title: CWC22-dependent pre-mRNA splicing and eIF4A3 binding enables global deposition of exon junction complexes.
abstract: In metazoan cells, spliced mRNAs are marked by the exon junction complex (EJC), a multi-protein complex that serves as a key regulator of post-transcriptional mRNA metabolism. Deposition of EJCs on mRNA is intimately linked to the splicing process. The spliceosomal protein CWC22 directly binds the core EJC-protein eIF4A3, guides it to the spliceosome and initiates EJC assembly. In addition, CWC22 is involved in the splicing process itself, but the molecular details of its dual function remain elusive. Here we analyze the mechanisms, by which CWC22 co-regulates pre-mRNA splicing and EJC assembly. We show that the core of CWC22 is sufficient to mediate both pre-mRNA splicing and EJC assembly. Nonetheless, both processes can be functionally uncoupled with an eIF4A3-binding deficient mutant of CWC22, which impedes EJC assembly. A C-terminal domain of CWC22 strongly enhances its spliceosomal interaction and likely regulates its function. High-throughput RNA-sequencing identifies global defects of pre-mRNA splicing and downregulation of diverse gene expression pathways in CWC22-depleted cells. We propose a model, in which CWC22 represents an integral component of the spliceosome and orchestrates pre-mRNA splicing and eIF4A3 binding to achieve global assembly of exon junction complexes.
authors: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany.
authors: Anna-Lena
authors: AL
authors: Steckelberg
authors: Cologne Center for Genomics (CCG), University of Cologne, D-50931 Cologne, Germany Institute of Human Genetics, University of Cologne, D-50931 Cologne, Germany.
authors: Janine
authors: J
authors: Altmueller
authors: Max Planck Institute for Biology of Ageing, Joseph-Stelzmann-Straße 9b, D-50931 Cologne, Germany.
authors: Christoph
authors: C
authors: Dieterich
authors: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany ngehring@uni-koeln.de.
authors: Niels H
authors: NH
authors: Gehring
mesh_terms: D001665:Binding Sites; D053487:DEAD-box RNA Helicases / Q000378:metabolism*; D039601:Eukaryotic Initiation Factor-4A / Q000378:metabolism*; D005091:Exons; D015870:Gene Expression; D057809:HEK293 Cells; D006801:Humans; D007438:Introns; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019696:Peptidylprolyl Isomerase; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012322:RNA Precursors / Q000378:metabolism*; D012326:RNA Splicing; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins; D017381:Spliceosomes / Q000378:metabolism*
chemical_list: C578086:CWC22 protein, human; D009687:Nuclear Proteins; D012322:RNA Precursors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D039601:Eukaryotic Initiation Factor-4A; C403493:EIF4A3 protein, human; D053487:DEAD-box RNA Helicases; D019696:Peptidylprolyl Isomerase