Figure 4

Trop2 is highly conserved among species and mTrop2 expression can stimulate the AP-1 transcription factor. (a) The percentage of sequence identity between the human and murine cytoplasmic tail of Trop2 is 84%. A similar conservation level is observed among other species. "+" indicates a.a. difference. (b) Expression of mTrop2 leads to activation of the AP-1 transcription factor. 293T cells were transiently co-transfected with an AP-1 SEAP reporter construct together with a vector control (pWPXLd) or mTrop2 expressing vector (pWPXLd-mTrop2). As a positive control a pSH-1 SEAP construct was used. Media was collected and processed for SEAP assays and a representative result from three separate experiments done in triplicate is shown as mean ± SD. The negative control was assigned a value of 1, and the other data was normalized to this value. At the time of media collection co-transfected 293T cells show a high level of mTrop2 expression as shown by flow cytometry. (c) Cell lysate from transfected 293T cells used in the AP-1 SEAP assay was harvested and used for immunoblotting to detect the levels of total and phosphorylated ERK1/2. (d) 293T cells were transiently co-transfected with an AP-1 SEAP reporter construct together with pWPXLd or pWPXLd-mTrop2. Cells were subsequently treated with media containing different concentrations of the MEK1 inhibitor PD98059 or DMSO. Media was collected and processed for SEAP assays. Results are shown as mean ± SD.