title: A novel intra-U1 snRNP cross-regulation mechanism: alternative splicing switch links U1C and U1-70K expression.
abstract: The U1 small nuclear ribonucleoprotein (snRNP)-specific U1C protein participates in 5' splice site recognition and regulation of pre-mRNA splicing. Based on an RNA-Seq analysis in HeLa cells after U1C knockdown, we found a conserved, intra-U1 snRNP cross-regulation that links U1C and U1-70K expression through alternative splicing and U1 snRNP assembly. To investigate the underlying regulatory mechanism, we combined mutational minigene analysis, in vivo splice-site blocking by antisense morpholinos, and in vitro binding experiments. Alternative splicing of U1-70K pre-mRNA creates the normal (exons 7-8) and a non-productive mRNA isoform, whose balance is determined by U1C protein levels. The non-productive isoform is generated through a U1C-dependent alternative 3' splice site, which requires an adjacent cluster of regulatory 5' splice sites and binding of intact U1 snRNPs. As a result of nonsense-mediated decay (NMD) of the non-productive isoform, U1-70K mRNA and protein levels are down-regulated, and U1C incorporation into the U1 snRNP is impaired. U1-70K/U1C-deficient particles are assembled, shifting the alternative splicing balance back towards productive U1-70K splicing, and restoring assembly of intact U1 snRNPs. Taken together, we established a novel feedback regulation that controls U1-70K/U1C homeostasis and ensures correct U1 snRNP assembly and function.
authors.affiliation: Institute of Biochemistry, Justus Liebig University of Giessen, Giessen, Germany.
authors.forename: Tanja Dorothe
authors.initials: TD
authors.lastname: Rösel-Hillgärtner
authors.forename: Lee-Hsueh
authors.initials: LH
authors.lastname: Hung
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Khrameeva
authors.forename: Patrick
authors.initials: P
authors.lastname: Le Querrec
authors.forename: Mikhail S
authors.initials: MS
authors.lastname: Gelfand
authors.forename: Albrecht
authors.initials: A
authors.lastname: Bindereif
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004252:DNA Mutational Analysis; D004625:Embryo, Nonmammalian; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D012322:RNA Precursors / Q000235:genetics*; D022821:RNA Splice Sites / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics*; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000235:genetics*; D017381:Spliceosomes / Q000235:genetics; D015027:Zebrafish / Q000235:genetics / Q000254:growth & development
chemical_list: D012322:RNA Precursors; D022821:RNA Splice Sites; D017412:Ribonucleoprotein, U1 Small Nuclear