Increased expression of Pold does not alter the canonical replication program in vivo

In vitro experiments utilising the reconstituted Saccharomyces cerevisiae eukaryotic replisome indicated that the efficiency of leading strand replication is impaired by a moderate increase in Pold concentration. It was hypothesised that the slower rate of the leading strand synthesis characteristic for reactions containing 2-fold and 4-fold increased quantities of Pold represented a consequence of a rare event, during which Pold stochastically outcompeted Pole and, in an inefficient manner, temporarily facilitated extension of the leading strand. Inspired by this observation, we aimed to determine whether increased Pold levels influence replication dynamics in vivo using the fission yeast Schizosaccharomyces pombe as a model system. We provide an evidence that 2-4-fold over-production of Pold does not significantly alter cell cycle progression and the dynamics of the three replicative polymerases: Pold, Pole and Pola. Our experiments do not disapprove the existence of opportunistic polymerase switches, however, the data indicate that, if stochastic replacement of Pole for Pold does occur in vivo, it represents a rare phenomenon that does not significantly influence canonical replication program. Overall design: Polymerase usage sequencing analysis of S. pombe mutants characterised by increased expression of polymerase delta.