Gene amplification and its consequences

Abstract

The discovery in the late 1970s of gene amplification in somatic cells has led to the realization that changes in gene copy number may occur both spontaneously and at high frequencies, and that the genomic variability that is thereby generated can provide the basis for a heritable advantage when cells or organisms are exposed to selection pressures (Schimke et al. 1986). Many types of genes, including the cellular protooncogenes, can participate in amplification events, and these events are generally associated with an overproduction of the corresponding gene product that then causes a change in cellular properties. Much of the current work in this field is centered on efforts to demonstrate the occurrence of amplification with diverse genes and organisms, including humans, as well as to establish conditions that may modulate frequencies of amplification. Recent work has also shown that changes in gene copy number may alter regulatory mechanisms within cells, resulting in abnormal patterns of gene expression. This report summarizes the results of such investigations, which were presented during the workshop entitled DNA Amplification at the 16th International Congress of Genetics in Toronto, Canada. The first two presentations described amplification events that occur in Saccharomyces cerevisiae. Dr. C. E. Paquin described her work with the alcohol dehydrogenase genes (ADHl-4) in yeast. She has shown that yeast strains lacking the ADHl gene (which encodes the fermentative isozyme of this family of proteins) are sensitive to antimycin A, an inhibitor of respiration. When exposed to antimycin A, the ADHl deletion mutants give rise to variants that are resistant to the antibiotic, of which approximately 1 % show changes in copy number of either ADH2 or ADH4. The ADH2 gene is normally repressed by the exposure of yeast to glucose. In the antimycin A resistant clone, however, an extra copy of the ADH2 gene has been generated with an altered promoter that permits expression in the presence of glucose, thereby allowing the yeast to survive by fermentation when exposed to both glucose and antibiotic. In an independently derived antimycin A resistant clone, the ADH4 gene was instead amplified to a high level. In this case, the additional genes were carried as extrachromosomal 2 1 -kb sequences, arranged as palindromes on a linear 42-kb DNA sequence. The ADH4 gene diverges significantly from the other members of the ADH family, and presumably encodes a protein that may have only a weak alcohol dehydrogenase activity; this divergence may explain why multiple copies of the gene were required to provide metabolic protection from antimycin A selection. The 42-kb linear DNA fragment is unusually small for a yeast chromosomal element. Although presumably this microchromosome contains telomeres at its ends, it apparently lacks centromeric sequences and very likely segregates randomly at mitosis. Paquin also described a novel selection scheme, employing coamplification of neighboring CUPl (conferring resistance to copper ions) and ADH genes, that will permit the easier detection of new antibiotic resistant mutants that have elevated ADH gene copy numbers. Such mutants should also increase the copy number of the adjacent CUPl gene, and would therefore show elevated resistance to copper, a phenotype that is more readily