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Pre‐initiation complex assembly functions as a molecular switch that splits the Mcm2‐7 double hexamer
Author(s) -
MiyazawaOnami Mayumi,
Araki Hiroyuki,
Tanaka Seiji
Publication year - 2017
Publication title -
embo reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 4.584
H-Index - 184
eISSN - 1469-3178
pISSN - 1469-221X
DOI - 10.15252/embr.201744206
Subject(s) - molecular genetics , genetics , random hexamer , library science , biology , computer science , microbiology and biotechnology , gene
Initiation of chromosomal DNA replication in eukaryotes involves two steps: licensing and firing. In licensing, a core component of the replicative helicase, the Mcm2–7 complex, is loaded onto replication origins as an inactive double hexamer, which is activated in the firing step by firing factors. A reaction intermediate called the pre‐initiation complex (pre‐ IC ) has been proposed to assemble transiently during firing, but the existence of the pre‐ IC has not yet been confirmed. Here, we show, by systematic chromatin immunoprecipitation, that a distinct intermediate that fits the definition of the pre‐ IC assembles during firing in the budding yeast Saccharomyces cerevisiae . Pre‐ IC assembly is observed in the absence of Mcm10, one of the firing factors, and is mutually dependent on all the firing factors whose association to replication origins is triggered by cyclin‐dependent kinase. In the pre‐ IC , the Mcm2–7 double hexamer is separated into single hexamers, as in the active helicase. Our data indicate that pre‐ IC assembly functions as an all‐or‐nothing molecular switch that splits the Mcm2–7 double hexamer.