title: Identification of cis-acting elements on positive-strand subgenomic mRNA required for the synthesis of negative-strand counterpart in bovine coronavirus.
abstract: It has been demonstrated that, in addition to genomic RNA, sgmRNA is able to serve as a template for the synthesis of the negative-strand [(-)-strand] complement. However, the cis-acting elements on the positive-strand [(+)-strand] sgmRNA required for (-)-strand sgmRNA synthesis have not yet been systematically identified. In this study, we employed real-time quantitative reverse transcription polymerase chain reaction to analyze the cis-acting elements on bovine coronavirus (BCoV) sgmRNA 7 required for the synthesis of its (-)-strand counterpart by deletion mutagenesis. The major findings are as follows. (1) Deletion of the 5'-terminal leader sequence on sgmRNA 7 decreased the synthesis of the (-)-strand sgmRNA complement. (2) Deletions of the 3' untranslated region (UTR) bulged stem-loop showed no effect on (-)-strand sgmRNA synthesis; however, deletion of the 3' UTR pseudoknot decreased the yield of (-)-strand sgmRNA. (3) Nucleotides positioned from -15 to -34 of the sgmRNA 7 3'-terminal region are required for efficient (-)-strand sgmRNA synthesis. (4) Nucleotide species at the 3'-most position (-1) of sgmRNA 7 is correlated to the efficiency of (-)-strand sgmRNA synthesis. These results together suggest, in principle, that the 5'- and 3'-terminal sequences on sgmRNA 7 harbor cis-acting elements are critical for efficient (-)-strand sgmRNA synthesis in BCoV.
authors: Graduate Institute of Veterinary Pathobiology, College of Veterinary Medicine, National Chung-Hsing University, Taichung 40227, Taiwan. ybu999@gmail.com.
authors: Po-Yuan
authors: PY
authors: Yeh
authors: Graduate Institute of Veterinary Pathobiology, College of Veterinary Medicine, National Chung-Hsing University, Taichung 40227, Taiwan. hwu2@dragon.nchu.edu.tw.
authors: Hung-Yi
authors: HY
authors: Wu
mesh_terms: D017938:Coronavirus, Bovine / Q000235:genetics*; D004252:DNA Mutational Analysis; D012333:RNA, Messenger / Q000235:genetics*; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics*; D050436:Regulatory Elements, Transcriptional; D017384:Sequence Deletion; D014158:Transcription, Genetic
chemical_list: D012333:RNA, Messenger; D012367:RNA, Viral