The promoter of rat multidrug resistance protein 3 (Mrp3) has been cloned and analyzed in the rat intestinal cell line (IEC-18 cells). A series of 5' deletion mutants of the Mrp3 promoter region were constructed and placed into the pGL3-Basic vector (luciferase reporter gene). Deletion analysis of the Mrp3 promoter identified a basal transcription element at -123/-106, two negative response regions at -2723/ -1128 and -530/-443, respectively, and two positive response regions at -1063/-943 and -302/-157. Further site-directed mutagenesis analysis and gel mobility shift assays provided evidence for Sp1 and Sp3 binding within -123/-105 regions. These studies indicate that Sp1 and Sp3 may be involved in the regulation of the rat Mrp3 gene. (C) 2002 Elsevier Science (USA).