title: Relationships and Evolution of Double-Stranded RNA Totiviruses of Yeasts Inferred from Analysis of L-A-2 and L-BC Variants in Wine Yeast Strain Populations.
abstract: Saccharomyces cerevisiae killer strains secrete a protein toxin active on nonkiller strains of the same (or other) yeast species. Different killer toxins, K1, K2, K28, and Klus, have been described. Each toxin is encoded by a medium-size (1.5- to 2.3-kb) M double-stranded RNA (dsRNA) located in the cytoplasm. M dsRNAs require L-A helper virus for maintenance. L-A belongs to the Totiviridae family, and its dsRNA genome of 4.6 kb codes for the major capsid protein Gag and a minor Gag-Pol protein, which form the virions that separately encapsidate L-A or the M satellites. Different L-A variants exist in nature; on average, 24% of their nucleotides are different. Previously, we reported that L-A-lus was specifically associated with Mlus, suggesting coevolution, and proposed a role of the toxin-encoding M dsRNAs in the appearance of new L-A variants. Here we confirm this by analyzing the helper virus in K2 killer wine strains, which we named L-A-2. L-A-2 is required for M2 maintenance, and neither L-A nor L-A-lus shows helper activity for M2 in the same genetic background. This requirement is overcome when coat proteins are provided in large amounts by a vector or in ski mutants. The genome of another totivirus, L-BC, frequently accompanying L-A in the same cells shows a lower degree of variation than does L-A (about 10% of nucleotides are different). Although L-BC has no helper activity for M dsRNAs, distinct L-BC variants are associated with a particular killer strain. The so-called L-BC-lus (in Klus strains) and L-BC-2 (in K2 strains) are analyzed.
  
  
  
  Killer strains of S. cerevisiae secrete protein toxins that kill nonkiller yeasts. The "killer phenomenon" depends on two dsRNA viruses: L-A and M. M encodes the toxin, and L-A, the helper virus, provides the capsids for both viruses. Different killer toxins exist: K1, K2, K28, and Klus, encoded on different M viruses. Our data indicate that each M dsRNA depends on a specific helper virus; these helper viruses have nucleotide sequences that may be as much as 26% different, suggesting coevolution. In wine environments, K2 and Klus strains frequently coexist. We have previously characterized the association of Mlus and L-A-lus. Here we sequence and characterize L-A-2, the helper virus of M2, establishing the helper virus requirements of M2, which had not been completely elucidated. We also report the existence of two specific L-BC totiviruses in Klus and K2 strains with about 10% of their nucleotides different, suggesting different evolutionary histories from those of L-A viruses.
authors.affiliation: Instituto de Biología Funcional y Genómica (IBFG), Consejo Superior de Investigaciones Científicas/Universidad de Salamanca, Salamanca, Spain.|Escuela Politécnica Superior de Zamora, Universidad de Salamanca, Zamora, Spain.
authors.forename: Nieves
authors.initials: N
authors.lastname: Rodríguez-Cousiño
authors.affiliation: Instituto de Biología Funcional y Genómica (IBFG), Consejo Superior de Investigaciones Científicas/Universidad de Salamanca, Salamanca, Spain mrosa@usal.es.
authors.forename: Rosa
authors.initials: R
authors.lastname: Esteban
mesh_terms: D000935:Antifungal Agents / Q000378:metabolism*; D002213:Capsid / Q000378:metabolism; D015685:Fusion Proteins, gag-pol / Q000235:genetics; D015683:Gene Products, gag / Q000235:genetics; D006378:Helper Viruses / Q000235:genetics*; D008928:Mitochondria / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000378:metabolism / Q000821:virology*; D012526:Satellite Viruses / Q000235:genetics; D019197:Totivirus / Q000235:genetics*
chemical_list: D000935:Antifungal Agents; D015685:Fusion Proteins, gag-pol; D015683:Gene Products, gag
keywords: L-A helper virus; double-stranded RNA virus; yeast killer toxins; yeast virus; yeast wine strains