Eukaryotic replication origins: Strength in flexibility

Abstract

The eukaryotic replicative DNA helicase, Mcm2-7, is loaded in inactive form as a double hexameric complex around double-stranded DNA. To ensure that replication origins fire no more than once per S phase, activation of the Mcm2-7 helicase is temporally separated from Mcm2-7 loading in the cell cycle. This 2-step mechanism requires that inactive Mcm2-7 complexes be maintained for variable periods of time in a topologically bound state on chromatin, which may create a steric obstacle to other DNA transactions. We have recently found in the budding yeast, Saccharomyces cerevisiae, that Mcm2-7 double hexamers can respond to collisions with transcription complexes by sliding along the DNA template. Importantly, Mcm2-7 double hexamers remain functional after displacement along DNA and support replication initiation from sites distal to the origin. These results reveal a novel mechanism to specify eukaryotic replication origin sites and to maintain replication origin competence without the need for Mcm2-7 reloading.

Keywords: DNA replication; Mcm2-7; ORC; chromosomes; replication origin.

Figure 1.

Activity states of the replicative DNA helicase at bacterial and eukaryotic replication origins. In bacteria the replicative DNA helicase is loaded in active form around single-stranded DNA at the origin (left). In eukaryotes, the replicative DNA helicase is loaded in inactive form around double-stranded DNA at the origin; activation of the helicase is temporally separated from helicase loading in the cell cycle (right). The initiator protein(s) direct the loading of the replicative DNA helicase. G1 and S indicate the G1 phase and S phase cell cycle stages in eukaryotes.

Figure 2.

Two levels of budding yeast origin flexibility. Mcm2-7 complexes are preferentially loaded at canonical origin sites that contain a high-affinity ORC binding site (top). Mutation of the high-affinity ORC binding site at a canonical origin can redirect ORC to lower-affinity binding sites in the template, resulting in Mcm2-7 loading at non-canonical origin sites (bottom left). Collisions between Mcm2-7 DHs and other DNA translocases at replication origins can induce the sliding of Mcm2-7 to non-canonical origin sites (bottom right)/ ARS: Autonomously replicating sequence.