Highly mutable sites for ICR-170-induced frameshift mutations are associated with potential DNA hairpin structures: studies with SUP4 and other Saccharomyces cerevisiae genes.

Abstract

The majority of the mutations induced by ICR-170 in both the CYCI gene (J. F. Ernst et al. Genetics 111:233-241, 1985) and the HIS4 gene (L. Mathison and M. R. Culbertson, Mol. Cell. Biol. 5:2247-2256 of the yeast Saccharomyces cerevisiae were recently shown to be single G-C base-pair insertions at monotonous runs of two or more G -C base pairs. However, not all sites were equally mutable; in both the CYCI and HIS4 genes there is a single highly mutable site where a G -C base pair is preferentially inserted at a sequence. Here we report the ICR-170 mutagen specificity at the SUP4-o tyrosine tRNA gene of yeast.C&netic fine structure analysis and representative DNA sequence determination of mutations revealed that there is also a single highly mutable site in SUP4-o and that the mutation is a G C base-pair insertion at a monotonous run of G C base pairs. Analysis of DNA sequences encompassing the regions of highly mutable sites for all three genes indicated that the mutable sites are at the bases of potential hairpin structures; this type of structure could not be found at any of the other, less mutable G-C runs in SUP4, CYCI, and HIS4. Based on these results and recent information regarding novel DNA structural conformations, we present a mechanism for ICR-170-induced mutagenesis: (i) ICR-170 preferentially binds to DNA in the j conformation; factors that increase the temporal stability of this structure, such as adjacent stem-and-loop formation, increase the frequency of ICR-170 binding; (ii) the observed mutagen specificity reflects formation of a preferred ICR-170 intercalative geometry at 41 sites; (iii) during replication or repair, ICR-170 remains associated with the single-stranded template; stuttering or strand slippage by the polymerization complex as it encounters the mutagen results in nucleotide duplication; (v) subsequent replication or mismatch repair fixes the insertion into the genome. This mechanism accounts for both the ICR-170 mutagenic specificity and the molecular basis of the highly mutable sites in S. cerevisiae.The mechanisms by which frameshift mutations occur have been the subject of intense interest since the original description of this class of mutations in 1%1 (7). Streisinger et al. (48) proposed a general model to account for frameshift mutagenesis, suggesting that these mutations occur in regions of strand discontinuity, for example, as occurs during replication or repair, and that the actual frameshift is generated within sequences of base-pair redundancy. Additions or deletions therefore arise as a result of strand slippage in either the primer or the template strand, respectively. This model accounted for the increased frequency of frameshift mutations induced with intercalating agents (27, 28) by suggesting that these compounds stabilize mispaired sequences. A corollary to this model is an explanation of the molecular basis of highly mutable sites, or "hotspots," initially described by Benzer (4). If mispairing is promoted in regions of monotonous stretches of base pairs, ...

Frameshift mutations induced by three classes of acridines in the lacZ reversion assay in Escherichia coli: Potency of responses and relationship to slipped mispairing models

“…QM exhibits enhanced induction of ϩG frameshifts relative to its nonreactive counterpart quinacrine (Tables II, IV). A tendency for acridine mustards to induce mainly ϩ1 frameshifts is also evident among mutations induced by ICR-170 in yeast and Neurospora [Burns et al, 1986;Hampsey et al, 1986] and by in the Hprt gene of TK6 human lymphoblasts [Taft et al, 1994]. No frameshifts were observed in A:T runs in the Hprt assay [Taft et al, 1994].…”