FIG. 9.

Proliferation of pLIG.Notch1IC.H cell lines after long-term estrogen withdrawal. (A) pLIG.Notch1IC.H-expressing cells were allowed to continue growing after 6 days without estrogen (as for Fig. 7A). After that, the cells were resuspended in fresh estrogen-free medium at 3 × 10⁴ per well and cell proliferation was monitored daily for an additional 6 days by MTT assay. (B) pLIG.hNotch1IC and pLIG.EBNA2 cells were allowed to grow without estrogen for 4 weeks. The resulting outgrowing rescued cells lines, Notch1IC.R and EBNA2.R, were plated at 2 × 10⁴ cells per well in the estrogen-free medium, and cell proliferation was monitored daily for 4 days as described for panel A. (C) Western blots of cellular extracts derived from cell lines grown continously without estrogen. Lane 1, pLIG.Notch1IC-rescued cells (Notch1IC.R in panel B); lane 2, pLIG.EBNA2-rescued cells (EBNA2.R in panel B); lane 3, EREB2.5 cells grown with estrogen. The upper panel shows a blot probed with a c-Myc antibody, and the lower panel shows a blot probed with an LMP-1 antibody. c-Myc and LMP-1 proteins are indicated by the arrows.