Amplification in rodent cells usually involves bridge-breakage-fusion (bbf) cycles initiated either by endto-end fusion of sister chromatids, or by chromosome breakage. In contrast, in human cells, resistance to the antimetabolite A-(phosphonacetyl)-L-aspartate (PALA) can be mediated by several different mechanisms that lead to overexpression of the target enzyme carbamyl-P synthetase, aspartate transcarbamylase, dihydro-orotase (CAD). Mechanisms involving bbf cycles account for only a minority of CAD amplification events in the human fibrosarcoma cell line HT 1080. Here, formation of a 2p isochromosome and overexpression of CAD by other types of amplification events (and even without amplification) are much more prevalent. Broken DNA is recognized by mammalian cells with intact damage-recognition pathways, as a signal to arrest or to die. Loss of these pathways by, for example, loss of p53 or pRb tumour suppressor function, or by increased expression of
ras
and
myc
oncogenes, causes non-permissive rat and human cells to become permissive both for amplification and for other manifestations of DNA damage. In cells that are already permissive, amplification can be stimulated by overexpressing oncogenes such as
c-myc
or
ras
, or by damaging DNA in a variety of ways. To supplement genetic analysis of amplification in mammalian cells, an amplification selection has been established in
Schizosaccharomyces pombe
. Selection with LiCl yields cells with amplified
sod2
genes in structures related to those observed in mammalian cells. The effect on amplification in
S. pombe
can now be tested for any mutation in a gene involved in repair of damaged DNA or in normal cellular responses to DNA damage.
Comparative analysis of different laser systems to study cellular responses to DNA damage in mammalian cells