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Bernard La Scola - One of the best experts on this subject based on the ideXlab platform.
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Isolation and genomic characterization of a new mimivirus of lineage B from a Brazilian river
Archives of Virology, 2020Co-Authors: Ludmila Karen Dos Santos Silva, Julien Andreani, Rodrigo Araújo Lima Rodrigues, Bernard La Scola, Anthony Levasseur, Ana Claudia Santos Pereira Andrade, Hiroyuki Hikida, Jônatas Santos AbrahãoAbstract:Since its discovery, the first identified giant virus associated with amoebae, Acanthamoeba polyphaga mimivirus (APMV), has been rigorously studied to understand the structural and genomic complexity of this virus. In this work, we report the isolation and genomic characterization of a new mimivirus of lineage B, named “Borely moumouvirus”. This new virus exhibits a structure and replicative cycle similar to those of other members of the family Mimiviridae . The genome of the new isolate is a linear double-strand DNA molecule of ~1.0 Mb, containing over 900 open reading frames. Genome annotation highlighted different translation system components encoded in the DNA of Borely moumouvirus, including aminoacyl-tRNA synthetases, translation factors, and tRNA molecules, in a distribution similar to that in other lineage B mimiviruses. Pan-genome analysis indicated an increase in the genetic arsenal of this group of viruses, showing that the family Mimiviridae is still expanding. Furthermore, phylogenetic analysis has shown that Borely moumouvirus is closely related to moumouvirus australiensis. This is the first mimivirus lineage B isolated from Brazilian territory to be characterized. Further prospecting studies are necessary for us to better understand the diversity of these viruses so a better classification system can be established.
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Role of the R349 Gene and Its Repeats in the MIMIVIRE Defense System
Frontiers in Microbiology, 2019Co-Authors: Said Mougari, Jônatas Santos Abrahão, Graziele Oliveira, Jacques Bou Khalil, Bernard La ScolaAbstract:MIMIVIRE is a defense system described in lineage A Mimivirus (Mimiviridae family) that mediates resistance against Zamilon virophage. It is composed of putative helicase and nuclease associated with a gene of unknown function called R349, which contains four 15 bp repeats homologous to the virophage sequence. In a previous study, the silencing of such genes restored virophage susceptibility. Moreover, the CRISPR Cas-4 like activity of the nuclease has recently been characterized. In this study, a recently isolated Mimivirus of lineage A with R349 gene lacking 3 of 4 repeats was demonstrated to be susceptible to Zamilon. To reinforce the importance of the R349 gene in the MIMIVIRE system, we developed and presented, for the first time to our knowledge, a protocol for Mimivirus genomic editing. By knocking out R349 gene in a Mimivirus lineage A, we observed the replication of Zamilon, indicating that this gene is critical in the resistance against this specific group of virophages.
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guarani virophage a new sputnik like isolate from a brazilian lake
Frontiers in Microbiology, 2019Co-Authors: Said Mougari, Meriem Bekliz, Fabrizio Di Pinto, Jônatas Santos Abrahão, Anthony Levasseur, Bernard La ScolaAbstract:Virophages are critical regulators of viral population dynamics and potential actors in the stability of the microbial networks. These small biological entities predate the replicative cycle of giant viruses, such as the members of the Mimiviridae family or their distant relatives, which produce within the cytoplasm of their host cells a viral factory harboring a complex biochemistry propitious to the growth of the smaller parasites. In this paper, we describe the isolation and the characterization of a new virophage, the eighth, that we named Guarani. We observed that Guarani exhibits a late replication cycle compared to its giant virus host. In addition, like all Sputnik strains, Guarani is able to infect the three lineages A, B and C of the Mimiviridae family, and affects the replication and the infectivity of its host virus. In terms of genetic content, Guarani has a 18,967 bp long double-stranded DNA genome encoding 22 predicted genes very similar to Sputnik genes, except for ORF19 and ORF12. The former is more related to Zamilon while the latter seems to be novel. The architecture of the Guarani genome is closely related to Sputnik and Zamilon strains, suggesting a common origin for all these virophages.
