Spectra of Spontaneous Frameshift Mutations at the hisD3052 Allele of Salmonella typhimurium in Four DNA Repair Backgrounds

Abstract

Abstract To characterize the hisD3052 −1 frameshift allele of Salmonella typhimurium, we analyzed ~6000 spontaneous revertants (rev) for a 2-base deletion hotspot within the sequence (CG)4, and we sequenced ~500 nonhotspot rev. The reversion target is a minimum of 76 bases (nucleotides 843–918) that code for amino acids within a nonconserved region of the histidinol dehydrogenase protein. Only 0.4–3.9% were true rev. Of the following classes, 182 unique second-site mutations were identified: hotspot, complex frameshifts requiring ΔuvrB + pKM101 (TA98-specific) or not (concerted), 1-base insertions, duplications, and nonhotspot deletions. The percentages of hotspot mutations were 13.8% in TA1978 (wild type), 24.5% in UTH8413 (pKM101), 31.6% in TA1538 (ΔuvrB), and 41.0% in TA98 (ΔuvrB, pKM101). The ΔuvrB allele decreased by three times the mutant frequency (MF, rev/108 survivors) of duplications and increased by about two times the MF of deletions. Separately, the ΔuvrB allele or pKM101 plasmid increased by two to three times the MF of hotspot mutations; combined, they increased this MF by five times. The percentage of 1-base insertions was not influenced by either ΔuvrB or pKM101. Hotspot deletions and TA98-specific complex frameshifts are inducible by some mutagens; concerted complex frameshifts and 1-base insertions are not; and there is little evidence for mutagen-induced duplications and nonhotspot deletions. Except for the base substitutions in TA98-specific complex frameshifts, all spontaneous mutations of the hisD3052 allele are likely templated. The mechanisms may involve (1) the potential of direct and inverted repeats to undergo slippage and misalignment and to form quasi-palindromes and (2) the interaction of these sequences with DNA replication and repair proteins.