Endo-reduplication in mouse liver after conditional mutation of ORC2 and combined mutation of ORC1 and ORC2

ABSTRACT

The six subunit Origin Recognition Complex (ORC) is essential for loading MCM2-7 at origins of DNA replication to promote initiation of DNA replication in organisms ranging from S. cerevisiae to humans. In rare instances, as in cancer cell-lines in culture with mutations in ORC1, ORC2 or ORC5, or in endo-reduplicating mouse hepatocytes in vivo without ORC1, DNA replication has been observed in the virtual absence of individual ORC subunits. Although ORC1 is dispensable in the mouse liver for endo-reduplication, because of the homology of ORC1 with CDC6, it could be argued that CDC6 was substituting for ORC1 to restore functional ORC. Here, we have created mice with a conditional deletion of ORC2, to demonstrate that mouse embryo fibroblasts require ORC2 for proliferation, but that the mouse hepatocytes can carry out DNA synthesis in vitro and endo-reduplicate in vivo, despite the deletion of ORC2. Combining the conditional mutation of ORC1 and ORC2 revealed that the mouse liver can still carry out endo-reduplication despite the deletion of the two genes, both during normal development and after partial hepatectomy. Since endo-reduplication, like normal S phase replication, requires the presence of MCM2-7 on the chromatin, these results suggest that in primary hepatocytes there is a mechanism to load sufficient MCM2-7 to carry out effective DNA replication despite the virtual absence of two subunits of ORC.