Figure 4.

PrimPol Functions during an Unperturbed S Phase

(A) His-tagged PrimPol (PrimPol at 12 ng/μl) was added to Xenopus egg extract supplemented with sperm nuclei. Extracts were treated with geminin (80 nM), roscovitine (0.5 mM), or aphidicolin (100 μg/ml) and incubated at 21°C. At the indicated times (minutes), chromatin was isolated and associated proteins subjected to western blot analysis with the antibodies indicated.

(B) Experiment in (A) was repeated at a 60 min time point, and the last lane corresponds to a sample preincubated with geminin followed by aphidicolin treatment.

(C) Analysis of metaphase aberrations in mock and aphidicolin-treated primary MEFs lacking PrimPol. Percentage and type of aberrations per chromosome are indicated. Examples of two chromatid breaks and two rearrangements from PrimPol-deficient cells are shown. See Figure S4 for details on the knockout cells.

(D) Model of PrimPol-mediated replication fork progression. Following DNA replication stalling (depicted on the leading strand), PrimPol could reprime DNA synthesis downstream of the lesion to allow DNA replication to continue. PrimPol can also catalyze TLS of some DNA lesions and could directly extend the stalled primer terminus facilitating replication fork progression. With regards to UV-damaged templates, PrimPol could function in the error-free extension from CPDs and the error-prone bypass of 6-4 PPs.