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Mutagenic Effects of 2-Deoxyribonolactone in Escherichia coli. An Abasic Lesion That Disobeys the A-Rule

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posted on 01.06.2004, 00:00 authored by Kelly M. Kroeger, Yu Lin Jiang, Yoke Wah Kow, Myron F. Goodman, Marc M. Greenberg
Abasic sites are often referred to as noninstructive lesions. The C1‘-oxidized abasic site (2-deoxyribonolactone, L) is produced by several DNA damaging agents, including γ-radiolysis and the neocarzinostatin chromophore (NCS). The effects of a C1‘-oxidized abasic site incorporated at a defined site in single-stranded plasmid were examined in SOS polymerase-proficient and -deficient Escherichia coli. For comparison, experiments utilizing plasmids containing an abasic site (AP) were carried out side by side. In contrast to plasmid containing AP, dA and dG were incorporated most often when plasmid containing L was replicated. The ratio of dG:dA incorporation depended upon local sequence and varied from 0.9 to 2.2. High levels of translesion incorporation of dA are consistent with previous observations that treatment of DNA with the neocarzinostatin chromophore resulted in large amounts of G·C → A·T transitions [Povirk and Goldberg (1986) Nucleic Acids Res. 14, 1417] and support the proposal that L is the source of these mutations. Both abasic lesions were 100% lethal in triple knockout cells lacking pol II, pol IV, and pol V. Analysis of translesion synthesis in repair-deficient cells revealed that pol V played a significant role in replication of L and AP. Significant levels of −1 frameshifts were formed in 5‘-d(CL) sequences in the presence of pol V and were the exclusive product in pol V-deficient cells. Frameshift products were not formed when the nucleotide on the 5‘-side of L was either dT or dG. Deleting pol II or pol IV had only modest effects on replication of L-containing plasmid but significantly decreased the amount of −1 frameshift product formed from an AP lesion. Experiments carried out side by side using otherwise identical plasmids containing an AP site illustrate the distinct properties of these two abasic lesions and that neither should be thought of as noninstructive.