Identification of a strand-related bias in the PCNA-mediated bypass of spontaneous lesions by yeast Poleta.
Translesion synthesis (TLS) DNA polymerases are specialized to bypass lesions that block replicative polymerases and prevent complete genome duplication. Current TLS models hypothesize that PCNA, the polymerase processivity clamp, is important for regulating the access and loading of the low fidelity TLS polymerases onto DNA in response to replication-blocking lesions. PCNA binds to the C-terminus of yeast Poleta, for example, and this interaction is required for cell survival after UV irradiation. Previously, we identified two spontaneous, Polzeta-dependent "complex" mutation hotspots using the lys2DeltaA746 frameshift reversion assay in repair-compromised cells. In the current study we observed an accumulation of Polzeta-dependent complex frameshifts at a third hotspot in Poleta-deficient cells. Interestingly, the sequence of this third hotspot is the reverse complement of the two hotspots previously identified, suggesting that the utilization of Polzeta and Poleta may be related to the position of the relevant lesion on either the leading- or lagging-strand template. Using the lys2DeltaA746 assay system, we investigated changes in the accumulation of complex events at hotspots when the direction of replication was reversed in repair-compromised cells with either wildtype Poleta, a deletion of Poleta, or a mutant of Poleta that cannot interact with PCNA. Our results suggest that there is a polymerase hierarchy between Poleta and Polzeta in the bypass of certain lesions and that the interaction of Poleta with PCNA is needed for some, but not all, spontaneous lesion bypass.
Abdulovic
Amy L
AL
Graduate Program in Biochemistry, Cell and Developmental Biology Program, Emory University, Atlanta, GA 30322, United States.
Minesinger
Brenda K
BK
Jinks-Robertson
Sue
S
D001483:Base Sequence; D004249:DNA Damage; D004260:DNA Repair; D004261:DNA Replication; D004271:DNA, Fungal / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D008969:Molecular Sequence Data; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D000099270:Y-Family DNA Polymerases
D004271:DNA, Fungal; D004259:DNA-Directed DNA Polymerase; D018809:Proliferating Cell Nuclear Antigen; C112338:Rad30 protein; D000099270:Y-Family DNA Polymerases