scholarly journals Recent Advances in Bunyavirus Reverse Genetics Research: Systems Development, Applications, and Future Perspectives

2021 ◽  
Vol 12 ◽  
Author(s):  
Fuli Ren ◽  
Shu Shen ◽  
Qiongya Wang ◽  
Gang Wei ◽  
Chaolin Huang ◽  
...  
Keyword(s):  
Viral Replication ◽  
Reverse Genetics ◽  
Rna Viruses ◽  
Current Knowledge ◽  
Evolutionary Process ◽  
Replication Cycle ◽  
Necessary Condition ◽  
Emerging Viruses ◽  
Host Interactions ◽  
Genetics Research

Bunyaviruses are members of the Bunyavirales order, which is the largest group of RNA viruses, comprising 12 families, including a large group of emerging and re-emerging viruses. These viruses can infect a wide variety of species worldwide, such as arthropods, protozoans, plants, animals, and humans, and pose substantial threats to the public. In view of the fact that a better understanding of the life cycle of a highly pathogenic virus is often a precondition for developing vaccines and antivirals, it is urgent to develop powerful tools to unravel the molecular basis of the pathogenesis. However, biosafety level −3 or even −4 containment laboratory is considered as a necessary condition for working with a number of bunyaviruses, which has hampered various studies. Reverse genetics systems, including minigenome (MG), infectious virus-like particle (iVLP), and infectious full-length clone (IFLC) systems, are capable of recapitulating some or all steps of the viral replication cycle; among these, the MG and iVLP systems have been very convenient and effective tools, allowing researchers to manipulate the genome segments of pathogenic viruses at lower biocontainment to investigate the viral genome transcription, replication, virus entry, and budding. The IFLC system is generally developed based on the MG or iVLP systems, which have facilitated the generation of recombinant infectious viruses. The MG, iVLP, and IFLC systems have been successfully developed for some important bunyaviruses and have been widely employed as powerful tools to investigate the viral replication cycle, virus–host interactions, virus pathogenesis, and virus evolutionary process. The majority of bunyaviruses is generally enveloped negative-strand RNA viruses with two to six genome segments, of which the viruses with bipartite and tripartite genome segments have mostly been characterized. This review aimed to summarize current knowledge on reverse genetic studies of representative bunyaviruses causing severe diseases in humans and animals, which will contribute to the better understanding of the bunyavirus replication cycle and provide some hints for developing designed antivirals.

scholarly journals RNA Virus-Encoded miRNAs: Current Insights and Future Challenges

2021 ◽  
Vol 12 ◽  
Author(s):  
Asuka Nanbo ◽  
Wakako Furuyama ◽  
Zhen Lin
Keyword(s):  
Viral Replication ◽  
Rna Viruses ◽  
Current Knowledge ◽  
Rna Virus ◽  
Transcriptional Level ◽  
Emerging Viruses ◽  
Wide Range ◽  
Future Challenges ◽  
Eukaryotic Gene ◽  
Potential Applications

MicroRNAs are small non-coding RNAs that regulate eukaryotic gene expression at the post-transcriptional level and affect a wide range of biological processes. Over the past two decades, numerous virus-encoded miRNAs have been identified. Some of them are crucial for viral replication, whereas others can help immune evasion. Recent sequencing-based bioinformatics methods have helped identify many novel miRNAs, which are encoded by RNA viruses. Unlike the well-characterized DNA virus-encoded miRNAs, the role of RNA virus-encoded miRNAs remains controversial. In this review, we first describe the current knowledge of miRNAs encoded by various RNA viruses, including newly emerging viruses. Next, we discuss how RNA virus-encoded miRNAs might facilitate viral replication, immunoevasion, and persistence in their hosts. Last, we briefly discuss the challenges in the experimental methodologies and potential applications of miRNAs for diagnosis and therapeutics.

scholarly journals Reverse Genetics System Demonstrates that Rotavirus Nonstructural Protein NSP6 Is Not Essential for Viral Replication in Cell Culture

2017 ◽  
Vol 91 (21) ◽  
Author(s):  
Satoshi Komoto ◽  
Yuta Kanai ◽  
Saori Fukuda ◽  
Masanori Kugita ◽  
Takahiro Kawagishi ◽  
...  
Keyword(s):  
Cell Culture ◽  
Viral Replication ◽  
Reverse Genetics ◽  
Nonstructural Protein ◽  
Growth Curves ◽  
Single Step ◽  
Replication Cycle ◽  
Nonstructural Proteins ◽  
Virus Growth ◽  
Severe Diarrhea

