A model to explain these observations. Components: ORC and MCM2-7 are protein complexes comprising the pre-replicative complex. Blue cylinders represent nucleosomes, with dark blue indicating closed/repressive chromatin and light blue indicating open/accessible chromatin. Red proteins are limiting replication initiation factors (such as Cdc45 and Sld3). Txn = transcription. Sequence of events: in G1 (not depicted), the limiting replication initiation factors (red circles) associate with the earliest-firing ORIs (top row). When S phase begins, these early ORIs fire and release the factors, which are then free to associate with other ORIs (though note that Cdc45 is a component of the replication fork, so can only be recycled after fork termination). The relative affinities of the remaining ORIs for these factors - and thus their relative firing times - are determined by the chromatin state near the ORI during S-phase. ORIs near genes highly transcribed in S phase (middle row) have an accessible chromatin structure and thus high affinity, so will tend to fire earlier than those with little nearby S-phase transcription and thus less accessible chromatin (bottom row). Although not shown here, subnuclear positioning could help determine ORI accessibility, either by influencing chromatin structure or through other mechanisms. Figure adapted from .