Concerted interaction between origin recognition complex (ORC), nucleosomes and replication origin DNA ensures stable ORC–origin binding

    loading  Checking for direct PDF access through Ovid

Abstract

Chromosomal replication origins, where DNA replication is initiated, are determined in eukaryotic cells by specific binding of a six-subunit origin recognition complex (ORC). Many biochemical analyses have showed the detailed properties of the ORC–DNA interaction. However, because of the lack of in vitro analysis, the molecular architecture of the ORC–chromatin interaction is unclear. Recently, mainly from in vivo analyses, a role of chromatin in the ORC–origin interaction has been reported, including the existence of a specific pattern of nucleosome positioning around origins and of a specific interaction between chromatin—or core histones—and Orc1, a subunit of ORC. Therefore, to understand how ORC establishes its interaction with origin in vivo, it is essential to know the molecular mechanisms of the ORC–chromatin interaction. Here, we show that ORC purified from yeast binds more stably to origin-containing reconstituted chromatin than to naked DNA and forms a nucleosome-free region at origins. Molecular imaging using atomic force microscopy (AFM) shows that ORC associates with the adjacent nucleosomes and forms a larger complex. Moreover, stable binding of ORC to chromatin requires linker DNA. Thus, ORC establishes its interaction with origin by binding to both nucleosome-free origin DNA and neighboring nucleosomes.

Related Topics

    loading  Loading Related Articles