Affiliation: | 1 Institute of Environmental Medicine, New York University Medical Center, 550 First Avenue, New York, NY 10016, USA 2 Joseph Gottstein Memorial Cancer Research Laboratory, Department of Pathology SM-30, University of Washingon, Seattle, WA 98195, USA 3 Laboratory of Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, U.S.A. |
Abstract: | We have addressed the possibility of terminal transferase involvement in somatic mutagenesis and the creation of N-region diversity, by measuring the ability of TdT to enhance single-base substitution mutagenesis during in vitro DNA synthesis. Using 3 independent assays we find that terminal transferase produces only a small increase in base-substitution mutagenesis when assayed in the presence of DNA polymerase-β. In the presence of either polymerase- or E. coli polymerase-I, however, no detectable increase in TdT-induced mutagenesis is seen. Furthermore, in an assay capable of detecting a variety of mutational events, terminal transferase primarily produces complex addition/deletion mutations, as well as a few multiple, tightly-clustered, single-base mutations. We conclude that the majority of the scattered single-base changes that occur during antibody gene differentiation are not catalyzed by terminal transferase, but instead result from another error-prone DNA synthetic process (possibly utilizing DNA polymerase-β). |