Base substitutions, frameshifts, and small deletions constitute ionizing radiation-induced point mutations in mammalian cells.

Abstract

The relative role of point mutations and large genomic rearrangements in ionizing radiation-induced mutagenesis has been an issue of long-standing interest. Recent studies using Southern blotting analysis permit the partitioning of ionizing radiation-induced mutagenesis in mammalian cells into detectable deletions and major genomic rearrangements and into point mutations. The molecular nature of these point mutations has been left unresolved; they may include base substitutions as well as small deletions, insertions, and frameshifts below the level of resolution of Southern blotting analysis. In this investigation, we have characterized a collection of ionizing radiation-induced point mutations at the endogenous adenine phosphoribosyltransferase (aprt) locus of Chinese hamster ovary cells at the DNA sequence level. Base substitutions represented =2/3 of the point mutations analyzed. Although the collection of mutants is relatively small, every possible type of base substitution event has been recovered. These mutations are well distributed throughout the coding sequence with only one multiple occurrence. Small deletions represented the remainder of characterized mutants; no insertions have been observed. Sequence-directed mechanisms mediated by direct repeats could account for some of the observed deletions, while others appear to be directly attributable to radiation-induced strand breakage.Although ionizing radiation was the first known mutagen (1), relatively little is known about the molecular mechanisms of ionizing radiation-induced mutagenesis in mammalian cells.

Mutagenesis by hydrogen peroxide treatment of mammalian cells: a molecular analysis

“…Thus, in the latter case repeated DNA sequences are not required. Similar sequence-directed mechanisms of deletion formation were later seen in a small pool of both spontaneous (32) and ionizing radiation-induced mutants (28) in mammalian cells. In the present study we have confirmed the presence of these two classes of deletion mutant in mammalian cells.…”

“…see ref. 28), the system is rapid and simple and both base changes and the two termini of deletions of 100 bp or less can be analysed from a much larger database than would otherwise be possible.…”

“…Strand breaks that occur in close proximity on the same DNA strand are probably directly responsible for the deletion of 1 or 2 bp from the five remaining small deletion mutants in the H 2 O 2 series (Table V, Since the hydroxyl radical generated intracellularly via a Fenton-type reaction is probably the active intermediate leading to genetic damage after H 2 O 2 treatment, it is instructive to compare our present data with that obtained in recent studies with ionizing radiation which generates hydroxy radical by the radiolysis of water. Of 51 mutants observed in the c 1 gene of lambda prophage, 11 were 1 and 2 bp frameshift deletions (36) and 4 out of 16 of the 'point mutations' arising in the endogenous adenine phosphoribosyl transferase gene of Chinese hamster ovary cells were small deletions (28).…”

Ionizing radiation-induced mutagenesis

“…For an endogenous gene this involves considerable labour of isolating, cloning and subcloning. The efficiency of the cloning procedure can be optimized by biological selection (Maniatis et al, 1982;Grosovsky et al, 1987 (Miller & Barnes, 1986). The sensitivity of this method might be further increased by including tetramethylammonium chloride in the hybridization buffers, which abolishes the preferential melting at A.T v G.C base-pairs, thus making the procedure independent of the base composition of the probe (Wood et al, 1985).…”

A pilot study into the effects of X-ray and computed tomography exposure on the amplification of DNA from bone

“…85). Others have shown that radiation breaks DNA into fragments and causes base-change mutations (Wolff, 1971;Liber et al, 1986;Grosovsky et al, 1988).…”