We found that, under historical climates, succession changed more area annually than fire by factors of 1.2 to 34, but one model simulated more landscape change from fire (factor of 0.1).
Nevertheless, lower pol-prim levels or weakened origin association that approaches below a minimal threshold may alter the temporal and spatial pattern of origin firing in S phase, as was shown for the key origin firing factors Cdc45 and Sld3 (Mantiero et al., 2011; Tanaka et al., 2011).
Fig 6B highlights the firing factors.
Next, we examined the recruitment of firing factors to pre-RCs in the S-phase extract.
As shown in Fig 2D, overproduced firing factors were recruited to the reconstituted pre-RCs in a DDK-dependent manner.
Briefly, firing factors (Dpb11, Sld2, Sld3, Sld7 and Cdc45) were overexpressed from galactose-inducible promoters and cells were synchronised in G1 phase with alpha factor.
Here, we show that plasmids containing pre-RCs assembled with purified proteins support complete and semi-conservative replication in extracts from budding yeast cells overexpressing firing factors.
Regardless, Mcm10 is present at levels 10-100 times lower than other firing factors, suggesting that this step may be limiting in our system.
Replication initiation in vivo requires CDK, which phosphorylates the firing factors Sld2 and Sld3 and triggers origin firing (Masumoto et al, 2002; Tanaka et al, 2007; Zegerman & Diffley, 2007).
Surprisingly, few proteins other than known replisome components and firing factors were found associated in a DDK-dependent manner (Supplementary Table S1), suggesting that most or all core factors involved in initiation and elongation have been identified.
DDK phosphorylation of Mcm2-7 does not by itself promote separation of the double hexamer, but is required for the recruitment of firing factors and replisome components in the extract.