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Table_1_Role of the R349 Gene and Its Repeats in the MIMIVIRE Defense System.XLSX
2019Co-Authors: Said Mougari, Jônatas Santos Abrahão, Graziele P. Oliveira, Jacques Bou Y. Khalil, Bernard La ScolaAbstract:MIMIVIRE is a defense system described in lineage A Mimivirus (Mimiviridae family) that mediates resistance against Zamilon virophage. It is composed of putative helicase and nuclease associated with a gene of unknown function called R349, which contains four 15 bp repeats homologous to the virophage sequence. In a previous study, the silencing of such genes restored virophage susceptibility. Moreover, the CRISPR Cas-4 like activity of the nuclease has recently been characterized. In this study, a recently isolated Mimivirus of lineage A with R349 gene lacking 3 of 4 repeats was demonstrated to be susceptible to Zamilon. To reinforce the importance of the R349 gene in the MIMIVIRE system, we developed and presented, for the first time to our knowledge, a protocol for Mimivirus genomic editing. By knocking out R349 gene in a Mimivirus lineage A, we observed the replication of Zamilon, indicating that this gene is critical in the resistance against this specific group of virophages.
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Table_1_Guarani Virophage, a New Sputnik-Like Isolate From a Brazilian Lake.XLSX
2019Co-Authors: Said Mougari, Meriem Bekliz, Fabrizio Di Pinto, Jonatas Abrahao, Anthony Levasseur, Bernard La ScolaAbstract:Virophages are critical regulators of viral population dynamics and potential actors in the stability of the microbial networks. These small biological entities predate the replicative cycle of giant viruses, such as the members of the Mimiviridae family or their distant relatives, which produce within the cytoplasm of their host cells a viral factory harboring a complex biochemistry propitious to the growth of the smaller parasites. In this paper, we describe the isolation and the characterization of a new virophage, the eighth, that we named Guarani. We observed that Guarani exhibits a late replication cycle compared to its giant virus host. In addition, like all Sputnik strains, Guarani is able to infect the three lineages A, B and C of the Mimiviridae family, and affects the replication and the infectivity of its host virus. In terms of genetic content, Guarani has a 18,967 bp long double-stranded DNA genome encoding 22 predicted genes very similar to Sputnik genes, except for ORF19 and ORF12. The former is more related to Zamilon while the latter seems to be novel. The architecture of the Guarani genome is closely related to Sputnik and Zamilon strains, suggesting a common origin for all these virophages.
Jean-michel Claverie - One of the best experts on this subject based on the ideXlab platform.
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Mimiviridae: An Expanding Family of Highly Diverse Large dsDNA Viruses Infecting a Wide Phylogenetic Range of Aquatic Eukaryotes
Viruses, 2018Co-Authors: Jean-michel Claverie, Chantal AbergelAbstract:Since 1998, when Jim van Etten’s team initiated its characterization, Paramecium bursaria Chlorella virus 1 (PBCV-1) had been the largest known DNA virus, both in terms of particle size and genome complexity. In 2003, the Acanthamoeba-infecting Mimivirus unexpectedly superseded PBCV-1, opening the era of giant viruses, i.e., with virions large enough to be visible by light microscopy and genomes encoding more proteins than many bacteria. During the following 15 years, the isolation of many Mimivirus relatives has made Mimiviridae one of the largest and most diverse families of eukaryotic viruses, most of which have been isolated from aquatic environments. Metagenomic studies of various ecosystems (including soils) suggest that many more remain to be isolated. As Mimiviridae members are found to infect an increasing range of phytoplankton species, their taxonomic position compared to the traditional Phycodnaviridae (i.e., etymologically “algal viruses”) became a source of confusion in the literature. Following a quick historical review of the key discoveries that established the Mimiviridae family, we describe its current taxonomic structure and propose a set of operational criteria to help in the classification of future isolates.
