@article{mbs:/content/journal/jgv/10.1099/jgv.0.000941, author = "Amroun, Abdennour and Priet, Stéphane and Querat, Gilles", title = "Toscana virus cap-snatching and initiation of transcription", journal= "Journal of General Virology", year = "2017", volume = "98", number = "11", pages = "2676-2688", doi = "https://doi.org/10.1099/jgv.0.000941", url = "https://www.microbiologyresearch.org/content/journal/jgv/10.1099/jgv.0.000941", publisher = "Microbiology Society", issn = "1465-2099", type = "Journal Article", keywords = "cap-snatching", keywords = "prime-and-realign", keywords = "Toscana virus", keywords = "Phenuiviridae", keywords = "phleboviruses", keywords = "transcription initiation", abstract = "Toscana virus (TOSV) is an arthropod-borne phlebovirus within the family Phenuiviridae in the order Bunyavirales. It seems to be an important agent of human meningoencephalitis in the warm season in the Mediterranean area. Because the polymerase of Bunyavirales lacks a capping activity, it cleaves short-capped RNA leaders derived from the host cell, and uses them to initiate viral mRNA synthesis. To determine the size and nucleotide composition of the host-derived RNA leaders, and to elucidate the first steps of TOSV transcription initiation, we performed a high-throughput sequencing of the 5′ end of TOSV mRNAs in infected cells at different times post-infection. Our results indicated that the viral polymerase cleaved the host-capped RNA leaders within a window of 11–16 nucleotides. A single population of cellular mRNAs could be cleaved at different sites to prime the synthesis of several viral mRNA species. The majority of the mRNA resulted from direct priming, but we observed mRNAs resulting from several rounds of prime-and-realign events. Our data suggest that the different rounds of the prime-and-realign mechanism result from the blocking of the template strand in a static position in the active site, leading to the slippage of the nascent strand by two nucleotides when the growing duplex is sorted out from the active site. To minimize this rate-limiting step, TOSV polymerase cleaves preferentially capped RNA leaders after GC, so as to greatly reduce the number of cycles of priming and realignment, and facilitate the separation of the growing duplex.", }