4 Activation of Pre-replication Complexes

Abstract

Eukaryotic organisms contain large genomes that must be duplicated rapidly and precisely in preparation for mitosis. Cells achieve rapid DNA replication by initiating DNA synthesis every 10–100 kb along the chromosome at sites called origins of DNA replication (reviewed in Chapter 2). The initiation of DNA synthesis at origins involves a remarkably complicated and highly regulated cascade of events that culminates in the assembly of the replisome, whose key components are a DNA helicase and DNA polymerases. The first event in the initiation of DNA replication takes place in the G₁ phase of the cell cycle and involves the assembly of a pre-replication complex (pre-RC) consisting of the origin recognition complex (ORC), Cdc6, Cdt1, and MCM2-7, the presumptive replicative DNA helicase. The mechanism of pre-RC assembly is reviewed in Chapter 3. Second, the replicative DNA helicase is activated, and origins are unwound. In the third and final phase, DNA polymerases are recruited (as described below, the distinction between the second and third steps is not absolute in budding yeast). This review focuses on the events of helicase activation, origin unwinding, and polymerase recruitment, which lie downstream from pre-RC formation. Among these steps, helicase activation seems to be the most complex, as it requires two protein kinases (CDK and DDK) and no fewer than six auxiliary factors (Mcm10, Dpb11, Sld2, GINS, Cdc45, and Sld3). We do not discuss why origins undergo only a single initiation event per cell cycle, as this is the topic of Chapters 15 and 16, nor...