Figure 2.

Induction of CLU/XIP8 after IR. Log-phase MCF-7 cells were treated with various doses of IR. (A) Increased steady-state expression of clu/xip8 transcripts in MCF-7 cells, 3 days after IR. Equivalent total RNA loading was monitored by ethidium bromide staining and 36B4 transcript levels as described (14). (B) Time-course induction of CLU/XIP8 proteins in MCF-7 cells after IR. Log-phase MCF-7 cells were treated with 10 Gy and whole-cell lysates were prepared. CLU/XIP8 proteins (60- and 40-kDa forms) were monitored by Western blot assays and detected by using the 41D antibody. Ku70 protein was detected by the N3H10 antibody and served as a loading control. (C) Increased steady-state levels of the 60-kDa precursor and ≈40-kDa secretory forms of CLU/XIP8 after various doses of IR. Whole-cell lysates from MCF-7 cells were prepared 3 days after IR. Ku70 levels remained unaltered. (D) Dose-response of CLU/XIP8 protein induction in MCF-7 cells after IR. Data in Fig. 3B, as well as other experiments using lower IR doses were compiled. Induction of nuclear CLU/XIP8 protein was not apparent until 1 Gy (not shown). Western blots of CLU/XIP8 compared with Ku70 or α-tubulin protein levels in control or IR-treated MCF-7 cells were quantified by densitometric scans and normalized for loading. Values for treated/control protein levels for each dose of IR then were graphed (mean ± SE). Data represent the results of experiments performed at least three times, each in duplicate.