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Preferential expression of a mutant allele of the amplified MDR 1 ( ABCB 1) gene in drug‐resistant variants of a human sarcoma
Author(s) -
Chen Kevin G.,
Lacayo Norman J.,
Durán George E.,
Wang Yan,
Bangs C. Dana,
Rea Susan,
Kovacs Mary,
Cherry Athena M.,
Brown J. Martin,
Sikic Branimir I.
Publication year - 2002
Publication title -
genes, chromosomes and cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.754
H-Index - 119
eISSN - 1098-2264
pISSN - 1045-2257
DOI - 10.1002/gcc.10067
Subject(s) - biology , allele , mutant , microbiology and biotechnology , southern blot , locus (genetics) , gene , genetics , amplicon , polymerase chain reaction
Activation of the MDR 1 ( ABCB 1) gene is a common event conferring multidrug resistance (MDR) in human cancers. We investigated MDR 1 activation in MDR variants of a human sarcoma line, some of which express a mutant MDR 1, which facilitated the study of allelic gene expression. Structural alterations of MDR 1, gene copy numbers, and allelic expression were analyzed by cytogenetic karyotyping, oligonucleotide hybridization, Southern blotting, polymerase chain reaction, and DNA heteroduplex assays. Both chromosome 7 alterations and several cytogenetic changes involving the 7q21 locus are associated with the development of MDR in these sarcoma cells. Multistep‐selected cells and their revertants contain three‐ to six‐fold MDR 1 gene amplification compared with that of the drug‐sensitive parental cell line MES‐SA and single‐step doxorubicin‐selected mutants. MDR 1 gene amplification precedes the emergence of a mutant allele in cells that were coselected with doxorubicin and a cyclosporin inhibitor of P‐glycoprotein (P‐gp). Allele‐specific oligonucleotide hybridization showed that the endogenous mutant allele was present as a single copy, with multiple copies of the normal allele. Reselection of revertant cells with doxorubicin in either the presence or the absence of the P‐gp inhibitor resulted in exclusive reexpression of the mutant MDR 1 allele, regardless of the presence of multiple wild‐type MDR 1 alleles. These data provide new insights into how multiple alleles are regulated in the amplicon of drug‐resistant cancer cells and indicate that increased expression of an amplified gene can result from selective transcription of a single mutant allele of the gene. © Wiley‐Liss, Inc.