Molecular analysis of mutations induced at the hisD3052 allele of Salmonella by single chemicals and complex mixtures.

Abstract

More single chemicals and complex environmental mixtures have been evaluated for mutagenicity at the hisD3052 allele of Salmonella, primarily in strain TA98, than in any other mutation assay. The development of colony probe hybridization procedures and the application of the polymerase chain reaction and direct DNA sequencing has permitted rapid molecular access to this allele. We discuss these techniques and the resulting mutation spectra that have been induced by a variety of environmental mutagens and complex mixtures. A common GC or CG deletion within a hot-spot region of the sequence dominates most of the spectra. In addition to this two-base deletion, we have recovered about 200 other types of mutations within the 72-base target for reversion of the hisD3052 allele. These include a variety of deletions (as large as 35 bases), duplications (as large as 46 bases), and complex mutations involving base substitutions. The quasipalindromic nature of the target sequence and its potential to form DNA secondary structures and slippage mismatches appear to be an important basis for the mutability of this allele.

Insights Into Mutagenesis Using Escherichia coli Chromosomal lacZ Strains That Enable Detection of a Wide Spectrum of Mutational Events

“…An important class of mutational hotspots includes short imperfect inverted repeat sequences, also known as quasipalindromes (QPs) (Figure 2A). These can be seen in mutational spectra in bacteriophage, bacteria, yeast, and in mutations responsible for several human genetic diseases (Stewart and Sherman 1974;Ripley 1982;De Boer and Ripley 1984;Mo et al 1991;Demarini et al 1993;Cebula 1995;Greenblatt et al 1996;Bissler 1998;Yoshiyama and Maki 2003;Schultz et al 2006). Genetic analysis of the mechanism of mutagenesis suggests that these hotspot mutations, which always improve the perfection of the inverted repeat sequence, occur by template-switch reactions during normal replication (Viswanathan et al 2000;Dutra and Lovett 2006), as proposed initially by Ripley (1982).…”

Induction of Mutation Spectra by Complex Mixtures: Approaches, Problems, and Possibilities

“…A related phenomenon is that all of the complex mixtures (as well as nearly all single compounds) that we have evaluated at the hisD3052 allele induce a significant increase in the frequency of the hot spot two-base deletion (18). Most (>90%) of the revertants induced in TA98 by combustion emissions and air samples (and their respective fractions) contain the hotspot deletion (18).…”

“…Most (>90%) of the revertants induced in TA98 by combustion emissions and air samples (and their respective fractions) contain the hotspot deletion (18). In this regard, these complex mixtures do not produce mutation spectra that are distinct from each other.…”

“…The approach to the molecular analysis described above is applicable to mutations induced at the hisD3052 allele by either single agents or complex mixtures Recently, we have begun to merge these two technologies for the purpose of identifying the mutations and generating mutation spectra induced by various types of complex mixtures, including an unfractionated and a neutral and base fraction of an ambient urban air sample (16), main-stream and side stream cigarette smoke condensate (17), and municipal waste incinerator partides (18). Our approach has involved the generation of mutation spectra from the unfractionated organic extract of a mixture as well as from selected mutagenic fractions from the mixture.…”

Mutation spectrum in Salmonella induced by environmental tobacco smoke

“…Molecular analysis of revertants of these strains has been simplified by the development of probe colony hybridization, polymerase chain reaction (PCR), and DNA sequence analysis. Details of the application of these methods to the analysis of revertants induced by complex mixtures have been discussed by DeMarini et al [1993DeMarini et al [ , 1994b.…”