Fig. 6. KLF4 and SP1 interact with IPO13 in the presence and absence of H₂O₂-induced oxidative stress.

HeLa cells transfected to co-express GFP or GFP-KLF4 (a–c) or -SP1 (d–f) with DSRED-IPO13. At 16 h post-transfection, cells were treated with or without 125 μM H₂O₂ for 1 h before lysis and incubation with or without GTPγS and immunoprecipitation (IP) using GFP-Trap (Chromotek). Input/IP samples were probed by Western blotting using mouse-anti-GFP (Roche) or rabbit anti-IMP13 (Protein Tech) antibodies and then imaged using the ChemiDoc Gel Imaging System (Biorad). b, c, e, f Densitometric analysis was performed on images such as those in a and d for binding of DSRED-IPO13 to GFP-KLF4 (a) or GFP-SP1 (d) under H₂O₂ treated conditions and untreated (UT) conditions with and without the addition of GTPγS. The amount of co-immunoprecipitated IPO13 was normalized to the amount of protein available for co-immunoprecipitation (as seen in the input panels). This value was normalized to the amount of immunoprecipitated GFP-KLF4 or –SP1. Pooled results representing the mean ± SEM (error bars) for IPO13 bound under H₂O₂ treated conditions relative to UT conditions (no GTPγS treatment - No Add.) (b n = 2 independent experiments, e n = 4 independent experiments) or with GTPγS treatment under UT and treated conditions (c n = 2 independent experiments, f n = 4 independent experiments) are shown. p values (two-tailed student’s t test) left to right: p = 0.0422 and p = 0.0375. Source data are provided as a Source Data file.