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crispr cas like system in giant viruses why mimivire is not likely to be an adaptive immune system
Virologica Sinica, 2016Co-Authors: Jean-michel Claverie, Chantal AbergelAbstract:Giant viruses from the Mimiviridae family (Mimivirus, Megavirus, etc.) replicate inside their Acanthamoeba host by the mean of a large intracytoplasmic virion factory within which DNA transcription and replication take place using the virus encoded machineries. Members of the Mimiviridae have been isolated in association with much smaller dsDNA viruses called "virophages" that replicate within the virion factory, behaving as parasites of the giant virus. In a recent work Levasseur et al. (2016) used a silencing approach to identify candidate genes possibly involved in the inhibition of the Zamilon virophage in a specific clade of Mimiviridae. Based on the presence of four copies of a short Zamilon sequence in one of the Mimivirus candidate gene, the authors proposed that resistance to virophages was conferred by a CRISPR-Cas-like system (called MIMIVIRE). Here we dispute this interpretation on the ground that 1) the simultaneous and co-localized replication of the virophage and Mimivirus genomes makes the acquisition of a nucleic acid-based immunity unlikely, 2) the Zamilonlike sequence allegedly acquired by Mimivirus are neither regularly spaced nor flanked by recognizable repeats, 3) the corresponding Zamilon sequence is devoid of distinct flanking sequences that may serve as Protospacer Adjacent Motifs (PAM) discriminating between the virophage and its host. We propose a simpler protein-based interaction model that explains the observed phenomena without having to extend the realm of adaptive immunity to the world of eukaryotic viruses, a revolutionary step that would require stronger experimental evidences.
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The Megavirus Chilensis Cu,Zn-Superoxide Dismutase: the First Viral Structure of a Typical Cellular Copper Chaperone-Independent Hyperstable Dimeric Enzyme
Journal of Virology, 2014Co-Authors: Audrey Lartigue, Chantal Abergel, Jean-michel Claverie, Bénédicte Burlat, Bruno Coutard, Florence Chaspoul, G. McfaddenAbstract:Unlabelled: Giant viruses able to replicate in Acanthamoeba castellanii penetrate their host through phagocytosis. After capsid opening, a fusion between the internal membranes of the virion and the phagocytic vacuole triggers the transfer in the cytoplasm of the viral DNA together with the DNA repair enzymes and the transcription machinery present in the particles. In addition, the proteome analysis of purified mimivirus virions revealed the presence of many enzymes meant to resist oxidative stress and conserved in the Mimiviridae. Megavirus chilensis encodes a predicted copper, zinc superoxide dismutase (Cu,Zn-SOD), an enzyme known to detoxify reactive oxygen species released in the course of host defense reactions. While it was thought that the metal ions are required for the formation of the active-site lid and dimer stabilization, megavirus chilensis SOD forms a very stable metal-free dimer. We used electron paramagnetic resonance (EPR) analysis and activity measurements to show that the supplementation of the bacterial culture with copper and zinc during the recombinant expression of Mg277 is sufficient to restore a fully active holoenzyme. These results demonstrate that the viral enzyme's activation is independent of a chaperone both for disulfide bridge formation and for copper incorporation and suggest that its assembly may not be as regulated as that of its cellular counterparts. A SOD protein is encoded by a variety of DNA viruses but is absent from mimivirus. As in poxviruses, the enzyme might be dispensable when the virus infects Acanthamoeba cells but may allow megavirus chilensis to infect a broad range of eukaryotic hosts. Importance: Mimiviridae are giant viruses encoding more than 1,000 proteins. The virion particles are loaded with proteins used by the virus to resist the vacuole's oxidative stress. The megavirus chilensis virion contains a predicted copper, zinc superoxide dismutase (Cu,Zn-SOD). The corresponding gene is present in some megavirus chilensis relatives but is absent from mimivirus. This first crystallographic structure of a viral Cu,Zn-SOD highlights the features that it has in common with and its differences from cellular SODs. It corresponds to a very stable dimer of the apo form of the enzyme. We demonstrate that upon supplementation of the growth medium with Cu and Zn, the recombinant protein is fully active, suggesting that the virus's SOD activation is independent of a copper chaperone for SOD generally used by eukaryotic SODs.
