Differential changes in genome structure and expression of the mdr gene family in multidrug-resistant murine erythroleukemia cell lines.
The genome structure and expression of mdr genes were examined in multidrug-resistant sublines of two different murine (DBA/2J) Friend erythroleukemia cell lines, PC4 and C7D, derived by stepwise exposure to increasing concentrations of adriamycin beginning with 5 ng/ml. The PC4 cell lines selected in higher drug concentrations (80-1280 ng/ml) demonstrated amplification of all three mdr genes with preferential amplification of mdr3. Overexpression of the mdr2 and mdr3 genes accompanied their genomic amplification; however, expression of mdr1 was not seen despite amplification. In the C7D cell lines selected with higher drug concentrations (40-160 ng/ ml), amplification and overexpression of mdr1 and mdr2 without mdr3 was observed. Increased expression of mdr1 occurred prior to gene amplification. The distribution of mdr-specific genes in micrococcal nuclease-generated chromatin fractions differing in transcriptionally active sequences and proteins was different between the parent and drug-resistant sublines. An enrichment (two- to threefold) of mdr3 genes in the H1-depleted mononucleosome fraction enriched for actively transcribed genes (e.g., globin) was detected by Southern analysis of chromatin fractions in PC4-80 cells (selected in 80 ng/ml of adriamycin and overexpressing mdr3), compared to the parental cells. mdr3 enrichment was also detected using a new PCR-based method, which examined mdr3 genes and repetitive sequences. Of note, the H1-depleted chromatin fraction from PC4-20 showed enrichment of the mdr3 gene, although mdr3 expression was not detected in the cell line. These studies showed a different pattern of gene amplification and overexpression in genetically related erythroleukemia cell lines selected for resistance to the same chemotherapeutic agent. A change in chromatin organization of mdr genes preceded overexpression and amplification of the mdr3 gene.
Fracasso
P M
PM
Department of Medicine, New England Medical Center, Boston, MA, USA.
Slapak
C A
CA
Nair
S
S
Modrak
D
D
Draper
M P
MP
LeCerf
J M
JM
Levy
S B
SB
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D002843:Chromatin / Q000648:ultrastructure; D004273:DNA, Neoplasm / Q000235:genetics; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D005784:Gene Amplification; D019450:Genes, MDR; D004915:Leukemia, Erythroblastic, Acute / Q000473:pathology; D051379:Mice
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D002843:Chromatin; D004273:DNA, Neoplasm