Inhibition or Absence of DNA Proofreading Exonuclease is Not Sufficient to Allow Copying of Pyrimidine Dimers

Abstract

Speculation about the role of the 3′ to 5′ proofreading exonuclease function of DNA polymerases during UV mutagenesis has led us to examine the extents of DNA synthesis performed by a variety of different DNA polymerases, upon intact and UV-irradiated OX174 DNA. The relative inhibition of DNA synthesis after UV-irradiation in the presence of the heavy metal ions Ag+ and Hg++, and use of the T4 pyrimidine dimer specific endonuclease, confirms that the majority of the UV-induced inhibition of DNA synthesis is due to the presence of pyrimidine dimers in the template DNA. Experiments with DNA polymerases lacking 3′ to 5′ exonuclease activity, or with inhibited or denatured 3′ to 5′ exonuclease activity, revealed that pyrimidine dimers constitute an absolute block for DNA synthesis, irrespective of 3′ to 5′ exonuclease proofreading activity. It is therefore concluded that, although it might be required, inhibition or absence of 3′ to 5′ proofreading activity is not sufficient to allow efficient transdimer synthesis.