| Multidrug resistance (MDR) is a barrier to successful cancer chemotherapy. Although MDR is highly associated with overexpression of ABC transporters, mechanisms behind their upregulation are not entirely understood. N1 TM* is a gamma-secretase-cleavable truncated transmembrane Notch1 protein and its cleaved form, intracellular Notch1 (N1 IC), is involved in transcriptional regulation involved in cellular development, differentiation, and apoptosis. Our early observation revealed an inverse relationship between the expression of ABCC1/MRP1 and N1 TM* in leukemic MDR cells. Therefore, we hypothesized that there are distinct Notch1 regulatory mechanisms controlling the overexpression of ABC transporters in MDR cancer cells.;To test whether Notch1 is involved in the overexpression of ABC transporters, we measured N1TM* in the MDR cells overexpressing representative ABC transporters, ABCB1/MDR1, ABCC1/MRP1, and ABCG2/BCRP. We confirmed that the inverse relationship between the expression of ABC transporters and N1 TM* is confined in MDR cells overexpressing ABCC1/MRP1 but not in cells overexpressing other ABC transporters. Furthermore, the measurement of PSEN1, a catalytic component of GS that is involved in generation of N IC from NTM*, showed upregulation of its level in the MDR cells overexpressing ABCC1/MRP1. Indeed, when we reduced N1IC levels in MCF7NP cells with either a gamma-secretase inhibitor or shRNA. We observed a reduction of ABCC1/MRP1. By contrast, ectopic expression of N1IC in MCF7/WT cells led to increased expression of ABCC1/MRP1 and drug resistance. With an ABCC1/MRP1 promoter construct, we observed its reduced transcriptional activity after blocking the generation of N1 IC; we also observed its induced transcriptional activity in stable cells ectopically overexpressing N1IC. Moreover, electrophoretic mobility shift (EMSA) and chromatin immunoprecipitation (ChIP) assays revealed an interaction between a specific promoter region of ABCC1/MRP1 and the N1 IC-activated transcription factor CBF1, suggesting that the regulation of ABCC1/MRP1 expression by Notch1 is mediated by CBF1. Finally, we examined the C-class of ABC transporters, and found that the promoter of ABCC4 and ABCC5 also contain a putative CBF1 binding element, and their expression was increased in cells with higher N1IC levels, but we did not see this for ABCC2, ABCC3, and ABCC6.;Overall, our results reveal a novel regulatory mechanism of ABCC1/MRP1 expression. |