ABSTRACT The use of overlapping open reading frames (ORFs) to synthesize more than one unique protein from a single mRNA has been described for several viruses. Segment 11 of the rotavirus genome encodes two nonstructural proteins, NSP5 and NSP6. The NSP6 ORF is present in the vast majority of rotavirus strains, and therefore the NSP6 protein would be expected to have a function in viral replication. However, there is no direct evidence of its function or requirement in the viral replication cycle yet. Here, taking advantage of a recently established plasmid-only-based reverse genetics system that allows rescue of recombinant rotaviruses entirely from cloned cDNAs, we generated NSP6-deficient viruses to directly address its significance in the viral replication cycle. Viable recombinant NSP6-deficient viruses could be engineered. Single-step growth curves and plaque formation of the NSP6-deficient viruses confirmed that NSP6 expression is of limited significance for RVA replication in cell culture, although the NSP6 protein seemed to promote efficient virus growth. IMPORTANCE Rotavirus is one of the most important pathogens of severe diarrhea in young children worldwide. The rotavirus genome, consisting of 11 segments of double-stranded RNA, encodes six structural proteins (VP1 to VP4, VP6, and VP7) and six nonstructural proteins (NSP1 to NSP6). Although specific functions have been ascribed to each of the 12 viral proteins, the role of NSP6 in the viral replication cycle remains unknown. In this study, we demonstrated that the NSP6 protein is not essential for viral replication in cell culture by using a recently developed plasmid-only-based reverse genetics system. This reverse genetics approach will be successfully applied to answer questions of great interest regarding the roles of rotaviral proteins in replication and pathogenicity, which can hardly be addressed by conventional approaches.

scholarly journals Exploitation of Host Factors and Cellular Pathways by Tombusviruses for the Biogenesis of the Viral Replication Organelles

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 18
Author(s):  
Peter D. Nagy
Keyword(s):  
Viral Replication ◽  
Rna Viruses ◽  
Plant Viruses ◽  
Complex Nature ◽  
Host Proteins ◽  
Yeast Saccharomyces Cerevisiae ◽  
Host Interactions ◽  
Genome Wide ◽  
Show Evidence ◽  
Surrogate Host

Plus-stranded RNA viruses recruit cellular vesicles and co-opt cellular proteins involved in cellular metabolism and lipid biosynthesis to build viral replicase complexes (VRCs) within the large viral replication compartments. We use tombusviruses (TBSV), which are small (+)RNA viruses, as model plant viruses to study virus replication, recombination, and virus–host interactions using yeast (Saccharomyces cerevisiae) as a surrogate host. Several systematic genome-wide screens and global proteomic and lipidomic approaches have led to the identification of ~500 host proteins/genes that are implicated in TBSV replication. We characterized the role of two-dozen co-opted host proteins, sterols, and phosphatidylethanolamine in tombusvirus VRC assembly and viral RNA synthesis. We provide evidence on the critical roles of phosphoinositides and co-opted membrane-shaping proteins in VRC formation. We also present data that tombusviruses hijack the glycolytic and fermentation pathways to obtain ATP, which is required for the biogenesis of the replication compartment. Finally, we show evidence that TBSV usurps COPII and endosomal vesicles to form a unique microenvironment involving peroxisomes and endoplasmic reticulum (ER) to support viral replication. These new insights highlight the amazingly complex nature of virus-host interactions.

scholarly journals Rapid reconstruction of SARS-CoV-2 using a synthetic genomics platform

2020 ◽  
Author(s):  
Tran Thi Nhu Thao ◽  
Fabien Labroussaa ◽  
Nadine Ebert ◽  
Philip V’kovski ◽  
Hanspeter Stalder ◽  
...  
Keyword(s):  
Reverse Genetics ◽  
Yeast Artificial Chromosome ◽  
Vaccine Development ◽  
Rna Viruses ◽  
Rna Virus ◽  
Functional Characterization ◽  
Clinical Samples ◽  
Emerging Viruses ◽  
Synthetic Dna ◽  
Synthetic Genomics

AbstractReverse genetics has been an indispensable tool revolutionising our insights into viral pathogenesis and vaccine development. Large RNA virus genomes, such as from Coronaviruses, are cumbersome to clone and to manipulate in E. coli hosts due to size and occasional instability1-3. Therefore, an alternative rapid and robust reverse genetics platform for RNA viruses would benefit the research community. Here we show the full functionality of a yeast-based synthetic genomics platform for the genetic reconstruction of diverse RNA viruses, including members of the Coronaviridae, Flaviviridae and Paramyxoviridae families. Viral subgenomic fragments were generated using viral isolates, cloned viral DNA, clinical samples, or synthetic DNA, and reassembled in one step in Saccharomyces cerevisiae using transformation associated recombination (TAR) cloning to maintain the genome as a yeast artificial chromosome (YAC). T7-RNA polymerase has been used to generate infectious RNA, which was then used to rescue viable virus. Based on this platform we have been able to engineer and resurrect chemically-synthetized clones of the recent epidemic SARS-CoV-24 in only a week after receipt of the synthetic DNA fragments. The technical advance we describe here allows to rapidly responding to emerging viruses as it enables the generation and functional characterization of evolving RNA virus variants - in real-time - during an outbreak.

