Inter-origin cooperativity of geminin action establishes an all-or-none switch for replication origin licensing

In metazoans, geminin functions as a molecular switch for preventing re-replication of chromosomal DNA. Geminin binds to and inhibits Cdt1, which is required for replication origin licensing, but little is known about the mechanisms underlying geminin's all-or-none action in licensing inhibition. Using Xenopus egg extract, we found that the all-or-none activity correlated with the formation of Cdt1 foci on chromatin, suggesting that multiple Cdt1-geminin complexes on origins cooperatively inhibit licensing. Based on experimental identification of licensing intermediates targeted by geminin and Cdt1, we developed a mathematical model of the licensing process. The model involves positive feedback owing to the cooperative action of geminin at neighboring origins and accurately accounts for the licensing activity mediated by geminin and Cdt1 in the extracts. The model also predicts that such cooperativity leads to clustering of licensing-inhibited origins, an idea that is supported by the experimentally measured distribution of inter-origin distances. We propose that geminin inhibits licensing through an inter-origin interaction, ensuring strict and coordinated control of multiple replication origins on chromosomes.