferon production or signaling. The results indicated that alphaviruses strongly differ in virus-host cell interactions, and the ability to cause CPE in tissue culture does not necessarily correlate with pathogenesis and strongly depends on the sequence of Viral Nonstructural Proteins.
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sindbis virus expression vectors packaging of rna replicons by using defective helper rnas
Journal of Virology, 1993Co-Authors: Peter J. Bredenbeek, Charles M. Rice, Ilya Frolov, Sondra SchlesingerAbstract:: Since the recovery of infectious RNA transcripts from full-length cDNA clones, alphavirus genome RNAs have been engineered to allow expression of heterologous RNAs and Proteins. The highest levels of expression of heterologous products are achieved when the Viral structural genes are replaced by the heterologous coding sequences. Such recombinant RNAs are self-replicating (replicons) and can be introduced into cells as naked RNA, but they require trans complementation to be packaged and released from cells as infectious virion particles. In this report, we describe a series of defective Sindbis virus helper RNAs which can be used for packaging Sindbis virus RNA replicons. The defective helper RNAs contain the cis-acting sequences required for replication as well as the subgenomic RNA promoter which drives expression of the structural protein genes. In cells cotransfected with both the replicon and defective helper RNAs, Viral Nonstructural Proteins translated from the replicon RNA allow replication and transcription of the defective helper RNA to produce the virion structural Proteins. A series of defective helper RNAs were compared for the ability to package the replicon RNA as well as for the ability to be replicated and packaged. One defective helper RNA not only packaged the replicon but also was itself encapsidated and would be useful under conditions in which extensive amplification is advantageous. Other defective helper RNAs were able to package the replicon efficiently but were packaged very poorly themselves. These helpers should be useful for applications in which expression of the Viral structural Proteins or virus spread is not desired.
Donghoon Chung - One of the best experts on this subject based on the ideXlab platform.
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benzamidine ml336 inhibits plus and minus strand rna synthesis of venezuelan equine encephalitis virus without affecting host rna production
Antiviral Research, 2020Co-Authors: Andrew M Skidmore, Robert S Adcock, Colleen B Jonsson, Jennifer E Golden, Donghoon ChungAbstract:Abstract Venezuelan equine encephalitis virus (VEEV) is an alphavirus that is endemic to the Americas. VEEV outbreaks occur periodically and cause encephalitis in both humans and equids. There are currently no therapeutics or vaccines for treatment of VEEV in humans. Our group has previously reported on the development of a benzamidine VEEV inhibitor, ML336, which shows potent antiViral activity in both in vitro and in vivo models of infection. In cell culture experiments, ML336 inhibits Viral RNA synthesis when added 2–4 h post-infection, and mutations conferring resistance occur within the Viral Nonstructural Proteins (nsP2 and nsP4). We hypothesized that ML336 targets an activity of the Viral replicase complex and inhibits Viral RNA synthesis. To test this hypothesis, we employed various biochemical and cellular assays. Using structural analogues of ML336, we demonstrate that the cellular antiViral activity of these compounds correlates with their inhibition of Viral RNA synthesis. For instance, the IC50 of ML336 for VEEV RNA synthesis inhibition was determined as 1.1 nM, indicating potent anti-RNA synthesis activity in the low nanomolar range. While ML336 efficiently inhibited VEEV RNA synthesis, a much weaker effect was observed against the Old World alphavirus Chikungunya virus (IC50 > 4 μM), agreeing with previous data from a cell based assay. Using a tritium incorporation assay, we demonstrated that there was no significant inhibition of cellular transcription. With a combination of fluorography, strand-specific qRT-PCR, and tritium incorporation, we demonstrated that ML336 inhibits the synthesis of the positive sense genomic, negative sense template, and subgenomic RNAs of VEEV. Based on these results, we propose that the mechanism of action for this class of antiViral compounds is inhibition of Viral RNA synthesis through interaction with the Viral replicase complex.
Paula Turkki - One of the best experts on this subject based on the ideXlab platform.
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Human Enterovirus Group B Viruses Rely on Vimentin Dynamics for Efficient Processing of Viral Nonstructural Proteins.
Journal of Virology, 2020Co-Authors: Paula Turkki, Mira Laajala, Malin Flodström-tullberg, Varpu MarjomäkiAbstract:We report that several viruses from the human enterovirus group B cause massive vimentin rearrangements during lytic infection. Comprehensive studies suggested that Viral protein synthesis was triggering the vimentin rearrangements. Blocking the host cell vimentin dynamics with beta, beta'-iminodipropionitrile (IDPN) did not significantly affect the production of progeny viruses and only moderately lowered the synthesis of structural Proteins such as VP1. In contrast, the synthesis of the Nonstructural Proteins 2A, 3C, and 3D was drastically lowered. This led to attenuation of the cleavage of the host cell substrates PABP and G3BP1 and reduced caspase activation, leading to prolonged cell survival. Furthermore, the localization of the Proteins differed in the infected cells. Capsid protein VP1 was found diffusely around the cytoplasm, whereas 2A and 3D followed vimentin distribution. Based on protein blotting, smaller amounts of Nonstructural Proteins did not result from proteasomal degradation but from lower synthesis without intact vimentin cage structure. In contrast, inhibition of Hsp90 chaperone activity, which regulates P1 maturation, lowered the amount of VP1 but had less effect on 2A. The results suggest that the vimentin dynamics regulate Viral Nonstructural protein synthesis while having less effect on structural protein synthesis or overall infection efficiency. The results presented here shed new light on differential fate of structural and Nonstructural Proteins of enteroviruses, having consequences on host cell survival.IMPORTANCE A virus needs the host cell in order to replicate and produce new progeny viruses. For this, the virus takes over the host cell and modifies it to become a factory for Viral Proteins. Irrespective of the specific virus family, these Proteins can be divided into structural and Nonstructural Proteins. Structural Proteins are the building blocks for the new progeny virions, whereas the Nonstructural Proteins orchestrate the takeover of the host cell and its functions. Here, we have shown a mechanism that viruses exploit in order to regulate the host cell. We show that Viral protein synthesis induces vimentin cages, which promote production of specific Viral Proteins that eventually control apoptosis and host cell death. This study specifies vimentin as the key regulator of these events and indicates that Viral Proteins have different fates in the cells depending on their association with vimentin cages.
