title: Enhancer-dependent 5'-splice site control of fruitless pre-mRNA splicing.
abstract: The Drosophila fruitless (fru) gene encodes a transcription factor that essentially regulates all aspects of male courtship behavior. The use of alternative 5'-splice sites generates fru isoforms that determine gender-appropriate sexual behaviors. Alternative splicing of fru is regulated by TRA and TRA2 and depends on an exonic splicing enhancer (fruRE) consisting of three 13-nucleotide repeat elements, nearly identical to those that regulate alternative sex-specific 3'-splice site choice in the doublesex (dsx) gene. dsx has provided a useful model system to investigate the mechanisms of enhancer-dependent 3'-splice site choice. However, little is known about enhancer-dependent regulation of alternative 5'-splice sites. The mechanisms of this process were investigated using an in vitro system in which recombinant TRA/TRA2 could activate the female-specific 5'-splice site of fru. Mutational analysis demonstrated that one 13-nucleotide repeat element within the fruRE is required and sufficient to activate the regulated female-specific splice site. As was established for dsx, the fruRE can be replaced by a short element encompassing tandem 13-nucleotide repeat elements, by heterologous splicing enhancers, and by artificially tethering a splicing activator to the pre-mRNA. Complementation experiments showed that Ser/Arg-rich proteins facilitate enhancer-dependent 5'-splice site activation. We conclude that splicing enhancers function similarly in activating regulated 5'- and 3'-splice sites. These results suggest that exonic splicing enhancers recruit multiple spliceosomal components required for the initial recognition of 5'- and 3'-splice sites.
authors.affiliation: Department of Microbiology and Molecular Genetics, College of Medicine, University of California, Irvine, California 92697-4025, USA.
authors.forename: Bianca J
authors.initials: BJ
authors.lastname: Lam
authors.forename: Arati
authors.initials: A
authors.lastname: Bakshi
authors.forename: Fatma Y
authors.initials: FY
authors.lastname: Ekinci
authors.forename: Jenny
authors.initials: J
authors.lastname: Webb
authors.forename: Brenton R
authors.initials: BR
authors.lastname: Graveley
authors.forename: Klemens J
authors.initials: KJ
authors.lastname: Hertel
mesh_terms: D020121:5' Untranslated Regions / Q000235:genetics*; D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D029721:Drosophila Proteins / Q000235:genetics*; D004331:Drosophila melanogaster / Q000235:genetics*; D004742:Enhancer Elements, Genetic; D005260:Female; D005786:Gene Expression Regulation; D009419:Nerve Tissue Proteins / Q000235:genetics*; D012322:RNA Precursors / Q000235:genetics*; D012727:Sex Characteristics; D014157:Transcription Factors / Q000235:genetics*
chemical_list: D020121:5' Untranslated Regions; D017931:DNA Primers; D029721:Drosophila Proteins; D009419:Nerve Tissue Proteins; D012322:RNA Precursors; D014157:Transcription Factors; C103226:fru protein, Drosophila