Bisulfite Induces Tandem Double CC .fwdarw. TT Mutations in Double-Stranded DNA. 2. Kinetics of Cytosine Deamination

Abstract

Deamination of cytosine to uracil in double-stranded DNA (ds DNA) by sodium bisulfite has been monitored with a sensitive genetic assay. In this system, reversion of a mutant in the lacZ alpha gene coding sequence of bacteriophage M13mp2 C141 was detected by employing an ung- bacterial strain defective in the enzyme uracil glycosylase. Within the 4-base target, it is possible to measure the rates of induction of C-->T, C-->A, C-->G, and CC-->TT mutations in DNA that has been incubated at physiological temperature and pH and then transfected into ung+ and ung- E. coli cells, respectively, for amplification and detection of the mutation. For concentrations of bisulfite from 1 to 50 mM, the reversion frequency in ung- cells increased linearly with time of incubation. The most interesting features of the bisulfite reaction were as follow: (1) Mutations were reduced 5-fold in ung+ cells, indicating ung is involved in repair of bisulfite-treated transforming DNA. (2) Sequencing of 157 revertants revealed that C-->T and tandem CC-->TT transition mutations comprised 100% of the mutations scored. (3) A unique finding was that, at the highest concentrations and longest incubation times, almost every mutant obtained in ds DNA exposed to bisulfite was found to be a CC-->TT tandem double mutation. (4) The high frequency of tandem double mutants is inconsistent with two random, independent mutational events and, coupled with the observed ung dependence, lends support to the concept of catalytic deamination, wherein bisulfite induces deamination in contiguous cytosines by a concerted mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)

Far‐UV‐lnduced Dimeric Photoproducts in Short Oligonucleotides: Sequence Effects

“…The latter mutation is a predominant event in the mutagenicity of solar light (41) and the induction of cancers associated with sun exposure (42). However, CC + TT tandem mutations are also observed after exposure to various agents including reactive oxygen species (43), peroxynitrite (44) and bisulfite (45). Because the latter damaging agents are most unlikely to induce the formation of dimeric pyrimidine photoproducts within DNA, the chemical structure of the far-UV-induced CC lesion involved in the induction of tandem mutation may be different from a photoproduct.…”

The genotoxic effect of potassium metabisulfite using chromosome aberration, sister chromatid exchange, micronucleus tests in human lymphocytes and chromosome aberration test in bone marrow cells of rats

“…The test substance is converted in aqueous solution to the bisulfite and sulfur dioxide such as sodium metabisulfite. Bisulfite (HSO 3 ) causes the deamination of cytosine in both double-stranded and single-stranded DNA and in RNA Simmons and Friedberg, 1979;Chen and Shaw, 1994]. Hydrolytic deamination has been shown under acidic conditions, but it is of minor importance at pH 5,3.…”

Tandem-base mutations occur in mouse liver and adipose tissue preferentially as G:C to T:A transversions and accumulate with age

“…TBM are associated most commonly with ultraviolet light, appearing as CC to TT transitions [Hutchinson, 1994]. Bisulfite [Chen and Shaw, 1994] and reactive oxygen species [Reid and Loeb, 1993] have also been shown to cause CC to TT TBM, while acetaldehyde [Matsuda et al, 1998] and cis-diamminedichloroplatinum (II) [Bubley et al, 1991] cause GG to TT TBM. All mice were housed in isolated air-filtered cages within a barrier facility, thereby minimizing any air-borne mutagens.…”