Separable roles for rent1/hUpf1 in altered splicing and decay of nonsense transcripts.
The mechanism by which disruption of reading frame can influence pre-messenger RNA (pre-mRNA) processing is poorly understood. We assessed the role of factors essential for nonsense-mediated mRNA decay (NMD) in nonsense-mediated altered splicing (NAS) with the use of RNA interference (RNAi) in mammalian cells. Inhibition of rent1/hUpf1 expression abrogated both NMD and NAS of nonsense T cell receptor beta transcripts. In contrast, inhibition of rent2/hUpf2 expression did not disrupt NAS despite achieving comparable stabilization of nonsense transcripts. We also demonstrate that NAS and NMD are genetically separable functions of rent1/hUpf1. Additionally, rent1/hUpf1 enters the nucleus where it may directly influence early events in mRNA biogenesis. This provides compelling evidence that NAS relies on a component of the nonsense surveillance machinery but is not an indirect consequence of NMD.
Mendell
Joshua T
JT
Institute of Genetic Medicine and Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, 858 Ross Building, 720 Rutland Avenue, Baltimore, MD 21205, USA.
ap Rhys
Colette M J
CM
Dietz
Harry C
HC
D021581:Active Transport, Cell Nucleus; D017398:Alternative Splicing; D019943:Amino Acid Substitution; D015152:Blotting, Northern; D002467:Cell Nucleus / Q000378:metabolism; D018389:Codon, Nonsense; D003593:Cytoplasm / Q000378:metabolism; D033722:Equilibrative-Nucleoside Transporter 2 / Q000235:genetics / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D020868:Gene Silencing; D019674:Genes, T-Cell Receptor beta; D006367:HeLa Cells; D006801:Humans; D009154:Mutation; D020365:RNA Helicases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D018389:Codon, Nonsense; D033722:Equilibrative-Nucleoside Transporter 2; D005231:Fatty Acids, Unsaturated; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; C438655:Slc29a2 protein, rat; D015534:Trans-Activators; D014157:Transcription Factors; C468286:UPF2 protein, human; D020365:RNA Helicases; C101828:UPF1 protein, human; C038753:leptomycin B