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. Author manuscript; available in PMC: 2013 Feb 26.
Published in final edited form as: Science. 2009 Sep 17;326(5953):722–726. doi: 10.1126/science.1176326

Fig. 1.

Fig. 1

IRP2 is stabilized under iron-replete conditions after siRNA-mediated suppression of SCFFBXL5. (A) IRP2 accumulation was assessed using the AlphaScreen assay (top) or by immunoblot analysis of endogenous IRP2 (bottom). Knockdown efficiency of the SKP1, CUL1, and RBX1 siRNAs is shown in fig. S2D. (B) IRP1 stabilization under iron-replete conditions after siRNA-mediated suppression of FBXL5, as measured by immunoblot analysis of HEK 293 cell lines stably transfected with either a myc-tagged wild-type (WT) or 3C>3S IRP1 Tet-inducible expression construct. (C) Measurement of RNA binding activity from lysates after siRNA-mediated suppression of FBXL5 in cells expressing HA-IRP2-FLAG. Because human IRP1-IRE and IRP2-IRE complexes migrate similarly, antibodies were added to supershift individual complexes. (D) Relative TfR1 and FBXL5 mRNA accumulation levels measured by qRT-PCR. Assays were performed in triplicate with data represented as the mean ± SE with P values determined using Student's unpaired t test (***P < 0.001).