Notch1 Regulates the Expression of the Multidrug Resistance Gene ABCC1/MRP1 in Cultured Cancer Cells

Multidrug resistance (MDR) is a barrier to successful cancer chemotherapy. Although MDR is associated with overexpression of ATP-binding cassette (ABC) membrane transporters, mechanisms behind their up-regulation are not entirely understood. The cleaved form of the Notch1 protein, intracellular Notch1 (N1(IC)), is involved in transcriptional regulation of genes. To test whether Notch1 is involved in the expression of multidrug resistance-associated protein 1 (ABCC1/MRP1; herein referred to as ABCC1), we measured N1(IC) and presenilin 1 (PSEN1), the catalytic subunit of γ-secretase required for Notch activation. We observed higher levels of N1(IC) and PSEN1 proteins as well as higher activity of N1(IC) in ABCC1-expressing MDR MCF7/VP cells compared with parental MCF7/WT cells. Reducing N1(IC) levels in MCF7/VP cells with either a γ-secretase inhibitor or shRNA led to reduction of ABCC1. By contrast, ectopic expression of N1(IC) in MCF7/WT cells led to increased expression of ABCC1 and associated drug resistance, consistent with expression of this transporter. Inhibition of ABCC1 reversed drug resistance of N1(IC)-overexpressing stable cells. Using an ABCC1 promoter construct, we observed both its reduced transcriptional activity after blocking the generation of N1(IC) and its increased transcriptional activity in stable cells overexpressing N1(IC). ChIP and gel-shift assays revealed an interaction between a specific promoter region of ABCC1 and the N1(IC)-activated transcription factor CBF1, suggesting that the regulation of ABCC1 expression by Notch1 is mediated by CBF1. Indeed, deletion or site-directed mutagenesis of these CBF1 binding sites within the ABCC1 promoter region attenuated promoter-reporter activity. Overall, our results reveal a unique regulatory mechanism of ABCC1 expression.