FIGURE 4:

The Ran gradient and Ubc9 import are sensitive to oxidative stress. (A) Endogenous Ubc9 distribution detected by IF in HeLa cells treated for 10 min with the indicated concentrations of H₂O₂. (B) Endogenous Ubc9 (red) and Ran (green) in HeLa cells undergo reversible changes in distribution in response to oxidative stress. HeLa cells were treated for 10 min with 200 μM H₂O₂ and then allowed to recover in fresh media for 30 and 60 min. (C) xy scatter plot of Ubc9 N/C and Ran N/C in untreated (black dots) and H₂O₂-treated HeLa cells (red dots). (D, E) Ubc9 and Ran N/C levels in HeLa cells treated with 200 μM H₂O₂ measured as a function of time. (F) Ubc9-Uba2 heterodimer detected by immunoblotting. HeLa cells were treated for 10 min with the indicated concentrations of H₂O₂ and immunoblotted for Ubc9. Gel samples were prepared without or with 100 mM dithiothreitol, as indicated. Scale bars, 20 μm.