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theme
2014Co-Authors: Audrey Lartigue, Jean-michel Claverie, Chantal AbergelAbstract:Giant viruses from the Mimiviridae family replicate entirely in their host cytoplasm where their genes are transcribed by a viral transcription apparatus. mRNA polyadenylation uniquely occurs at hairpin-forming palindromic sequences terminating viral transcripts. Here we show that a conserved gene cluster both en-code the enzyme responsible for the hairpin cleavage and the viral polyA polymerases (vPAP). Unexpect-edly, the vPAPs are homodimeric and uniquely self-processive. The vPAP backbone structures exhibit a symmetrical architecture with two subdomains shar-ing a nucleotidyltransferase topology, suggesting that vPAPs originate from an ancestral duplication. A Poxvirus processivity factor homologue encoded by Megavirus chilensis displays a conserved 5′-GpppA 2′O methyltransferase activity but is also able to in-ternally methylate the mRNAs ’ polyA tails. These findings elucidate how the arm wrestling between hosts and their viruses to access the translation ma-chinery is taking place in Mimiviridae
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Mimivirus and its Virophage
Annual Review of Genetics, 2009Co-Authors: Jean-michel Claverie, Chantal AbergelAbstract:Mimivirus, a virus infecting amoebae of the acanthamoeba genus, is the prototype member of the Mimiviridae, the latest addition to the family of the nucleocytoplasmic large DNA viruses, already including the Poxviridae, the Iridoviridae, the Asfarviridae, and the Phycodnaviridae. Because of the size of its particle—a fiber-covered icosahedral protein capsid 0.75 μm in diameter—Mimivirus was initially mistaken for a parasitic bacterium. Its 1.2-Mb genome sequence encodes more than 900 proteins, many of them associated with functions never before encountered in a virus, such as four aminoacyl-tRNA synthetases. These findings revived the debate about the origin of DNA viruses and their possible role in the emergence of the eukaryotic nucleus. The recent isolation of a new type of satellite virus, called a virophage, associated with a second strain of Mimivirus, confirmed its unique position within the virus world. Post-genomic studies are now in progress, slowly shedding some light on the physiology of the m...
Chantal Abergel - One of the best experts on this subject based on the ideXlab platform.
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Mimiviridae: An Expanding Family of Highly Diverse Large dsDNA Viruses Infecting a Wide Phylogenetic Range of Aquatic Eukaryotes
Viruses, 2018Co-Authors: Jean-michel Claverie, Chantal AbergelAbstract:Since 1998, when Jim van Etten’s team initiated its characterization, Paramecium bursaria Chlorella virus 1 (PBCV-1) had been the largest known DNA virus, both in terms of particle size and genome complexity. In 2003, the Acanthamoeba-infecting Mimivirus unexpectedly superseded PBCV-1, opening the era of giant viruses, i.e., with virions large enough to be visible by light microscopy and genomes encoding more proteins than many bacteria. During the following 15 years, the isolation of many Mimivirus relatives has made Mimiviridae one of the largest and most diverse families of eukaryotic viruses, most of which have been isolated from aquatic environments. Metagenomic studies of various ecosystems (including soils) suggest that many more remain to be isolated. As Mimiviridae members are found to infect an increasing range of phytoplankton species, their taxonomic position compared to the traditional Phycodnaviridae (i.e., etymologically “algal viruses”) became a source of confusion in the literature. Following a quick historical review of the key discoveries that established the Mimiviridae family, we describe its current taxonomic structure and propose a set of operational criteria to help in the classification of future isolates.
