Figure 5.
Polκ mediates faithful bypass of unhooked aldehyde ICLs, independently of Rev1. (A) Western blot analysis of mock, Polκ and Rev1 (ΔPolκΔRev1) depleted extract, and ΔPolκ ΔRev1 extract supplemented with the indicated WT or mutant Polκ proteins alongside a titration of undepleted extract. (B, C) Quantifications of lesion bypass assays on pICL-AARED (D) and pICL-ACRRED (E) repair reactions in mock, ΔPolκ ΔRev1 double depleted extract, and ΔPolκΔRev1 extract supplemented with indicated Polκ WT or mutant proteins. Based on gels in Supplementary Fig. S9C and D. (D) Distribution and frequency of nucleotide misincorporation of pICL-AARED repair products from Mock, Rev1, or Polκ depleted extract. Distributions of all mutation types (deletions, SNVs, and WT) are indicated in Supplementary Fig. S10A. (E) Model of the role of Polκ in aldehyde-ICL repair via the FA pathway. Polκ is the main insertion polymerase for unhooked aldehyde-ICL adducts during FA-mediated repair. For this, Polκ depends on its PIP domains and catalytic activity. The extension step is performed by either Rev1–Polζ or Polκ, which likely both are present at the lesion.