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human enterovirus group b viruses rely on vimentin dynamics for efficient processing of Viral Nonstructural Proteins
Journal of Virology, 2019Co-Authors: Paula Turkki, Mira Laajala, Malin Flodstromtullberg, Varpu MarjomäkiAbstract:: We report that several viruses from the human enterovirus group B cause massive vimentin rearrangements during lytic infection. Comprehensive studies suggested that Viral protein synthesis was triggering the vimentin rearrangements. Blocking the host cell vimentin dynamics with IDPN did not significantly affect the production of progeny viruses and only moderately lowered the synthesis of structural Proteins such as VP1. In contrast, the synthesis of the non-structural Proteins 2A, 3C, and 3D was drastically lowered. This led to attenuation of the cleavage of the host cell substrates PABP and G3BP1 and reduced caspase activation, thus leading to prolonged cell survival. Furthermore, the localization of the Proteins differed in the infected cells. Capsid protein VP1 was found diffusely around the cytoplasm, whereas 2A and 3D followed vimentin distribution. Based on protein blotting, lower amounts of non-structural Proteins did not result from proteasomal degradation, but from lower synthesis without intact vimentin cage structure. In contrast, inhibition of Hsp90 chaperone activity, which regulates P1 maturation, lowered the amount of VP1, but had less effect on 2A. The results suggest that, the vimentin dynamics regulate Viral non-structural protein synthesis while having no effect on structural protein synthesis or overall infection efficiency. The results presented here shed new light on differential fate of structural and non-structural Proteins of enteroviruses, having consequences on host cell survival.Importance A virus needs the host cell in order to replicate and produce new progeny viruses. For this, the virus takes over the host cell and modifies it to become a factory for Viral Proteins. Irrespective of the specific virus family, these Proteins can be divided into structural and non-structural Proteins. Structural Proteins are the building blocks for the new progeny virions, whereas the non-structural Proteins orchestrate the take-over of the host cell and its functions. Here we have shown a mechanism that viruses exploit in order to regulate the host cell. We show that Viral protein synthesis induces vimentin cages, which promote production of specific Viral Proteins that eventually control apoptosis and the host cell death. This study specifies vimentin as the key regulator of these events and indicates that Viral Proteins have different fates in the cells depending on their association with vimentin cages.
Andrew M Skidmore - One of the best experts on this subject based on the ideXlab platform.
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benzamidine ml336 inhibits plus and minus strand rna synthesis of venezuelan equine encephalitis virus without affecting host rna production
Antiviral Research, 2020Co-Authors: Andrew M Skidmore, Robert S Adcock, Colleen B Jonsson, Jennifer E Golden, Donghoon ChungAbstract:Abstract Venezuelan equine encephalitis virus (VEEV) is an alphavirus that is endemic to the Americas. VEEV outbreaks occur periodically and cause encephalitis in both humans and equids. There are currently no therapeutics or vaccines for treatment of VEEV in humans. Our group has previously reported on the development of a benzamidine VEEV inhibitor, ML336, which shows potent antiViral activity in both in vitro and in vivo models of infection. In cell culture experiments, ML336 inhibits Viral RNA synthesis when added 2–4 h post-infection, and mutations conferring resistance occur within the Viral Nonstructural Proteins (nsP2 and nsP4). We hypothesized that ML336 targets an activity of the Viral replicase complex and inhibits Viral RNA synthesis. To test this hypothesis, we employed various biochemical and cellular assays. Using structural analogues of ML336, we demonstrate that the cellular antiViral activity of these compounds correlates with their inhibition of Viral RNA synthesis. For instance, the IC50 of ML336 for VEEV RNA synthesis inhibition was determined as 1.1 nM, indicating potent anti-RNA synthesis activity in the low nanomolar range. While ML336 efficiently inhibited VEEV RNA synthesis, a much weaker effect was observed against the Old World alphavirus Chikungunya virus (IC50 > 4 μM), agreeing with previous data from a cell based assay. Using a tritium incorporation assay, we demonstrated that there was no significant inhibition of cellular transcription. With a combination of fluorography, strand-specific qRT-PCR, and tritium incorporation, we demonstrated that ML336 inhibits the synthesis of the positive sense genomic, negative sense template, and subgenomic RNAs of VEEV. Based on these results, we propose that the mechanism of action for this class of antiViral compounds is inhibition of Viral RNA synthesis through interaction with the Viral replicase complex.
