Inhibition of gene amplification in telomerase deficient immortalized mouse embryonic fibroblasts

Mutations in genes important for the preservation of genome stability can increase the frequency of gene amplification, a process relevant to tumor development. To investigate whether telomerase, the enzyme deputed to telomere maintenance, also plays a role in gene amplification, we studied the amplification of the carbamyl-P-synthetase, aspartate transcarbamilase, dihydro-orotase (CAD) gene in immortalized embryonic fibroblasts derived from telomerase knockout mice (mTERC(-/-)) of the first and of the sixth generation. As expected, in 9 out of 10 N-(phosphonacetyl)-L-aspartate (PALA) resistant clones derived from wild-type cells, CAD was amplified; in contrast, in none of the 30 PALA resistant clones isolated from the three mTERC(-/-) cell lines we could detect CAD amplification, indicating that, in the absence of telomerase activity, gene amplification is inhibited. The causal relationship between mTERC deficiency and lack of gene amplification was demonstrated by the restoration of CAD gene amplification in two of the three deficient cell lines transfected with mTERC. The lack of amplification in mTERC deficient cells could be related to a defect in the stabilization of the ends of the amplified chromosomes in the absence of telomerase, to a more general effect of telomerase in the regulation of gene expression, including genes involved in amplification, or to a possible interaction of the telomerase RNA with proteins involved in gene amplification.