Specific gene amplification associated with consistent chromosomal abnormality in independently established multidrug-resistant Chinese hamster ovary cells

Abstract

Multidrug-resistant (MDR) Chinese hamster ovary (CHO) cell lines were established by selection for resistance to the toxicities of vinblastine (VB) and Adriamycin (AD) in progressively increasing drug concentrations. These cell lines have amplified the DNA sequence that has previously been shown to be amplified in another MDR CHO cell line which was selected with vincristine (VC). An overproduced 4.5 kb mRNA was detected in these MDR cell lines. We report here that the levels of DNA amplification and the 4.5 kb transcript do not correlate with the levels of drug resistance, suggesting that either translational control for the expression of the amplified gene is involved or multiple genes are participating in conferring drug resistance in these cell lines. The amplified DNA sequence was used as a probe and localized by in situ hybridization to chromosome 1q 26-28 (middle portion of the long arm) in the drug-sensitive CHO line, but proximal to the telomere of chromosome 1q in both VB- and AD-selected MDR cell lines. This is consistent with results that have been previously reported for the VC-selected MDR cell lines. Cytogenetic analyses revealed abnormal chromosomal banding patterns or homogeneously staining regions (HSR) between 1q 26-28 and the 1q ter in these independently established MDR lines. These results, taken together, suggest that chromosomal rearrangements leading to gene translocation have consistently accompanied gene amplification in these MDR cell lines. The mechanisms of translocation and its implication in multidrug resistance in these cell lines are discussed.

Chromosome breakage at a major fragile site associated with P-glycoprotein gene amplification in multidrug-resistant CHO cells.

“…These results, consistent with our initial observation (35), indicate that the amplified DNAs in the three independently established MDR CHO cell lines are distally localized on the chromosome where the single-copy sequence resides. These observations are also consistent with the fact that many drug-resistant Chinese hamster variants contain amplified DNA (22,(45)(46)(47).…”

“…Southern blot hybridizations have shown that both P-gp (35,42) and gene class 5 sequences (unpublished data) are coamplified 30-to 40- Indeed, we found that the Z7 marker chromosome (M) gained an additional segment with a size approximating that of the missing part in chromosome lq. Furthermore, the Giemsabanding pattern in the Z7 addition matched that of the distal portion of chromosome lq.…”

“…Furthermore, the Giemsabanding pattern in the Z7 addition matched that of the distal portion of chromosome lq. No loss or gain of other chromosome segments was evident in these two MDR cells (35,42).…”

“…The cell lines VCR15, VBR1.5, and ADR10 were originally established by stepwise selection protocols involving progressively increasing concentrations of vincristine (VC), vinblastine (VB), and adriamycin to final concentrations of 15, 1.5, and 10 ,ug/ml, respectively (35,42 14,1994 on of the long arm) in most of these metaphases ( Fig. 2A and B, arrowheads) (2,35). Three distinct hybridization zones were found in the amplified chromosome segments for both genes by double-color FISH ( Fig.…”

“…(i) The amplified DNA localized distally to the single-copy target gene in the same chromosomal arm has been described for many drug-resistant hamster cells selected with various cytotoxic agents (22, 35, 38-40, 45-47, 51 deletion of the chromosome arm containing amplified DNA has been noted (35). (iii) Frequent sister chromatid fusion and formation of dicentric chromosomes have also been observed to occur in many drug-resistant hamster cell lines (17,22,39,46,51).…”