title: An intronic RNA structure modulates expression of the mRNA biogenesis factor Sus1.
abstract: Sus1 is a conserved protein involved in chromatin remodeling and mRNA biogenesis. Unlike most yeast genes, the SUS1 pre-mRNA of Saccharomyces cerevisiae contains two introns and is alternatively spliced, retaining one or both introns in response to changes in environmental conditions. SUS1 splicing may allow the cell to control Sus1 expression, but the mechanisms that regulate this process remain unknown. Using in silico analyses together with NMR spectroscopy, gel electrophoresis, and UV thermal denaturation experiments, we show that the downstream intron (I2) of SUS1 forms a weakly stable, 37-nucleotide stem-loop structure containing the branch site near its apical loop and the 3' splice site after the stem terminus. A cellular assay revealed that two of four mutants containing altered I2 structures had significantly impaired SUS1 expression. Semiquantitative RT-PCR experiments indicated that all mutants accumulated unspliced SUS1 pre-mRNA and/or induced distorted levels of fully spliced mRNA relative to wild type. Concomitantly, Sus1 cellular functions in histone H2B deubiquitination and mRNA export were affected in I2 hairpin mutants that inhibited splicing. This work demonstrates that I2 structure is relevant for SUS1 expression, and that this effect is likely exerted through modulation of splicing.
authors.affiliation: Gene Expression and RNA Metabolism Laboratory, Centro de Investigación Príncipe Felipe, Valencia 46012, Spain Facultad de Medicina, Universidad Católica de Valencia, Valencia 46001, Spain.
authors.forename: Ali
authors.initials: A
authors.lastname: AbuQattam
authors.affiliation: Facultad de Medicina, Universidad Católica de Valencia, Valencia 46001, Spain.
authors.forename: José
authors.initials: J
authors.lastname: Gallego
authors.affiliation: Gene Expression and RNA Metabolism Laboratory, Centro de Investigación Príncipe Felipe, Valencia 46012, Spain.
authors.forename: Susana
authors.initials: S
authors.lastname: Rodríguez-Navarro
mesh_terms: D006358:Hot Temperature; D007438:Introns; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D012326:RNA Splicing; D012331:RNA, Fungal / Q000737:chemistry*; D012333:RNA, Messenger / Q000096:biosynthesis*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*
chemical_list: D009687:Nuclear Proteins; D012331:RNA, Fungal; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; C481300:Sus1 protein, S cerevisiae
keywords: NMR; RNA; Sus1; gene expression; splicing; structure; thermal stability; yeast