title: Activation-Dependent TRAF3 Exon 8 Alternative Splicing Is Controlled by CELF2 and hnRNP C Binding to an Upstream Intronic Element.
abstract: Cell-type-specific and inducible alternative splicing has a fundamental impact on regulating gene expression and cellular function in a variety of settings, including activation and differentiation. We have recently shown that activation-induced skipping of TRAF3 exon 8 activates noncanonical NF-κB signaling upon T cell stimulation, but the regulatory basis for this splicing event remains unknown. Here we identifycis- andtrans-regulatory elements rendering this splicing switch activation dependent and cell type specific. Thecis-acting element is located 340 to 440 nucleotides upstream of the regulated exon and acts in a distance-dependent manner, since altering the location reduces its activity. A small interfering RNA screen, followed by cross-link immunoprecipitation and mutational analyses, identified CELF2 and hnRNP C astrans-acting factors that directly bind the regulatory sequence and together mediate increased exon skipping in activated T cells. CELF2 expression levels correlate with TRAF3 exon skipping in several model systems, suggesting that CELF2 is the decisive factor, with hnRNP C being necessary but not sufficient. These data suggest an interplay between CELF2 and hnRNP C as the mechanistic basis for activation-dependent alternative splicing of TRAF3 exon 8 and additional exons and uncover an intronic splicing silencer whose full activity depends on the precise location more than 300 nucleotides upstream of the regulated exon.
authors.affiliation: Freie Universität Berlin, Institute of Chemistry and Biochemistry, Berlin, Germany.
authors.forename: Astrid-Solveig
authors.initials: AS
authors.lastname: Schultz
authors.affiliation: Freie Universität Berlin, Institute of Chemistry and Biochemistry, Berlin, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Preussner
authors.affiliation: Max Delbrück Center for Molecular Medicine, Berlin, Germany.
authors.forename: Mario
authors.initials: M
authors.lastname: Bunse
authors.affiliation: Department of Biochemistry and Molecular Biology, Institute for Medical Research Israel-Canada, Hebrew University-Hadassah Medical School, Ein Karem, Jerusalem, Israel.
authors.forename: Rotem
authors.initials: R
authors.lastname: Karni
authors.affiliation: Freie Universität Berlin, Institute of Chemistry and Biochemistry, Berlin, Germany florian.heyd@fu-berlin.de.
authors.forename: Florian
authors.initials: F
authors.lastname: Heyd
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D001665:Binding Sites; D000067878:CELF Proteins / Q000378:metabolism*; D005091:Exons / Q000235:genetics*; D057809:HEK293 Cells; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C / Q000378:metabolism*; D006801:Humans; D007438:Introns / Q000235:genetics*; D008213:Lymphocyte Activation / Q000235:genetics*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D011072:Poly U / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D034741:RNA, Small Interfering / Q000378:metabolism; D037941:Silencer Elements, Transcriptional / Q000235:genetics; D013601:T-Lymphocytes / Q000276:immunology; D048008:TNF Receptor-Associated Factor 3 / Q000235:genetics* / Q000378:metabolism
chemical_list: D000067878:CELF Proteins; C117363:CELF2 protein, human; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C; D009419:Nerve Tissue Proteins; D034741:RNA, Small Interfering; D048008:TNF Receptor-Associated Factor 3; D011072:Poly U
keywords: RNA binding proteins; RNA splicing