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Chromosome rearrangements associated with CAD gene amplification. Experiments with cell hybrids
S Viaggi1, M Nüsse, L Ottaggio
1Laboratory of Mutagenesis, IST, Genoa, Italy.
Abstract:
Resistance to phosphonacetyl-L-aspartate (PALA) is caused by CAD gene amplification. The marker chromosome of a PALA-resistant cell line containing a homogeneously staining region with amplified CAD gene was introduced into PALA-sensitive Chinese hamster cells by microcell-mediated chromosome transfer. Two monochromosomal hybrids containing the marker chromosome in addition to the normal chromosome complement of sensitive cells and 1 tetraploid hybrid containing the complete genomes of donor (resistant) and recipient (sensitive) cells were studied in detail. It was shown that (i) the presence of the marker chromosome was both a necessary and a sufficient condition for the expression of the PALA-resistant phenotype; (ii) the marker chromosome underwent rearrangements in the monochromosomal hybrids, with preferential loss of non-amplified chromosomal regions, while it was not rearranged in the tetraploid hybrid; (iii) unlike the original PALA-resistant cells obtained after long-term selection in the presence of PALA, the PALA-resistant hybrids did not show chromosomal aberrations of other than the marker chromosome. This result indicates that chromosomal aberrations may be due to the selective procedure and is not an inherent property of cells containing amplified genes.
Insights
Resistance to phosphonacetyl-L-aspartate (PALA) is linked to CAD gene amplification. Introducing a marker chromosome carrying amplified CAD genes conferred PALA resistance, demonstrating its necessity and sufficiency for the phenotype.
Area of Science:
- Molecular Biology
- Genetics
- Cell Biology
Background:
- Phosphonacetyl-L-aspartate (PALA) resistance in cells is often associated with amplification of the CAD gene.
- Understanding the genetic basis of drug resistance is crucial for developing effective cancer therapies.
Purpose of the Study:
- To investigate the role of a specific marker chromosome containing amplified CAD genes in conferring PALA resistance.
- To analyze the stability and behavior of the marker chromosome in different cellular contexts.
Main Methods:
- Microcell-mediated chromosome transfer was used to introduce a marker chromosome from PALA-resistant cells into PALA-sensitive Chinese hamster cells.
- Characterization of monochromosomal and tetraploid hybrids through detailed cytogenetic analysis.
Main Results:
- The presence of the marker chromosome was both necessary and sufficient for PALA resistance.
- The marker chromosome underwent rearrangements in monochromosomal hybrids, with loss of non-amplified regions, but remained stable in tetraploid hybrids.
- PALA-resistant hybrids lacked the chromosomal aberrations observed in the original resistant cell line, suggesting these aberrations are artifacts of the selection process.
Conclusions:
- CAD gene amplification on a specific marker chromosome is the primary driver of PALA resistance.
- The stability of amplified gene regions may depend on the cellular context (e.g., ploidy).
- Chromosomal aberrations in drug-resistant cells may arise from the selection process rather than being an inherent property of gene amplification.