scholarly journals Infection of Mammals and Mosquitoes by Alphaviruses: Involvement of Cell Death

Cells ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 2612
Author(s):  
Lucie Cappuccio ◽  
Carine Maisse
Keyword(s):  
Cell Death ◽  
Current Knowledge ◽  
Blood Feeding ◽  
Mosquito Vector ◽  
Global Public Health ◽  
Emerging Viruses ◽  
Pathological Effect ◽  
Host Interactions ◽  
Regulated Cell Death

Alphaviruses, such as the chikungunya virus, are emerging and re-emerging viruses that pose a global public health threat. They are transmitted by blood-feeding arthropods, mainly mosquitoes, to humans and animals. Although alphaviruses cause debilitating diseases in mammalian hosts, it appears that they have no pathological effect on the mosquito vector. Alphavirus/host interactions are increasingly studied at cellular and molecular levels. While it seems clear that apoptosis plays a key role in some human pathologies, the role of cell death in determining the outcome of infections in mosquitoes remains to be fully understood. Here, we review the current knowledge on alphavirus-induced regulated cell death in hosts and vectors and the possible role they play in determining tolerance or resistance of mosquitoes.

scholarly journals BTV-vector-host interaction: the epidemiological triangle in disease transmission

2018 ◽  
Vol 74 (1) ◽  
pp. 5996-2018
Author(s):  
WIESŁAW NIEDBALSKI
Keyword(s):  
Experimental Infection ◽  
Disease Transmission ◽  
Reverse Genetics ◽  
Replication Cycle ◽  
Host Interactions ◽  
Host Interaction ◽  
Infection Models ◽  
Single Animal ◽  
Virus Replication Cycle ◽  
New Generation

Understanding the interaction between the bluetongue virus (BTV), the Culicoides vector and the ruminant host is essential to control bluetongue (BT). This triangle of interaction can be understood individually at the level of the virus, the level of vector and the host level. BTV-vector-host interactions involve physiological and ecological mechanisms, and they have evolved under a specific set of environmental conditions. Recent advances in understanding this interaction include increased knowledge of the virus replication cycle, BTV immunology and pathogenesis in the vertebrate host, as well as the virulence and pathogenicity features of newly discovered BTV serotypes. To understand the virus-host-vector interaction, new molecular biology techniques and experimental infection biology methods have been widely used. The next-generation sequencing, the establishment of a reverse genetics system for the virus, and development of novel infection models and refinement of the existing BTV experimental infection methodologies have proven very helpful. This progress in biotechnology has also made it possible to develop new-generation BTV vaccines, such as disabled infectious single cycle (DISC) vaccines and disabled infectious single animal (DISA) vaccines. However, several questions still need to be answered, such as those concerning cellular pathways involved in the induction of innate immunity and the function of NS4 in the BTV replication cycle. In addition, the identities of specific molecular determinants and the role of quasi-species diversity in determining BTV phenotype are still unclear and should be better explained..

scholarly journals Povidone-Iodine Attenuates Viral Replication in Ocular Cells: Implications for Ocular Transmission of RNA Viruses

Biomolecules ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 753
Author(s):  
Sneha Singh ◽  
Onkar B. Sawant ◽  
Shahzad I. Mian ◽  
Ashok Kumar
Keyword(s):  
Epithelial Cells ◽  
Viral Replication ◽  
Povidone Iodine ◽  
Rna Viruses ◽  
Retinal Pigment ◽  
Retinal Pigment Epithelial Cells ◽  
Host Cells ◽  
Eye Banking ◽  
Antiviral Effects ◽  
Infected Cells

Several RNA viruses, including SARS-CoV-2, can infect or use the eye as an entry portal to cause ocular or systemic diseases. Povidone-Iodine (PVP-I) is routinely used during ocular surgeries and eye banking as a cost-effective disinfectant due to its broad-spectrum antimicrobial activity, including against viruses. However, whether PVP-I can exert antiviral activities in virus-infected cells remains elusive. In this study, using Zika (ZIKV) and Chikungunya (CHIKV) virus infection of human corneal and retinal pigment epithelial cells, we report antiviral mechanisms of PVP-I. Our data showed that PVP-I, even at the lowest concentration (0.01%), drastically reduced viral replication in corneal and retinal cells without causing cellular toxicity. Antiviral effects of PVP-I against ZIKV and CHIKV were mediated by direct viral inactivation, thus attenuating the ability of the virus to infect host cells. Moreover, one-minute PVP-I exposure of infected ocular cells drastically reduced viral replication and the production of infectious progeny virions. Furthermore, viral-induced (CHIKV) expression of inflammatory genes (TNF-α, IL-6, IL-8, and IL1β) were markedly reduced in PVP-I treated corneal epithelial cells. Together, our results demonstrate potent antiviral effects of PVP-I against ZIKV and CHIKV infection of ocular cells. Thus, a low dose of PVP-I can be used during tissue harvesting for corneal transplants to prevent potential transmission of RNA viruses via infected cells.

scholarly journals Reverse Genetics for Peste des Petits Ruminants Virus: Current Status and Lessons to Learn from Other Non-segmented Negative-Sense RNA Viruses

2018 ◽  
Vol 33 (6) ◽  
pp. 472-483 ◽  
Author(s):  
Alfred Niyokwishimira ◽  
Yongxi Dou ◽  
Bang Qian ◽  
Prajapati Meera ◽  
Zhidong Zhang
Keyword(s):  
Reverse Genetics ◽  
Rna Viruses ◽  
Current Status ◽  
Peste Des Petits Ruminants ◽  
Negative Sense

scholarly journals Inhibitory and complementary therapeutic effect of sweet lime (Citrus limetta) against RNA-viruses

2021 ◽  
Vol 7 (1) ◽  
pp. 37-44
Author(s):  
Sulagna Ray Pal ◽  
Swapan Banerjee
Keyword(s):  
Therapeutic Effect ◽  
Rna Viruses ◽  
Current Knowledge ◽  
Western World ◽  
Therapeutic Effects ◽  
Neuroprotective Effects ◽  
Nutrient Contents ◽  
Therapeutic Benefits ◽  
Sweet Lime ◽  
Antiviral Activities

Sweet lime (), known as 'Mousambi' or 'Mosambi' in India, is one of the best citrus fruits regarding its nutrient contents. Its bioactive compounds (BAC) are exclusively used for multiple clinical applications considering many therapeutic benefits not only in Asian countries but also in the western world. The fruit pulp and juice are the best sources of ascorbic acid, B-vitamins, amino acids, and other secondary metabolites. Specifically, polyphenols such as flavanones, hesperetin, naringenin, and chlorogenic acid are highly rich in the fruit. The nutrients in sweet lime altogether provide significant anti-inflammatory, antioxidant, anti-cancer, and neuroprotective effects. The purpose of this study is to review and analyze the inhibitory and complementary therapeutic effects of sweet lime's pulp and juices to inhibit the virulence caused by RNA viruses, mainly SARS-CoV-2. This review study was designed based on extensive online searches of relevant open-access literature available in the best quality and reliable databases by using specific keywords and boolean operators. After a rigorous review, we found that flavanones in the fruit can alter or inhibit the polyproteins (pp1a and pp1b) responsible for viral replication. Therefore, sweet lime has potentialities to provide an inhibitory and a complementary therapeutic effect against RNA viruses, mainly SARS-CoV-2. About the antiviral activities, more clinical trials are needed to prove its efficacy; however, reviewing current knowledge, is one of the potent antioxidant, inflammatory fruits available and affordable almost worldwide.

scholarly journals Regulation of Viral Restriction by Post-Translational Modifications

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2197
Author(s):  
Célia Chamontin ◽  
Guillaume Bossis ◽  
Sébastien Nisole ◽  
Nathalie J. Arhel ◽  
Ghizlane Maarifi
Keyword(s):  
Viral Replication ◽  
Defense Mechanisms ◽  
Current Knowledge ◽  
Restriction Factors ◽  
Intrinsic Immunity ◽  
Post Translational Modifications ◽  
Major Mechanism ◽  
Wide Range ◽  
Subsequent Degradation ◽  
Global Vision

Intrinsic immunity is orchestrated by a wide range of host cellular proteins called restriction factors. They have the capacity to interfere with viral replication, and most of them are tightly regulated by interferons (IFNs). In addition, their regulation through post-translational modifications (PTMs) constitutes a major mechanism to shape their action positively or negatively. Following viral infection, restriction factor modification can be decisive. Palmitoylation of IFITM3, SUMOylation of MxA, SAMHD1 and TRIM5α or glycosylation of BST2 are some of those PTMs required for their antiviral activity. Nonetheless, for their benefit and by manipulating the PTMs machinery, viruses have evolved sophisticated mechanisms to counteract restriction factors. Indeed, many viral proteins evade restriction activity by inducing their ubiquitination and subsequent degradation. Studies on PTMs and their substrates are essential for the understanding of the antiviral defense mechanisms and provide a global vision of all possible regulations of the immune response at a given time and under specific infection conditions. Our aim was to provide an overview of current knowledge regarding the role of PTMs on restriction factors with an emphasis on their impact on viral replication.

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