title: Structural and functional analysis of four non-coding Y RNAs from Chinese hamster cells: identification, molecular dynamics simulations and DNA replication initiation assays.
abstract: BACKGROUND
  The genes coding for Y RNAs are evolutionarily conserved in vertebrates. These non-coding RNAs are essential for the initiation of chromosomal DNA replication in vertebrate cells. However thus far, no information is available about Y RNAs in Chinese hamster cells, which have already been used to detect replication origins and alternative DNA structures around these sites. Here, we report the gene sequences and predicted structural characteristics of the Chinese hamster Y RNAs, and analyze their ability to support the initiation of chromosomal DNA replication in vitro.
  
  
  RESULTS
  We identified DNA sequences in the Chinese hamster genome of four Y RNAs (chY1, chY3, chY4 and chY5) with upstream promoter sequences, which are homologous to the four main types of vertebrate Y RNAs. The chY1, chY3 and chY5 genes were highly conserved with their vertebrate counterparts, whilst the chY4 gene showed a relatively high degree of diversification from the other vertebrate Y4 genes. Molecular dynamics simulations suggest that chY4 RNA is structurally stable despite its evolutionarily divergent predicted stem structure. Of the four Y RNA genes present in the hamster genome, we found that only the chY1 and chY3 RNA were strongly expressed in the Chinese hamster GMA32 cell line, while expression of the chY4 and chY5 RNA genes was five orders of magnitude lower, suggesting that they may in fact not be expressed. We synthesized all four chY RNAs and showed that any of these four could support the initiation of DNA replication in an established human cell-free system.
  
  
  CONCLUSIONS
  These data therefore establish that non-coding chY RNAs are stable structures and can substitute for human Y RNAs in a reconstituted cell-free DNA replication initiation system. The pattern of Y RNA expression and functionality is consistent with Y RNAs of other rodents, including mouse and rat.
authors.affiliation: Departamento de Biotecnologia, Genética e Biologia Celular, Universidade Estadual de Maringá, Av. Colombo 5790, Maringá, Paraná, 87020-900, Brazil. qalneto@gmail.com.
authors.forename: Quirino Alves
authors.initials: QA
authors.lastname: de Lima Neto
authors.affiliation: Departamento de Biotecnologia, Genética e Biologia Celular, Universidade Estadual de Maringá, Av. Colombo 5790, Maringá, Paraná, 87020-900, Brazil. junior.fduarte@gmail.com.
authors.forename: Francisco Ferreira
authors.initials: FF
authors.lastname: Duarte Junior
authors.affiliation: Departamento de Bioquímica, Universidade Estadual de Maringá, UEM, Paraná, 87020-900, Brazil. psabueno@gmail.com.
authors.forename: Paulo Sérgio Alves
authors.initials: PS
authors.lastname: Bueno
authors.affiliation: Departamento de Bioquímica, Universidade Estadual de Maringá, UEM, Paraná, 87020-900, Brazil. favseixas@gmail.com.
authors.forename: Flavio Augusto Vicente
authors.initials: FA
authors.lastname: Seixas
authors.affiliation: Department of Zoology, University of Cambridge, Downing Street, Cambridge, CB2 3EJ, UK. mpcross@stanford.edu.
authors.forename: Madzia Pauline
authors.initials: MP
authors.lastname: Kowalski
authors.affiliation: Department of Zoology, University of Cambridge, Downing Street, Cambridge, CB2 3EJ, UK. eyemen.kheir@gmail.com.
authors.forename: Eyemen
authors.initials: E
authors.lastname: Kheir
authors.affiliation: Department of Zoology, University of Cambridge, Downing Street, Cambridge, CB2 3EJ, UK. tk218@cam.ac.uk.
authors.forename: Torsten
authors.initials: T
authors.lastname: Krude
authors.affiliation: Departamento de Biotecnologia, Genética e Biologia Celular, Universidade Estadual de Maringá, Av. Colombo 5790, Maringá, Paraná, 87020-900, Brazil. aparecidafernandez@gmail.com.
authors.forename: Maria Aparecida
authors.initials: MA
authors.lastname: Fernandez
mesh_terms: D000818:Animals; D002460:Cell Line; D003198:Computer Simulation; D003412:Cricetulus; D004261:DNA Replication; D005786:Gene Expression Regulation; D016678:Genome; D006860:Hydrogen Bonding; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009690:Nucleic Acid Conformation; D022661:RNA, Untranslated / Q000737:chemistry* / Q000235:genetics*; D013329:Structure-Activity Relationship
chemical_list: D022661:RNA, Untranslated