title: A novel 3' splice site recognition by the two zinc fingers in the U2AF small subunit.
abstract: The pre-mRNA splicing reaction of eukaryotic cells has to be carried out extremely accurately, as failure to recognize the splice sites correctly causes serious disease. The small subunit of the U2AF heterodimer is essential for the determination of 3' splice sites in pre-mRNA splicing, and several single-residue mutations of the U2AF small subunit cause severe disorders such as myelodysplastic syndromes. However, the mechanism of RNA recognition is poorly understood. Here we solved the crystal structure of the U2AF small subunit (U2AF23) from fission yeast, consisting of an RNA recognition motif (RRM) domain flanked by two conserved CCCH-type zinc fingers (ZFs). The two ZFs are positioned side by side on the β sheet of the RRM domain. Further mutational analysis revealed that the ZFs bind cooperatively to the target RNA sequence, but the RRM domain acts simply as a scaffold to organize the ZFs and does not itself contact the RNA directly. This completely novel and unexpected mode of RNA-binding mechanism by the U2AF small subunit sheds light on splicing errors caused by mutations of this highly conserved protein.
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, Tsurumi-ku, Yokohama 230-0045, Japan; Drug Design Group, Kanagawa Academy of Science and Technology (KAST), Takatsu-ku, Kawasaki 213-0012, Japan;
authors.forename: Hisashi
authors.initials: H
authors.lastname: Yoshida
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, Tsurumi-ku, Yokohama 230-0045, Japan; Drug Design Group, Kanagawa Academy of Science and Technology (KAST), Takatsu-ku, Kawasaki 213-0012, Japan;
authors.forename: Sam-Yong
authors.initials: SY
authors.lastname: Park
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, Tsurumi-ku, Yokohama 230-0045, Japan;
authors.forename: Takashi
authors.initials: T
authors.lastname: Oda
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, Tsurumi-ku, Yokohama 230-0045, Japan;
authors.forename: Taeko
authors.initials: T
authors.lastname: Akiyoshi
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, Tsurumi-ku, Yokohama 230-0045, Japan;
authors.forename: Mamoru
authors.initials: M
authors.lastname: Sato
authors.affiliation: Center for Life Science Technologies, Yokohama Institute, RIKEN, Tsurumi-ku, Yokohama 230-0045, Japan;
authors.forename: Mikako
authors.initials: M
authors.lastname: Shirouzu
authors.affiliation: Center for Life Science Technologies, Yokohama Institute, RIKEN, Tsurumi-ku, Yokohama 230-0045, Japan;
authors.forename: Kengo
authors.initials: K
authors.lastname: Tsuda
authors.affiliation: Center for Life Science Technologies, Yokohama Institute, RIKEN, Tsurumi-ku, Yokohama 230-0045, Japan; Faculty of Pharmacy and Research Institute of Pharmaceutical Science, Musashino University, Nishitokyo-shi, Tokyo 202-8585, Japan;
authors.forename: Kanako
authors.initials: K
authors.lastname: Kuwasako
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, Tsurumi-ku, Yokohama 230-0045, Japan;
authors.forename: Satoru
authors.initials: S
authors.lastname: Unzai
authors.affiliation: Center for Life Science Technologies, Yokohama Institute, RIKEN, Tsurumi-ku, Yokohama 230-0045, Japan; Faculty of Pharmacy and Research Institute of Pharmaceutical Science, Musashino University, Nishitokyo-shi, Tokyo 202-8585, Japan;
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Muto
authors.affiliation: Department of Biochemistry, Shimane University School of Medicine, Izumo 693-8501, Japan.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Urano
authors.affiliation: Department of Biochemistry, Shimane University School of Medicine, Izumo 693-8501, Japan.
authors.forename: Eiji
authors.initials: E
authors.lastname: Obayashi
mesh_terms: D020816:Amino Acid Motifs; D001665:Binding Sites; D004252:DNA Mutational Analysis; D008958:Models, Molecular; D009687:Nuclear Proteins / Q000737:chemistry*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D022821:RNA Splice Sites; D012261:Ribonucleoproteins / Q000737:chemistry*; D012568:Schizosaccharomyces / Q000737:chemistry / Q000502:physiology*; D000072668:Splicing Factor U2AF; D016335:Zinc Fingers / Q000502:physiology*
chemical_list: D009687:Nuclear Proteins; D022821:RNA Splice Sites; D012261:Ribonucleoproteins; D000072668:Splicing Factor U2AF
keywords: 3′ splice site; RNA splicing; U2AF small subunit