Lethal and mutagenic effects of nitrosoguanidine on synchronized Chlamydomonas

The lethal and mutagenic effects of 5 mug/ml N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) were maximal during the nuclear S-period of synchronously grown Chlamydomonas reinhardtii. This was revealed by a 50% drop in survival and a 50- to 100-fold increase in the recovery of slow-growth mutants (up to 40% of the survivors) which were first recognized as small colonies on agar medium. Partial characterization of these isolates revealed about 50% to be stable on subculture, and several were demonstrated to be either acetate-dependent, dark-lethal (light-dependent), or acetate-sensitive mutants. There was no significant increase of lethality or of slow-growth mutants correlated with treatment during the chloroplast DNA replication phase of the cell-cycle. The results of genetic analysis with 13 mutants induced during the nuclear S-period were consistent with their nuclear origin. These analyses were hampered by the high proportion of lethality among the progeny of most crosses. It is concluded that the enhanced mutant induction among nuclear S-phase cells may indicate preferential mutagenesis of replication fork DNA and induction of multiple-closely-linked mutations, as in some bacteria. Consequently, for C. reinhardtii, caution should be exercised in drawing relationships between abnormal behavioral and biochemical phenotypes in MNNG-induced mutants.