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Series GSE60045 Query DataSets for GSE60045
Status Public on Aug 15, 2014
Title A post-translational regulatory switch on UPF1 controls targeted mRNA degradation.
Organism Homo sapiens
Experiment type Expression profiling by high throughput sequencing
Summary Nonsense-mediated mRNA decay (NMD) controls the quality of eukaryotic gene expression and also degrades physiologic mRNAs. How NMD targets are identified is incompletely understood. A central NMD factor is the ATP-dependent RNA helicase UPF1. Neither the distance in space between the termination codon and the poly(A) tail nor the binding of steady-state, largely hypophosphorylated UPF1 is a discriminating marker of cellular NMD targets, unlike for PTC-containing reporter mRNAs when compared to their PTC-free counterparts. Here, we map phosphorylated UPF1 (p-UPF1) binding sites using transcriptome-wide footprinting or DNA oligonucleotide-directed mRNA cleavage to report that p-UPF1 provides the first reliable cellular NMD-target marker. p-UPF1 is enriched on NMD target 3'UTRs along with SMG5 and SMG7 but not SMG1 or SMG6. Immunoprecipitations of UPF1 variants deficient in various aspects of the NMD process in parallel with FRET experiments reveal that ATPase/helicase-deficient UPF1 manifests high levels of RNA binding and disregulated hyperphosphorylation, whereas wild-type UPF1 releases from nonspecific RNA interactions in an ATP hydrolysis-dependent mechanism until an NMD target is identified. 3'UTR-associated UPF1 undergoes regulated phosphorylation on NMD targets, providing a binding platform for mRNA degradative activities. p-UPF1 binding to NMD target 3'UTRs is stabilized by SMG5 and SMG7. Our results help to explain why steady-state UPF1 binding is not a marker for cellular NMD substrates and how this binding is transformed to induce mRNA decay.
Overall design RIP-seq experiments for p-UPF1, control IPs using rabbit IgG and additional control sample without IP were performed in duplicates
Contributor(s) Kurosaki T, Li W, Hoque M, Popp MW, Ermolenko DN, Tian B, Maquat LE
Citation(s) 25184677
Submission date Aug 04, 2014
Last update date May 15, 2019
Contact name Wencheng Li
Organization name PTC Therapeutics
Street address 100 Corporate Count
City South Plainfield
State/province NJ - New Jersey
ZIP/Postal code 07080
Country USA
Platforms (1)
GPL16791 Illumina HiSeq 2500 (Homo sapiens)
Samples (6)
GSM1464145 Ctl_1
GSM1464146 Ctl_2
GSM1464147 IgG_1
BioProject PRJNA257379
SRA SRP045217

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Supplementary file Size Download File type/resource
GSE60045_RAW.tar 3.5 Mb (http)(custom) TAR (of BED)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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