Establishment and Function of Chromatin Architecture at Eukaryotic Chromosome Replication Origins

Abstract

The Origin Recognition Complex (ORC) is essential for the initiation of eukaryotic chromosome replication as it loads the replicative helicase, the minichromosome maintenance (MCM) complex, at replication origins1. These origins display a stereotypic chromatin structure. They are nucleosome depleted at ORC binding sites and flanked by regularly spaced nucleosome arrays2,3. Although discovered over a decade ago, we still do not know how this chromatin struture is established and if it matters for replication. Here we show that ORC has an essential role in generating the origin chromatin structure. By genome-wide biochemical reconstitution we screened ~ 400 individual origins and 17 purified chromatin factors from budding yeast and found that ORC establishes both nucleosome depletion over origins as well as flanking nucleosome arrays by orchestrating the four chromatin remodelers INO80, ISW1a, ISW2 and Chd1. The importance of ORC’s chromatin organizing activity was underscored by ORC mutations that maintained classical MCM loader but lost array generation activity. These mutations were lethal in vivo. Our results demonstrate that ORC, besides its canonical role as MCM loader, has a second essential function as master regulator of chromatin structure at replication origins, which is critical for chromosome replication.