Single-stranded positive RNA ((+) ssRNA) viruses include several important human pathogens. Some members are responsible for large outbreaks, such as Zika virus, West Nile virus, SARS-CoV, and SARS-CoV-2, while others are endemic, causing an enormous global health burden. Since vaccines or specific treatments are not available for most viral infections, the discovery of direct-acting antivirals (DAA) is an urgent need. Still, the low-throughput nature of and biosafety concerns related to traditional antiviral assays hinders the discovery of new inhibitors. With the advances of reverse genetics, reporter replicon systems have become an alternative tool for the screening of DAAs. Herein, we review decades of the use of (+) ssRNA viruses replicon systems for the discovery of antiviral agents. We summarize different strategies used to develop those systems, as well as highlight some of the most promising inhibitors identified by the method. Despite the genetic alterations introduced, reporter replicons have been shown to be reliable systems for screening and identification of viral replication inhibitors and, therefore, an important tool for the discovery of new DAAs.
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http://dx.doi.org/10.3390/v12060598 | DOI Listing |
Virology
December 2024
The Centre for Infection and Immunity Studies, School of Medicine, Sun Yat-sen University, Shenzhen, Guangdong, 518107, China. Electronic address:
The knowledge on the life cycle of flaviviruses is still incomplete, and no direct-acting antivirals against their infections are clinically available. Herein, by screening via a Zika virus (ZIKV) replicon assay, we found that the N-terminus of NS2A exhibited great tolerance to the insertions of different split fluorescent proteins (split-FPs). Furthermore, both ZIKV and dengue virus encoding a split-FP-tagged NS2A propagated efficiently, and the split-FP-tagged ZIKVs had good genetic stability.
View Article and Find Full Text PDFMethods Mol Biol
December 2024
Viral Special Pathogens Branch, Division of High-Consequence Pathogens & Pathology, Centers for Disease Control & Prevention, Atlanta, GA, USA.
Reverse genetic systems are powerful tools in molecular virology that allow the generation of infectious recombinant virus and the manipulation of viral genomes. Reverse genetic systems enable the incorporation of reporter genes, facilitating many virological assays, including high-throughput screening. Additionally, reverse genetic systems can be used to introduce targeted mutations into the viral genome, allowing investigations of viral genetic elements and protein functions in virus pathogenesis and biology.
View Article and Find Full Text PDFBiochemistry (Mosc)
November 2024
Belozersky Institute of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow, 119992, Russia.
To design a safe cellular system for testing inhibitors targeting the RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2, a genetic construct was engineered containing viral cDNA with two blocks of reporter genes while the genes encoding structural S, E, and M proteins were absent. The first reporter block, consisting of Renilla luciferase and green fluorescent protein (Rluc-GFP), was located upstream of the SARS-CoV-2 5'-UTR. Meanwhile, the second block represented by firefly luciferase and red fluorescent protein (Fluc-RFP) was positioned downstream of the transcription regulatory sequence (TRS-N).
View Article and Find Full Text PDFVet Res
September 2024
College of Veterinary Medicine, Konkuk University, Seoul, Republic of Korea.
Feline infectious peritonitis (FIP) is a lethal, immune-mediated disease in cats caused by feline infectious peritonitis virus (FIPV), a biotype of feline coronavirus (FCoV). In contrast to feline enteric coronavirus (FECV), which exclusively infects enterocytes and causes diarrhea, FIPV specifically targets macrophages, resulting in the development of FIP. The transmission and infection mechanisms of this complex, invariably fatal disease remain unclear, with no effective vaccines or approved drugs for its prevention or control.
View Article and Find Full Text PDFHortic Res
September 2024
Nulla Bio Inc., 501 Jinjudaero, Jinju 660-701, Republic of Korea.
-mediated transformation remains a cornerstone of plant biology, fueling advancements in molecular genetics, new genomic techniques (NGTs), and the biotech industry. However, recalcitrant crops and technical hurdles persist as bottlenecks. The goal was to develop super-infective ternary vector systems that integrate a novel salicylic acid-degrading enzyme, GABA, and ethylene-degrading enzymes, targeting the transformation of crops by neutralizing plant defense system on .
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