Figure 4.

Transcriptional activation by CD44. (A) COS-7 cells were transfected with plasmids as indicated, and the transcriptional activity from the cotransfected reporter plasmids was measured. The activity of the TRE reporter plasmids was evaluated by transfecting COS-7 cells with RasV12 as a positive control. CD44ICD revealed ∼50% of the TRE-mediated transcriptional activity induced by overexpressed RasV12 protein under the experimental condition (insets). (B) Nucleotides mutated in Mut-TRE reporter plasmids are shown in italics and underlined. (C) COS-7 cells were transfected with a cDNA encoding full-length CD44 (lanes 2 and 3), CD44Δ287–290 constructs (lane 4) or with empty vector (lane 1). 36 h after transfection, the cells were incubated for an additional 12 h in the absence (lanes 1, 2, and 4) or presence of 100 μM BB2516 (lane 3). Blots were probed with anti-CD44cyto Ab. Asterisk indicates endogenous full-length CD44. (D) COS-7 cells were cotransfected with the TRE reporter plasmid together with an expression vector encoding full-length CD44, oncogenic Ras, or the corresponding empty vector. After 32 h, the cells were incubated for an additional 16 h in serum-free medium containing BB2516 (100 μM) or DMSO as a vehicle control before measurement of luciferase activity. Luciferase activity was expressed as fold induction relative to the value for cells transfected with the empty vector. (E) COS-7 cells were cotransfected with TRE reporter plasmid together with full-length CD44, CD44Δ287–290, or the corresponding empty vector. Data are expressed as indicated in Fig. 3 D.