Overexpression of p-glycoprotein results in the multidrug resistant (mdr) phenotype in which cells become cross-resistant to a number of structurally and functionally unrelated drugs. The physiological function of this protein is yet unknown. Due to its location on the bile canalicular surface of hepatocytes, we investigated the possible regulation of mdr expression by manipulations of biliary flow. Induction of cholestasis by surgical ligation of the bile duct or chemical treatment with `-naphthylisothiocyanate (ANIT) caused an increase in mdr expression in both rodents and primates. The increase in mdr mRNA was a result of increased gene transcription. We extended our previous investigations on the regulation of mdr expression by xenobiotics to include extrahepatic organs in the rat and to the liver of primates. We observed increased expression in the rat kidney, small intestine and lung by methylcholanthrene and/or 2-acetylaminofluorene (2-AAF). Enhanced expression of mdr mRNA was observed in monkey livers after administration of 2-AAF; this increase was demonstrated to be the result of increased transcription. To further investigate the regulation of the mdr1b gene, we cloned and sequenced the 5' upstream region of this gene and are conducting functional promotor assays. This region has a high degree of sequence identity to the mouse mdr1b promotor region and contains several previously identified cis-acting transcriptional elements including CAAT, TATA AP-1 and SP-1. A 3.2 kb cDNA clone of a second member of the rat mdr gene family, mdr2, was also isolated and sequenced; it has over 90% identity to the mouse mdr2 gene. The remainder of this gene, approximately 800 bp, is being cloned. We have continued work on possible contributions of message stability to the level of steady state mdr mRNA. Analysis of the 3' end of the human MDR1 gene has indicated that the entire 3' UTR region may be important in the determination of the half life of the mdr mRNA and that this effect may be due to secondary structure requirements.