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crispr cas like system in giant viruses why mimivire is not likely to be an adaptive immune system
Virologica Sinica, 2016Co-Authors: Jean-michel Claverie, Chantal AbergelAbstract:Giant viruses from the Mimiviridae family (Mimivirus, Megavirus, etc.) replicate inside their Acanthamoeba host by the mean of a large intracytoplasmic virion factory within which DNA transcription and replication take place using the virus encoded machineries. Members of the Mimiviridae have been isolated in association with much smaller dsDNA viruses called "virophages" that replicate within the virion factory, behaving as parasites of the giant virus. In a recent work Levasseur et al. (2016) used a silencing approach to identify candidate genes possibly involved in the inhibition of the Zamilon virophage in a specific clade of Mimiviridae. Based on the presence of four copies of a short Zamilon sequence in one of the Mimivirus candidate gene, the authors proposed that resistance to virophages was conferred by a CRISPR-Cas-like system (called MIMIVIRE). Here we dispute this interpretation on the ground that 1) the simultaneous and co-localized replication of the virophage and Mimivirus genomes makes the acquisition of a nucleic acid-based immunity unlikely, 2) the Zamilonlike sequence allegedly acquired by Mimivirus are neither regularly spaced nor flanked by recognizable repeats, 3) the corresponding Zamilon sequence is devoid of distinct flanking sequences that may serve as Protospacer Adjacent Motifs (PAM) discriminating between the virophage and its host. We propose a simpler protein-based interaction model that explains the observed phenomena without having to extend the realm of adaptive immunity to the world of eukaryotic viruses, a revolutionary step that would require stronger experimental evidences.
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The Megavirus Chilensis Cu,Zn-Superoxide Dismutase: the First Viral Structure of a Typical Cellular Copper Chaperone-Independent Hyperstable Dimeric Enzyme
Journal of Virology, 2014Co-Authors: Audrey Lartigue, Chantal Abergel, Jean-michel Claverie, Bénédicte Burlat, Bruno Coutard, Florence Chaspoul, G. McfaddenAbstract:Unlabelled: Giant viruses able to replicate in Acanthamoeba castellanii penetrate their host through phagocytosis. After capsid opening, a fusion between the internal membranes of the virion and the phagocytic vacuole triggers the transfer in the cytoplasm of the viral DNA together with the DNA repair enzymes and the transcription machinery present in the particles. In addition, the proteome analysis of purified mimivirus virions revealed the presence of many enzymes meant to resist oxidative stress and conserved in the Mimiviridae. Megavirus chilensis encodes a predicted copper, zinc superoxide dismutase (Cu,Zn-SOD), an enzyme known to detoxify reactive oxygen species released in the course of host defense reactions. While it was thought that the metal ions are required for the formation of the active-site lid and dimer stabilization, megavirus chilensis SOD forms a very stable metal-free dimer. We used electron paramagnetic resonance (EPR) analysis and activity measurements to show that the supplementation of the bacterial culture with copper and zinc during the recombinant expression of Mg277 is sufficient to restore a fully active holoenzyme. These results demonstrate that the viral enzyme's activation is independent of a chaperone both for disulfide bridge formation and for copper incorporation and suggest that its assembly may not be as regulated as that of its cellular counterparts. A SOD protein is encoded by a variety of DNA viruses but is absent from mimivirus. As in poxviruses, the enzyme might be dispensable when the virus infects Acanthamoeba cells but may allow megavirus chilensis to infect a broad range of eukaryotic hosts. Importance: Mimiviridae are giant viruses encoding more than 1,000 proteins. The virion particles are loaded with proteins used by the virus to resist the vacuole's oxidative stress. The megavirus chilensis virion contains a predicted copper, zinc superoxide dismutase (Cu,Zn-SOD). The corresponding gene is present in some megavirus chilensis relatives but is absent from mimivirus. This first crystallographic structure of a viral Cu,Zn-SOD highlights the features that it has in common with and its differences from cellular SODs. It corresponds to a very stable dimer of the apo form of the enzyme. We demonstrate that upon supplementation of the growth medium with Cu and Zn, the recombinant protein is fully active, suggesting that the virus's SOD activation is independent of a copper chaperone for SOD generally used by eukaryotic SODs.
