Regulation of Crm1 expression in cancer cells

Crm1 (or exportin 1) is a member of the karyopherin β protein family that mediates the nuclear export of cargo proteins and certain RNAs from the nucleus into the cytoplasm. We previously identified increased expression of Crm1 mRNA and protein in cervical tumours compared to normal epithelium, and increased Crm1 expression in transformed fibroblasts compared to normal fibroblasts. Moreover, Crm1 overexpression in cancer/transformed cells was found to be necessary for cell survival, implicating Crm1 as an attractive target for cancer therapy. We hypothesized that the increased expression of Crm1 in cancer/transformed cells derives from altered Crm1 promoter activity, yet no studies to date have investigated what regulates the Crm1 promoter. This study therefore set out to clone and characterize the Crm1 promoter. The cloned Crm1 promoter (-1405 to +99) was significantly more active in cancer and transformed cells compared to normal, correlating well with endogenous Crm1 protein levels. In order to define promoter regions responsible for the differential activity, deletion constructs were generated, and the −175 to +99 region was identified as important. Mutation of two CCAAT boxes and a GC box in this region significantly diminished Crm1 promoter activity in transformed and cancer cells, while having only a minor effect on Crm1 promoter activity in normal cells. In addition, ChIP assays revealed in vivo binding of NFY/CBP and Sp1 transcription factors to the CCAAT boxes and GC box of the Crm1 promoter in cancer and transformed cells, while binding was undetectable in normal cells. Interestingly, NFY and Sp1 expression levels were considerably elevated in transformed and cancer cells compared to normal. Taken together, we show that NFY- and Sp1-mediated activation of the Crm1 promoter contributes to Crm1 overexpression in cancer and transformed cells.