Figure 5.

ULK1 facilitates PARP1 redistribution and activation. (a) Localization of PARP1 was monitored in Ulk1^+/+ and Ulk1^−/− MEFs transfected with PARP1-GFP with or without exposure to 500 μM H₂O₂ for 1 h. After H₂O₂ exposure, PARP1 was depleted from the nucleoli in ULK1 WT MEFs but not in the Ulk1-KO MEFs. Representative images are shown. Scale bars=10 μM. (b) The percentage of cells showing nucleolar PARP1 was quantified in Ulk1^+/+ and Ulk1^−/− MEFs transfected with PARP1-GFP for 1 h after exposure to UV radiation or treatment with 500 μM H₂O₂ (n>100 cells per condition from three independent experiments). (c and d) Representative immunoblots and graphs (mean±S.D.) show the levels of PARylated proteins in Ulk1^+/+ and Ulk1^−/− MEFs treated with 500 μM H₂O₂ for the indicated time periods. (e and f) Ulk1^+/+ and Ulk1^−/−MEFs, and Ulk1^−/−MEFs stably expressing ULK1 were incubated with or without 500 μM H₂O₂ for 10 min before preparation of extracts for immunoblot detection of PARylated proteins. Representative immunoblots and graphs (mean±S.D.) show the levels of PARylated proteins. Each condition was replicated in triplicate, unless otherwise indicated. *P<0.001