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theme
2014Co-Authors: Audrey Lartigue, Jean-michel Claverie, Chantal AbergelAbstract:Giant viruses from the Mimiviridae family replicate entirely in their host cytoplasm where their genes are transcribed by a viral transcription apparatus. mRNA polyadenylation uniquely occurs at hairpin-forming palindromic sequences terminating viral transcripts. Here we show that a conserved gene cluster both en-code the enzyme responsible for the hairpin cleavage and the viral polyA polymerases (vPAP). Unexpect-edly, the vPAPs are homodimeric and uniquely self-processive. The vPAP backbone structures exhibit a symmetrical architecture with two subdomains shar-ing a nucleotidyltransferase topology, suggesting that vPAPs originate from an ancestral duplication. A Poxvirus processivity factor homologue encoded by Megavirus chilensis displays a conserved 5′-GpppA 2′O methyltransferase activity but is also able to in-ternally methylate the mRNAs ’ polyA tails. These findings elucidate how the arm wrestling between hosts and their viruses to access the translation ma-chinery is taking place in Mimiviridae
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Preliminary crystallographic analysis of a possible transcription factor encoded by the mimivirus L544 gene.
Acta Crystallographica Section F-structural Biology and Crystallization Communications, 2011Co-Authors: Alexandre Ciaccafava, Sandra Jeudy, Pascal Mansuelle, Chantal AbergelAbstract:Mimivirus is the prototype of a new family (the Mimiviridae) of nucleocytoplasmic large DNA viruses (NCLDVs), which already include the Poxviridae, Iridoviridae, Phycodnaviridae and Asfarviridae. Mimivirus specifically replicates in cells from the genus Acanthamoeba. Proteomic analysis of purified mimivirus particles revealed the presence of many subunits of the DNA-directed RNA polymerase II complex. A fully functional pre-transcriptional complex appears to be loaded in the virions, allowing mimivirus to initiate transcription within the host cytoplasm immediately upon infection independently of the host nuclear apparatus. To fully understand this process, a systematic study of mimivirus proteins that are predicted (by bioinformatics) or suspected (by proteomic analysis) to be involved in transcription was initiated by cloning and expressing them in Escherichia coli in order to determine their three-dimensional structures. Here, preliminary crystallographic analysis of the recombinant L544 protein is reported. The crystals belonged to the orthorhombic space group C2221 with one monomer per asymmetric unit. A MAD data set was used for preliminary phasing using the selenium signal present in a selenomethionine-substituted protein crystal.
Jônatas Santos Abrahão - One of the best experts on this subject based on the ideXlab platform.
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Isolation and genomic characterization of a new mimivirus of lineage B from a Brazilian river
Archives of Virology, 2020Co-Authors: Ludmila Karen Dos Santos Silva, Julien Andreani, Rodrigo Araújo Lima Rodrigues, Bernard La Scola, Anthony Levasseur, Ana Claudia Santos Pereira Andrade, Hiroyuki Hikida, Jônatas Santos AbrahãoAbstract:Since its discovery, the first identified giant virus associated with amoebae, Acanthamoeba polyphaga mimivirus (APMV), has been rigorously studied to understand the structural and genomic complexity of this virus. In this work, we report the isolation and genomic characterization of a new mimivirus of lineage B, named “Borely moumouvirus”. This new virus exhibits a structure and replicative cycle similar to those of other members of the family Mimiviridae . The genome of the new isolate is a linear double-strand DNA molecule of ~1.0 Mb, containing over 900 open reading frames. Genome annotation highlighted different translation system components encoded in the DNA of Borely moumouvirus, including aminoacyl-tRNA synthetases, translation factors, and tRNA molecules, in a distribution similar to that in other lineage B mimiviruses. Pan-genome analysis indicated an increase in the genetic arsenal of this group of viruses, showing that the family Mimiviridae is still expanding. Furthermore, phylogenetic analysis has shown that Borely moumouvirus is closely related to moumouvirus australiensis. This is the first mimivirus lineage B isolated from Brazilian territory to be characterized. Further prospecting studies are necessary for us to better understand the diversity of these viruses so a better classification system can be established.
