FIG 2.

LMP1 promotes cell proliferation and focus formation via IGF1R. (A) LMP1-expressing 293T cells were incubated with the indicated micromolar concentrations of AG1024 (AG) or PPP (P) for 1 day. Cell lysates were immunoprecipitated (IP) with anti-pTyr and probed with anti-IGF1R to specifically detect phospho-IGF1R. Numerical values indicate fold reduction in pIGF1R compared to that in a dimethyl sulfoxide (DMSO) vehicle control. In parallel, total cell lysates (TCLs) were probed for IGF1R, LMP1, and GAPDH. (B to D) MTS assays were used to assess the proliferation of LMP1-positive cells relative to pBabe cell proliferation over a 2-day period (B). In the same time span, cells were exposed to DMSO vehicle control, AG1024 (C), or PPP (D), and proliferation was measured. (E to H) MCF10a cells were sparsely seeded and given the indicated treatment for 10 days before foci were stained and quantitated (E). The focus number and size of LMP1-expressing cells were calculated relative to those for pBabe vector controls (F). Four hours after seeding, cells were treated with AG1024 or PPP, and the relative number (G) and size (H) of foci were subsequently assessed. In panels C, D, G, and H, the fraction of treated cell growth or focus formation was calculated relative to that for untreated cells. The proportion of LMP1-positive cells was then calculated relative to that for pBabe vector control cells. All drug doses are micromolar. *, LMP1-positive cells are significantly different than pBabe cells (P < 0.05).