Amplification and overexpression of the mouse mdr 1a gene in nine independently derived multidrug-resistant SEWA murine cell lines.

Abstract

Many different drugs may be used in selecting cells for multidrug resistance (MDR). Enhanced expression and/or gene amplification is known to cause overproduction of membrane-bound 170,000 P-glycoproteins, responsible for the MDR. In rodents, the P-glycoproteins are encoded by a small gene family: mdr 1a, mdr 1b, and mdr 2. To evaluate the relationship between the pattern of MDR and the selecting drug, nine MDR sublines were independently selected from a sensitive mouse tumor cell line, SEWATC13K, using three different drugs. Each MDR subline displayed amplification of one or more of the three mdr genes, but only one, mdr 1a, was consistently overexpressed. Thus, our results indicate that the pattern of mdr gene amplification and overexpression is independent of the selective agent. Furthermore, in four of the MDR sublines, where all three mdr genes had been originally amplified, pulsed field gel electrophoresis (PFGE) revealed that amplification of mdr 1a, only, was a second step of gene amplification. In addition, the gene for the calcium-binding protein, sorcin, was coamplified in eight of the nine MDR sublines. The sorcin gene was overexpressed in seven of these eight sublines. Finally, hybridizations with a probe homologous with a putative region of RFLP (restriction fragment length polymorphism), indicated that the amplified sequences originate from one or the other of the two homologous chromosomes with no preference.