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Role of the R349 Gene and Its Repeats in the MIMIVIRE Defense System
Frontiers in Microbiology, 2019Co-Authors: Said Mougari, Jônatas Santos Abrahão, Graziele Oliveira, Jacques Bou Khalil, Bernard La ScolaAbstract:MIMIVIRE is a defense system described in lineage A Mimivirus (Mimiviridae family) that mediates resistance against Zamilon virophage. It is composed of putative helicase and nuclease associated with a gene of unknown function called R349, which contains four 15 bp repeats homologous to the virophage sequence. In a previous study, the silencing of such genes restored virophage susceptibility. Moreover, the CRISPR Cas-4 like activity of the nuclease has recently been characterized. In this study, a recently isolated Mimivirus of lineage A with R349 gene lacking 3 of 4 repeats was demonstrated to be susceptible to Zamilon. To reinforce the importance of the R349 gene in the MIMIVIRE system, we developed and presented, for the first time to our knowledge, a protocol for Mimivirus genomic editing. By knocking out R349 gene in a Mimivirus lineage A, we observed the replication of Zamilon, indicating that this gene is critical in the resistance against this specific group of virophages.
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guarani virophage a new sputnik like isolate from a brazilian lake
Frontiers in Microbiology, 2019Co-Authors: Said Mougari, Meriem Bekliz, Fabrizio Di Pinto, Jônatas Santos Abrahão, Anthony Levasseur, Bernard La ScolaAbstract:Virophages are critical regulators of viral population dynamics and potential actors in the stability of the microbial networks. These small biological entities predate the replicative cycle of giant viruses, such as the members of the Mimiviridae family or their distant relatives, which produce within the cytoplasm of their host cells a viral factory harboring a complex biochemistry propitious to the growth of the smaller parasites. In this paper, we describe the isolation and the characterization of a new virophage, the eighth, that we named Guarani. We observed that Guarani exhibits a late replication cycle compared to its giant virus host. In addition, like all Sputnik strains, Guarani is able to infect the three lineages A, B and C of the Mimiviridae family, and affects the replication and the infectivity of its host virus. In terms of genetic content, Guarani has a 18,967 bp long double-stranded DNA genome encoding 22 predicted genes very similar to Sputnik genes, except for ORF19 and ORF12. The former is more related to Zamilon while the latter seems to be novel. The architecture of the Guarani genome is closely related to Sputnik and Zamilon strains, suggesting a common origin for all these virophages.
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Image_1_Role of the R349 Gene and Its Repeats in the MIMIVIRE Defense System.TIF
2019Co-Authors: Said Mougari, Jônatas Santos Abrahão, Graziele P. Oliveira, Jacques Bou Y. Khalil, Bernard La ScolaAbstract:MIMIVIRE is a defense system described in lineage A Mimivirus (Mimiviridae family) that mediates resistance against Zamilon virophage. It is composed of putative helicase and nuclease associated with a gene of unknown function called R349, which contains four 15 bp repeats homologous to the virophage sequence. In a previous study, the silencing of such genes restored virophage susceptibility. Moreover, the CRISPR Cas-4 like activity of the nuclease has recently been characterized. In this study, a recently isolated Mimivirus of lineage A with R349 gene lacking 3 of 4 repeats was demonstrated to be susceptible to Zamilon. To reinforce the importance of the R349 gene in the MIMIVIRE system, we developed and presented, for the first time to our knowledge, a protocol for Mimivirus genomic editing. By knocking out R349 gene in a Mimivirus lineage A, we observed the replication of Zamilon, indicating that this gene is critical in the resistance against this specific group of virophages.
