title: A novel assay identifies transcript elongation roles for the Nup84 complex and RNA processing factors.
abstract: To clarify the role of a number of mRNA processing factors in transcription elongation, we developed an in vivo assay for direct analysis of elongation on chromatin. The assay relies on two substrates containing two G-less cassettes separated by either a long and GC-rich or a short and GC-poor DNA sequence (G-less-based run-on (GLRO) assay). We demonstrate that PAF, THSC/TREX-2, SAGA, the exosome component Rrp6 and two subunits of cleavage factor IA (Rna14 and Rna15) are required for efficient transcription elongation, in contrast to some results obtained using other assays. Next, we undertook a mutant screen and found out that the Nup84 nucleoporin complex is also required for transcription elongation, as confirmed by the GLRO assay and RNA polymerase II chromatin immunoprecipitations. Therefore, in addition to showing that the GLRO assay is a sensitive and reliable method for the analysis of elongation in vivo, this study provides evidence for a new role of the Nup84 complex and a number of mRNA processing factors in transcription elongation that supports a connection of pre-mRNA processing and nuclear export with transcription elongation.
authors.affiliation: Centro Andaluz de Biología Molecular y Medicina Regenerativa CABIMER, Universidad de Sevilla-CSIC, Sevilla, Spain.
authors.forename: Cristina
authors.initials: C
authors.lastname: Tous
authors.forename: Ana G
authors.initials: AG
authors.lastname: Rondón
authors.forename: María
authors.initials: M
authors.lastname: García-Rubio
authors.forename: Cristina
authors.initials: C
authors.lastname: González-Aguilera
authors.forename: Rosa
authors.initials: R
authors.lastname: Luna
authors.forename: Andrés
authors.initials: A
authors.lastname: Aguilera
mesh_terms: D047369:Chromatin Immunoprecipitation; D004251:DNA Transposable Elements; D055786:Gene Knockout Techniques; D008967:Molecular Biology / Q000379:methods; D016254:Mutagenesis, Insertional; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism*; D012331:RNA, Fungal / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
chemical_list: D004251:DNA Transposable Elements; C097894:NUP84 protein, S cerevisiae; D028861:Nuclear Pore Complex Proteins; D012331:RNA, Fungal; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins