title: Global analysis of pre-mRNA subcellular localization following splicing inhibition by spliceostatin A.
abstract: Spliceostatin A (SSA) is a methyl ketal derivative of FR901464, a potent antitumor compound isolated from a culture broth of Pseudomonas sp no. 2663. These compounds selectively bind to the essential spliceosome component SF3b, a subcomplex of the U2 snRNP, to inhibit pre-mRNA splicing. However, the mechanism of SSA's antitumor activity is unknown. It is noteworthy that SSA causes accumulation of a truncated form of the CDK inhibitor protein p27 translated from CDKN1B pre-mRNA, which is involved in SSA-induced cell-cycle arrest. However, it is still unclear whether pre-mRNAs are uniformly exported from the nucleus following SSA treatment. We performed RNA-seq analysis on nuclear and cytoplasmic fractions of SSA-treated cells. Our statistical analyses showed that intron retention is the major consequence of SSA treatment, and a small number of intron-containing pre-mRNAs leak into the cytoplasm. Using a series of reporter plasmids to investigate the roles of intronic sequences in the pre-mRNA leakage, we showed that the strength of the 5' splice site affects pre-mRNA leakage. Additionally, we found that the level of pre-mRNA leakage is related to transcript length. These results suggest that the strength of the 5' splice site and the length of the transcripts are determinants of the pre-mRNA leakage induced by SF3b inhibitors.
authors.affiliation: Chemical Genetics Laboratory, RIKEN, Wako, Saitama 351-0198, Japan.|Division of Gene Expression Mechanism, Institute for Comprehensive Medical Science, Fujita Health University, Kutsukake-cho, Toyoake, Aichi 470-1192, Japan.
authors.forename: Rei
authors.initials: R
authors.lastname: Yoshimoto
authors.affiliation: Chemical Genetics Laboratory, RIKEN, Wako, Saitama 351-0198, Japan.|Frontier Research Core for Life Sciences, University of Toyama, Toyama-shi, Toyama 930-0194, Japan.|Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama-shi, Toyama 930-0194, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Kaida
authors.affiliation: RIKEN Center for Life Science Technologies, Division of Genomic Technologies, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Masaaki
authors.initials: M
authors.lastname: Furuno
authors.affiliation: RIKEN Center for Life Science Technologies, Division of Genomic Technologies, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.|National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894, USA.
authors.forename: A Maxwell
authors.initials: AM
authors.lastname: Burroughs
authors.affiliation: RIKEN Center for Life Science Technologies, Division of Genomic Technologies, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Shohei
authors.initials: S
authors.lastname: Noma
authors.affiliation: RIKEN Center for Life Science Technologies, Division of Genomic Technologies, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Harukazu
authors.initials: H
authors.lastname: Suzuki
authors.affiliation: Chemical Genetics Laboratory, RIKEN, Wako, Saitama 351-0198, Japan.
authors.forename: Yumi
authors.initials: Y
authors.lastname: Kawamura
authors.affiliation: RIKEN Preventive Medicine and Diagnosis Innovation Program (PMI), Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Yoshihide
authors.initials: Y
authors.lastname: Hayashizaki
authors.affiliation: Division of Gene Expression Mechanism, Institute for Comprehensive Medical Science, Fujita Health University, Kutsukake-cho, Toyoake, Aichi 470-1192, Japan.
authors.forename: Akila
authors.initials: A
authors.lastname: Mayeda
authors.affiliation: Chemical Genetics Laboratory, RIKEN, Wako, Saitama 351-0198, Japan.|Japan Science and Technology Corporation, CREST Research Project, Kawaguchi, Saitama 332-0012, Japan.
authors.forename: Minoru
authors.initials: M
authors.lastname: Yoshida
mesh_terms: D002467:Cell Nucleus / Q000235:genetics; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics*; D003593:Cytoplasm / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D009369:Neoplasms / Q000235:genetics*; D011714:Pyrans / Q000494:pharmacology*; D012322:RNA Precursors / Q000235:genetics; D012326:RNA Splicing; D017423:Sequence Analysis, RNA / Q000379:methods*; D013141:Spiro Compounds / Q000494:pharmacology*
chemical_list: C436291:CDKN1B protein, human; D011714:Pyrans; D012322:RNA Precursors; D013141:Spiro Compounds; C553684:spliceostatin A; D050760:Cyclin-Dependent Kinase Inhibitor p27
keywords: RNA-seq; pre-mRNA nuclear retention; pre-mRNA splicing; spliceostatin A