title: Formaldehyde-induced mutagenesis in Saccharomyces cerevisiae: molecular properties and the roles of repair and bypass systems.
abstract: Although DNA-protein cross-links (DPCs) pose a significant threat to genome stability, they remain a poorly understood class of DNA lesions. To define genetic impacts of DPCs on eukaryotic cells in molecular terms, we used a sensitive Saccharomyces cerevisiae frameshift-detection assay to analyze mutagenesis by formaldehyde (HCHO), and its response to nucleotide excision repair (NER) and translesion DNA synthesis (TLS). Brief exposure to HCHO was mutagenic for NER-defective rad14 strains but not for a corresponding RAD14 strain, nor for a rad14 strain lacking both Polζ and Polη TLS polymerases. This confirmed that HCHO-generated DNA lesions can trigger error-prone TLS and are substrates for the NER pathway. Sequencing revealed that HCHO-induced single-base-pair insertions occurred primarily at one hotspot; most of these insertions were also complex, changing an additional base-pair nearby. Most of the HCHO-induced mutations required both Polζ and Polη, providing a striking example of cooperativity between these two TLS polymerases during bypass of a DNA lesion formed in vivo. The similar molecular properties of HCHO-induced and spontaneous complex +1 insertions detected by this system suggest that DPCs which form in vivo during normal metabolism may contribute characteristic events to the spectra of spontaneous mutations in NER-deficient cells.
authors.affiliation: Department of Biological Sciences, University of Cincinnati, Cincinnati OH 45221-0006, USA. grogandw@ucmail.uc.edu
authors.forename: Dennis
authors.initials: D
authors.lastname: Grogan
authors.forename: Sue
authors.initials: S
authors.lastname: Jinks-Robertson
mesh_terms: D001483:Base Sequence; D004249:DNA Damage; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D005557:Formaldehyde / Q000633:toxicity*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D000099270:Y-Family DNA Polymerases
chemical_list: D045643:DNA Repair Enzymes; D004259:DNA-Directed DNA Polymerase; D005557:Formaldehyde; D029701:Saccharomyces cerevisiae Proteins; C072975:RAD14 protein, S cerevisiae; C425929:DNA polymerase zeta; C112338:Rad30 protein; D000099270:Y-Family DNA Polymerases