title: Two distinct sites are essential for virulent infection and support of variant satellite RNA replication in spontaneous beet black scorch virus variants.
abstract: Spontaneous point mutations of virus genomes are important in RNA virus evolution and often result in modifications of their biological properties. Spontaneous variants of beet black scorch virus (BBSV) and its satellite (sat) RNA were generated from cDNA clones by serial propagation in Chenopodium amaranticolor and Nicotiana benthamiana. Inoculation with recombinant RNAs synthesized in vitro revealed BBSV variants with divergent infectious phenotypes that affected either symptom expression or replication of satRNA variants. Sequence alignments showed a correlation between the phenotypes and distinct BBSV genomic loci in the 3'UTR or in the domain encoding the viral replicase. Comparative analysis between a virulent variant, BBSV-m294, and the wild-type (wt) BBSV by site-directed mutagenesis indicated that a single-nucleotide substitution of a uridine to a guanine at nt 3477 in the 3'UTR was responsible for significant increases in viral pathogenicity. Gain-of-function analyses demonstrated that the ability of the BBSV variants to support replication of variant satRNAs was mainly determined by aa 516 in the P82 replicase. In this case, an arginine substitution for a glutamine residue was essential for high levels of replication, and alterations of other residues surrounding position 516 in the wtBBSV isolate led to only minor phenotypic effects. These results provide evidence that divergence of virus functions affecting pathogenicity and supporting parasitic replication can be determined by a single genetic site, either a nucleotide or an amino acid. The results suggest that complex interactions occur between virus and associated satRNAs during virus evolution.
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Jin
authors.initials: J
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Xianbing
authors.initials: X
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Lindan
authors.initials: L
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Dawei
authors.initials: D
authors.lastname: Li
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Chenggui
authors.initials: C
authors.lastname: Han
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Ziding
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Jialin
authors.initials: J
authors.lastname: Yu
mesh_terms: D020413:3' Untranslated Regions; D001483:Base Sequence; D027461:Beta vulgaris / Q000821:virology*; D027462:Chenopodium / Q000821:virology; D014644:Genetic Variation; D016297:Mutagenesis, Site-Directed; D010935:Plant Diseases / Q000821:virology*; D019244:RNA, Satellite / Q000096:biosynthesis* / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000821:virology; D025641:Tobacco necrosis satellite virus / Q000235:genetics; D019183:Tombusviridae / Q000235:genetics* / Q000472:pathogenicity* / Q000502:physiology; D014774:Virulence / Q000235:genetics
chemical_list: D020413:3' Untranslated Regions; D019244:RNA, Satellite; D012324:RNA-Dependent RNA Polymerase