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Specific gene amplification associated with consistent chromosomal abnormality in independently established
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.
Insights
Multidrug-resistant Chinese hamster ovary (CHO) cell lines show amplified DNA and mRNA. However, gene amplification levels don't correlate with drug resistance, suggesting complex gene expression regulation or multiple genes involved.
Area of Science:
- Molecular Biology
- Genetics
- Cell Biology
Background:
- Multidrug resistance (MDR) in cancer therapy is a significant challenge.
- Chinese hamster ovary (CHO) cell lines are widely used models for studying drug resistance mechanisms.
Purpose of the Study:
- To investigate the genetic basis of multidrug resistance in CHO cell lines selected for resistance to vinblastine (VB) and Adriamycin (AD).
- To explore the correlation between gene amplification, mRNA expression, and drug resistance levels.
- To identify chromosomal alterations associated with multidrug resistance.
Main Methods:
- Establishment of MDR CHO cell lines through stepwise drug selection.
- Detection of DNA amplification and mRNA levels using molecular probes.
- In situ hybridization to localize amplified DNA sequences on chromosomes.
- Cytogenetic analysis to identify chromosomal abnormalities.
Main Results:
- Established MDR CHO cell lines exhibited amplification of a specific DNA sequence and overproduction of a 4.5 kb mRNA.
- The levels of DNA amplification and 4.5 kb transcript did not correlate with the degree of drug resistance.
- In situ hybridization localized the amplified DNA sequence to chromosome 1q, with a shift in location in MDR lines.
- Cytogenetic analysis revealed homogeneously staining regions (HSR) and chromosomal rearrangements in MDR lines.
Conclusions:
- Gene amplification and mRNA overexpression alone do not fully explain the observed multidrug resistance.
- Translational control or the involvement of multiple genes may contribute to MDR.
- Chromosomal rearrangements, including translocations, consistently accompany gene amplification in these MDR CHO cell lines.
- These findings highlight the complex genetic and chromosomal alterations underlying multidrug resistance.