IDENTIFICATION OF A NEW P-GLYCOPROTEIN-LIKE ATP-BINDING CASSETTE TRANSPORTER GENE THAT IS OVEREXPRESSED DURING HEPATOCARCINOGENESIS

The liver is remarkably insensitive to a variety of cytotoxins and exp resses a number of known drug resistance genes. To isolate new P-glyco protein (Pgp)-related genes, we screened a normal rat liver cDNA libra ry at low stringency with a MDR1 cDNA fragment containing the P-loop a nd ATP binding site We isolated a novel cDNA closely related to the Pg ps that is dramatically increased in hepatic neoplasia and refer to it as P-glycoprotein-related protein (PRP). The predicted protein shows PRP to be a member of the ATP-Binding Cassette (ABC) family of protein s, and a multisequence comparison of the nucleotide binding domain and the ABC family signature sequences reveals that PRP sequences are hig hly conserved with the greatest similarity to the yeast heavy metal tr ansporter encoded by hmt1, However, the hydropathy plot analysis sugge sts that PRP does not have any prominent membrane-spanning domains and thus is not typical of ABC transporters. The PRP transcript is detect ed in many normal tissues. In the H35 hepatoma cell line, PRP was over expressed compared to normal liver, Southern blot analysis of DNA from the H35 rat hepatoma cells reveals that the PRP gene was amplified co mpared to normal liver. The orotic acid model of hepatocarcinogenesis was used to determine if during stepwise progression to liver cancer, PRP changed with hepatocarcinogenesis. At the hyperplastic nodule stag e, PRP expression was increased over its expression in normal surround ing liver. More dramatic increases in PRP expression were found in fra nk hepatic carcinomas, Cumulatively, these studies are the first to li nk a novel ABC family member to the hepatic neoplastic process, a role that may be recapitulated in other cells, considering the ubiquitous expression of PRP.