The exon junction complex is detected on CBP80-bound but not eIF4E-bound mRNA in mammalian cells: dynamics of mRNP remodeling.
Newly spliced mRNAs in mammalian cells are characterized by a complex of proteins at exon-exon junctions. This complex recruits Upf3 and Upf2, which function in nonsense-mediated mRNA decay (NMD). Both Upf proteins are detected on mRNA bound by the major nuclear cap-binding proteins CBP80/CBP20 but not mRNA bound by the major cytoplasmic cap-binding protein eIF4E. These and other data indicate that NMD targets CBP80-bound mRNA during a 'pioneer' round of translation, but whether nuclear eIF4E also binds nascent but dead-end transcripts is unclear. Here we provide evidence that nuclear CBP80 but not nuclear eIF4E is readily detected in association with intron-containing RNA and the C-terminal domain of RNA polymerase II. Consistent with this evidence, we demonstrate that RNPS1, Y14, SRm160, REF/Aly, TAP, Upf3X and Upf2 are detected in the nuclear fraction on CBP80-bound but not eIF4E-bound mRNA. Each of these proteins is also detected on CBP80-bound mRNA in the cytoplasmic fraction, indicating a presence on mRNA after export. The dynamics of mRNP composition before and after mRNA export are discussed.
Lejeune
Fabrice
F
Department of Biochemistry and Biophysics, School of Medicine and Dentistry, 601 Elmwood Avenue, Box 712, University of Rochester, Rochester, NY 14642, USA.
Ishigaki
Yasuhito
Y
Li
Xiaojie
X
Maquat
Lynne E
LE
D000818:Animals; D034961:Antigens, Nuclear; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics; D001692:Biological Transport; D019556:COS Cells / Q000378:metabolism; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D039561:Eukaryotic Initiation Factor-4E; D005091:Exons / Q000235:genetics; D005914:Globins / Q000235:genetics; D007438:Introns / Q000235:genetics; D046911:Macromolecular Substances; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins / Q000378:metabolism; D010448:Peptide Initiation Factors / Q000378:metabolism*; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D039381:RNA Cap-Binding Proteins; D012315:RNA Caps / Q000378:metabolism*; D012319:RNA Polymerase II / Q000378:metabolism; D012322:RNA Precursors / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012261:Ribonucleoproteins; D013539:Simian virus 40 / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection
D034961:Antigens, Nuclear; D000952:Antigens, Polyomavirus Transforming; D004268:DNA-Binding Proteins; D039561:Eukaryotic Initiation Factor-4E; D046911:Macromolecular Substances; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; D010448:Peptide Initiation Factors; C406243:RBM8A protein, human; D039381:RNA Cap-Binding Proteins; D012315:RNA Caps; D012322:RNA Precursors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; C084538:RNPS1 protein, human; D012261:Ribonucleoproteins; C111710:SRRM1 protein, human; D014157:Transcription Factors; D005914:Globins; D012319:RNA Polymerase II