title: AG-dependent 3'-splice sites are predisposed to aberrant splicing due to a mutation at the first nucleotide of an exon.
abstract: In pre-mRNA splicing, a conserved AG/G at the 3'-splice site is recognized by U2AF(35). A disease-causing mutation abrogating the G nucleotide at the first position of an exon (E(+1)) causes exon skipping in GH1, FECH and EYA1, but not in LPL or HEXA. Knockdown of U2AF(35) enhanced exon skipping in GH1 and FECH. RNA-EMSA revealed that wild-type FECH requires U2AF(35) but wild-type LPL does not. A series of artificial mutations in the polypyrimidine tracts of GH1, FECH, EYA1, LPL and HEXA disclosed that a stretch of at least 10-15 pyrimidines is required to ensure normal splicing in the presence of a mutation at E(+1). Analysis of nine other disease-causing mutations at E(+1) detected five splicing mutations. Our studies suggest that a mutation at the AG-dependent 3'-splice site that requires U2AF(35) for spliceosome assembly causes exon skipping, whereas one at the AG-independent 3'-splice site that does not require U2AF(35) gives rise to normal splicing. The AG-dependence of the 3'-splice site that we analyzed in disease-causing mutations at E(+1) potentially helps identify yet unrecognized splicing mutations at E(+1).
authors.affiliation: Division of Neurogenetics, Center for Neurological Diseases and Cancer, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya 466-8550, Japan.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Fu
authors.forename: Akio
authors.initials: A
authors.lastname: Masuda
authors.forename: Mikako
authors.initials: M
authors.lastname: Ito
authors.forename: Jun
authors.initials: J
authors.lastname: Shinmi
authors.forename: Kinji
authors.initials: K
authors.lastname: Ohno
mesh_terms: D001665:Binding Sites; D004194:Disease / Q000235:genetics; D015536:Down-Regulation; D005091:Exons; D015894:Genome, Human; D057809:HEK293 Cells; D006801:Humans; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism; D009711:Nucleotides / Q000737:chemistry; D011743:Pyrimidines / Q000737:chemistry; D012322:RNA Precursors / Q000378:metabolism; D022821:RNA Splice Sites; D012326:RNA Splicing; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism; D000072668:Splicing Factor U2AF
chemical_list: D009687:Nuclear Proteins; D009711:Nucleotides; D011743:Pyrimidines; D012322:RNA Precursors; D022821:RNA Splice Sites; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D000072668:Splicing Factor U2AF; C076753:U2AF1 protein, human; C000607494:U2AF2 protein, human