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. Author manuscript; available in PMC: 2010 Jan 1.
Published in final edited form as: Free Radic Biol Med. 2008 Oct 7;46(1):33–41. doi: 10.1016/j.freeradbiomed.2008.09.022

Figure 1. P21 is detected in the nucleus and cytoplasm during persistent expression.

Figure 1

H1299 cells with inducible expression of EGFp21 were cultured in the presence of doxycycline for 24 hours (day 0) then incubated in room air (RA) or hyperoxia (O2) for 2 days. (A) Endogenous fluorescence of the EGFp21 transgene during culture in room air or hyperoxia. (B) Nuclear (N) and cytoplasmic (C) H1299 cell lysates after 0 and 2 days culture after doxycycline treatment to induce EGFp21 expression immunoblotted for EGFp21, histone H3 and GAPDH. The NLS of EGFp21 (RKR140–142) was mutated (AAA140–142) to generate EGFp21ΔNLS, which was stably transfected into parental H1299 cells. H1299 cell with inducible expression of either EGFp21 or EGFp21ΔNLS were cultured in the presence of doxycycline for 24 hrs and (C) endogenous fluorescence was monitored and (D) protein was fractionated into nuclear or cytoplasmic fractions and immunoblotted for the p21 transgene with Histone H3 and GAPDH used as fractionation controls. Immunoblots shown are representative of at least three separate experiments with similar results.