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. 2016 Nov 17;7:13498. doi: 10.1038/ncomms13498

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Copyright © 2016, The Author(s)

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(a) The boxed sequence from the 3′-UTR of the IFN-γ mRNA indicates a similar GRE sequence. Sequences of IFN-γ-GRE-Bio and mutated IFN-γ-GRE-Bio were used for binding reactions. (b) mRNA pull down assay was performed by mixing IFN-γ-GRE-Bio or mIFN-γ-GRE-Bio with total cell extracts from LX-2 cells. Precipitates were prepared for Western blotting using SoftLink Soft Release avidin resin. (c,d) Cold IFN-γ-GRE probes or mIFN-γ-GRE probes with different concentrations were used to compete for the binding between IFN-γ-GRE-Bio and CUGBP1. (e) The experiment shown in d was performed thrice, and the binding bands were quantified using a phosphorimager. (f) LX-2 cells were transfected with si-CUGBP1 or control siRNA for 48 h and treated with actinomycin D for the indicated times. Cells were collected for quantitative PCR analyses.