title: A short conserved motif in ALYREF directs cap- and EJC-dependent assembly of export complexes on spliced mRNAs.
abstract: The export of messenger RNAs (mRNAs) is the final of several nuclear posttranscriptional steps of gene expression. The formation of export-competent mRNPs involves the recruitment of export factors that are assumed to facilitate transport of the mature mRNAs. Using in vitro splicing assays, we show that a core set of export factors, including ALYREF, UAP56 and DDX39, readily associate with the spliced RNAs in an EJC (exon junction complex)- and cap-dependent manner. In order to elucidate how ALYREF and other export adaptors mediate mRNA export, we conducted a computational analysis and discovered four short, conserved, linear motifs present in RNA-binding proteins. We show that mutation in one of the new motifs (WxHD) in an unstructured region of ALYREF reduced RNA binding and abolished the interaction with eIF4A3 and CBP80. Additionally, the mutation impaired proper localization to nuclear speckles and export of a spliced reporter mRNA. Our results reveal important details of the orchestrated recruitment of export factors during the formation of export competent mRNPs.
authors.affiliation: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany.
authors.forename: Agnieszka M
authors.initials: AM
authors.lastname: Gromadzka
authors.affiliation: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany.
authors.forename: Anna-Lena
authors.initials: AL
authors.lastname: Steckelberg
authors.affiliation: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany.
authors.forename: Kusum K
authors.initials: KK
authors.lastname: Singh
authors.affiliation: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany.
authors.forename: Kay
authors.initials: K
authors.lastname: Hofmann
authors.affiliation: Institute for Genetics, University of Cologne, D-50674 Cologne, Germany ngehring@uni-koeln.de.
authors.forename: Niels H
authors.initials: NH
authors.lastname: Gehring
mesh_terms: D020816:Amino Acid Motifs; D001692:Biological Transport; D003001:Cloning, Molecular; D017124:Conserved Sequence; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D039601:Eukaryotic Initiation Factor-4A / Q000235:genetics / Q000378:metabolism; D005091:Exons; D015870:Gene Expression; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D039502:Nuclear Cap-Binding Protein Complex / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
chemical_list: C105242:ALYREF protein, human; D039502:Nuclear Cap-Binding Protein Complex; D009687:Nuclear Proteins; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D012261:Ribonucleoproteins; D014157:Transcription Factors; C026452:messenger ribonucleoprotein; D039601:Eukaryotic Initiation Factor-4A; C498428:DDX39A protein, human; C487886:DDX39B protein, human; C403493:EIF4A3 protein, human; D053487:DEAD-box RNA Helicases