title: Ty1 Integrase Interacts with RNA Polymerase III-specific Subcomplexes to Promote Insertion of Ty1 Elements Upstream of Polymerase (Pol) III-transcribed Genes.
abstract: Retrotransposons are eukaryotic mobile genetic elements that transpose by reverse transcription of an RNA intermediate and are derived from retroviruses. The Ty1 retrotransposon of Saccharomyces cerevisiae belongs to the Ty1/Copia superfamily, which is present in every eukaryotic genome. Insertion of Ty1 elements into the S. cerevisiae genome, which occurs upstream of genes transcribed by RNA Pol III, requires the Ty1 element-encoded integrase (IN) protein. Here, we report that Ty1-IN interacts in vivo and in vitro with RNA Pol III-specific subunits to mediate insertion of Ty1 elements upstream of Pol III-transcribed genes. Purification of Ty1-IN from yeast cells followed by mass spectrometry (MS) analysis identified an enrichment of peptides corresponding to the Rpc82/34/31 and Rpc53/37 Pol III-specific subcomplexes. GFP-Trap purification of multiple GFP-tagged RNA Pol III subunits from yeast extracts revealed that the majority of Pol III subunits co-purify with Ty1-IN but not two other complexes required for Pol III transcription, transcription initiation factors (TF) IIIB and IIIC. In vitro binding studies with bacterially purified RNA Pol III proteins demonstrate that Rpc31, Rpc34, and Rpc53 interact directly with Ty1-IN. Deletion of the N-terminal 280 amino acids of Rpc53 abrogates insertion of Ty1 elements upstream of the hot spot SUF16 tRNA locus and abolishes the interaction of Ty1-IN with Rpc37. The Rpc53/37 complex therefore has an important role in targeting Ty1-IN to insert Ty1 elements upstream of Pol III-transcribed genes.
authors.affiliation: From the Department of Biochemistry and Molecular Biology, Wine Research Centre, and.
authors.forename: Stephanie
authors.initials: S
authors.lastname: Cheung
authors.affiliation: Wine Research Centre, and.
authors.forename: Lina
authors.initials: L
authors.lastname: Ma
authors.affiliation: Centre for High-Throughput Biology, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada and.
authors.forename: Patrick H W
authors.initials: PH
authors.lastname: Chan
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan 115.
authors.forename: Hui-Lan
authors.initials: HL
authors.lastname: Hu
authors.affiliation: From the Department of Biochemistry and Molecular Biology, Centre for High-Throughput Biology, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada and.
authors.forename: Thibault
authors.initials: T
authors.lastname: Mayor
authors.affiliation: Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan 115.
authors.forename: Hung-Ta
authors.initials: HT
authors.lastname: Chen
authors.affiliation: From the Department of Biochemistry and Molecular Biology, Wine Research Centre, and vmeasday@mail.ubc.ca.
authors.forename: Vivien
authors.initials: V
authors.lastname: Measday
mesh_terms: D019426:Integrases / Q000737:chemistry / Q000502:physiology*; D016254:Mutagenesis, Insertional; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D021122:Protein Subunits / Q000378:metabolism; D012320:RNA Polymerase III / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018626:Retroelements; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D014158:Transcription, Genetic
chemical_list: D021122:Protein Subunits; D018626:Retroelements; D029701:Saccharomyces cerevisiae Proteins; D019426:Integrases; C508373:RPC53 protein, S cerevisiae; D012320:RNA Polymerase III
keywords: RNA polymerase III; Saccharomyces cerevisiae; Ty1 element; integrase; retrotransposon; retrovirus; tRNA gene; transposable element (TE)