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Table_1_Role of the R349 Gene and Its Repeats in the MIMIVIRE Defense System.XLSX
2019Co-Authors: Said Mougari, Jônatas Santos Abrahão, Graziele P. Oliveira, Jacques Bou Y. Khalil, Bernard La ScolaAbstract:MIMIVIRE is a defense system described in lineage A Mimivirus (Mimiviridae family) that mediates resistance against Zamilon virophage. It is composed of putative helicase and nuclease associated with a gene of unknown function called R349, which contains four 15 bp repeats homologous to the virophage sequence. In a previous study, the silencing of such genes restored virophage susceptibility. Moreover, the CRISPR Cas-4 like activity of the nuclease has recently been characterized. In this study, a recently isolated Mimivirus of lineage A with R349 gene lacking 3 of 4 repeats was demonstrated to be susceptible to Zamilon. To reinforce the importance of the R349 gene in the MIMIVIRE system, we developed and presented, for the first time to our knowledge, a protocol for Mimivirus genomic editing. By knocking out R349 gene in a Mimivirus lineage A, we observed the replication of Zamilon, indicating that this gene is critical in the resistance against this specific group of virophages.
G. Mcfadden - One of the best experts on this subject based on the ideXlab platform.
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The Megavirus Chilensis Cu,Zn-Superoxide Dismutase: the First Viral Structure of a Typical Cellular Copper Chaperone-Independent Hyperstable Dimeric Enzyme
Journal of Virology, 2014Co-Authors: Audrey Lartigue, Chantal Abergel, Jean-michel Claverie, Bénédicte Burlat, Bruno Coutard, Florence Chaspoul, G. McfaddenAbstract:Unlabelled: Giant viruses able to replicate in Acanthamoeba castellanii penetrate their host through phagocytosis. After capsid opening, a fusion between the internal membranes of the virion and the phagocytic vacuole triggers the transfer in the cytoplasm of the viral DNA together with the DNA repair enzymes and the transcription machinery present in the particles. In addition, the proteome analysis of purified mimivirus virions revealed the presence of many enzymes meant to resist oxidative stress and conserved in the Mimiviridae. Megavirus chilensis encodes a predicted copper, zinc superoxide dismutase (Cu,Zn-SOD), an enzyme known to detoxify reactive oxygen species released in the course of host defense reactions. While it was thought that the metal ions are required for the formation of the active-site lid and dimer stabilization, megavirus chilensis SOD forms a very stable metal-free dimer. We used electron paramagnetic resonance (EPR) analysis and activity measurements to show that the supplementation of the bacterial culture with copper and zinc during the recombinant expression of Mg277 is sufficient to restore a fully active holoenzyme. These results demonstrate that the viral enzyme's activation is independent of a chaperone both for disulfide bridge formation and for copper incorporation and suggest that its assembly may not be as regulated as that of its cellular counterparts. A SOD protein is encoded by a variety of DNA viruses but is absent from mimivirus. As in poxviruses, the enzyme might be dispensable when the virus infects Acanthamoeba cells but may allow megavirus chilensis to infect a broad range of eukaryotic hosts. Importance: Mimiviridae are giant viruses encoding more than 1,000 proteins. The virion particles are loaded with proteins used by the virus to resist the vacuole's oxidative stress. The megavirus chilensis virion contains a predicted copper, zinc superoxide dismutase (Cu,Zn-SOD). The corresponding gene is present in some megavirus chilensis relatives but is absent from mimivirus. This first crystallographic structure of a viral Cu,Zn-SOD highlights the features that it has in common with and its differences from cellular SODs. It corresponds to a very stable dimer of the apo form of the enzyme. We demonstrate that upon supplementation of the growth medium with Cu and Zn, the recombinant protein is fully active, suggesting that the virus's SOD activation is independent of a copper chaperone for SOD generally used by eukaryotic SODs.