Topic
RNA-dependent RNA polymerase
About: RNA-dependent RNA polymerase is a research topic. Over the lifetime, 13904 publications have been published within this topic receiving 767954 citations. The topic is also known as: RdRp & RNA replicase.
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TL;DR: The three-dimensional crystal structure at 3.3 Å resolution of a decameric, annular ribonucleoprotein complex of the RSV nucleoprotein (N) bound to RNA is determined, which mimics one turn of the viral helical nucleocapsid complex, which serves as template for viral RNA synthesis.
Abstract: The respiratory syncytial virus (RSV) is an important human pathogen, yet neither a vaccine nor effective therapies are available to treat infection. To help elucidate the replication mechanism of this RNA virus, we determined the three-dimensional (3D) crystal structure at 3.3 A resolution of a decameric, annular ribonucleoprotein complex of the RSV nucleoprotein (N) bound to RNA. This complex mimics one turn of the viral helical nucleocapsid complex, which serves as template for viral RNA synthesis. The RNA wraps around the protein ring, with seven nucleotides contacting each N subunit, alternating rows of four and three stacked bases that are exposed and buried within a protein groove, respectively. Combined with electron microscopy data, this structure provides a detailed model for the RSV nucleocapsid, in which the bases are accessible for readout by the viral polymerase. Furthermore, the nucleoprotein structure highlights possible key sites for drug targeting.
301 citations
TL;DR: It is demonstrated that the NS3 and NS5 proteins interact in vivo in dengue virus type 2-infected monkey kidney (CV-1) cells and in HeLa cells coinfected with recombinant vaccinia viruses encoding these proteins as shown by coimmunoprecipitations and immunoblotting methods.
300 citations
TL;DR: Findings reveal how Cas13a of type VI CRISPR-Cas systems defend against RNA phages and set the stage for its development as a tool for RNA manipulation.
300 citations
TL;DR: It is estimated that an active HCV replicase complex consists of one negative-strand RNA, two to ten positive-stranded RNAs, and several hundred nonstructural protein copies, which might be required as structural components of the vesicular compartments that are the site of HCV replication.
Abstract: The hepatitis C virus (HCV) encodes a large polyprotein; therefore, all viral proteins are produced in equimolar amounts regardless of their function. The aim of our study was to determine the ratio of nonstructural proteins to RNA that is required for HCV RNA replication. We analyzed Huh-7 cells harboring full-length HCV genomes or subgenomic replicons and found in all cases a >1,000-fold excess of HCV proteins over positive- and negative-strand RNA. To examine whether all nonstructural protein copies are involved in RNA synthesis, we isolated active HCV replication complexes from replicon cells and examined them for their content of viral RNA and proteins before and after treatment with protease and/or nuclease. In vitro replicase activity, as well as almost the entire negative- and positive-strand RNA, was resistant to nuclease treatment, whereas <5% of the nonstructural proteins were protected from protease digest but accounted for the full in vitro replicase activity. In consequence, only a minor fraction of the HCV nonstructural proteins was actively involved in RNA synthesis at a given time point but, due to the high amounts present in replicon cells, still representing a huge excess compared to the viral RNA. Based on the comparison of nuclease-resistant viral RNA to protease-resistant viral proteins, we estimate that an active HCV replicase complex consists of one negative-strand RNA, two to ten positive-strand RNAs, and several hundred nonstructural protein copies, which might be required as structural components of the vesicular compartments that are the site of HCV replication.
300 citations
TL;DR: It is concluded that Spot 42 RNA is an antisense RNA that acts to differentially regulate genes that are expressed from the same transcription unit and underline the importance of antisense control in bacterial gene regulation.
Abstract: The physiological role of Escherichia coli Spot 42 RNA has remained obscure, even though the 109-nucleotide RNA was discovered almost three decades ago. Structural features of Spot 42 RNA and previous work suggested to us that the RNA might be a regulator of discoordinate gene expression of the galactose operon, a control that is only understood at the phenomenological level. The effects of controlled expression of Spot 42 RNA or deleting the gene (spf) encoding the RNA supported this hypothesis. Down-regulation of galK expression, the third gene in the gal operon, was only observed in the presence of Spot 42 RNA and required growth conditions that caused derepression of the spf gene. Subsequent biochemical studies showed that Spot 42 RNA specifically bound at the galK Shine-Dalgarno region of the galETKM mRNA, thereby blocking ribosome binding. We conclude that Spot 42 RNA is an antisense RNA that acts to differentially regulate genes that are expressed from the same transcription unit. Our results reveal an interesting mechanism by which the expression of a promoter distal gene in an operon can be modulated and underline the importance of antisense control in bacterial gene regulation.
299 citations