Investigation of the Causes of Breast Cancer at the Cellular Level: Isolation of In Vivo Binding Sites of the Human Origin Recognition Complex
Annual summary rept. 1 Jul 1999-1 Jul 2002
COLD SPRING HARBOR LAB NY
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We study the process of DNA replication in proliferating human cells. Our efforts are directed to the identification and characterization of proteins that promote DNA replication initiators as well as the DNA sequences recognized by them replicators . We have focused in a group of initiator proteins called Origin Recognition Complex ORC, Cdc6, and the Mini-Chromosome Maintenance MCM proteins. hOrclp, the largest subunit of ORC, appears to be a critical factor for the coordination of DNA replication with the cell division cycle. hOrclp levels are higher between the exit of mitosis and the end of Cl, the stage at which initiator proteins are assembled at the origins of replication. As cells enter S phase, hOrclp is polyubiquitinated on chromatin and degraded by the proteasome. hOrclp destruction is signaled in part by the SCFskP2 ubiquitin conjugating machinery. The controlled destruction of hOrclp likely contributes to avoid DNA overreplication, a common phenomenon in cancer cells. Our efforts to identify DNA replicator sequences using chromatin immunoprecipitation ChIP assays support the hypothesis that human origins of replication may not be defined by short specific DNA sequences but rather by higher-order chromatin structures.
- Genetic Engineering and Molecular Biology
- Medicine and Medical Research