ORC associates with the Nup107-160 subcomplex, coupling nucleoporins to replication initiation
Authors: Logan Richards 1; Christopher Lord 1; Mary-Lauren Benton 2; John Capra 1,3; Jared Nordman 1
Affiliations: 1) Department of Biological Sciences, Vanderbilt University, Nashville, TN ; 2) Department of Computer Science, Baylor University, Waco, TX ; 3) Bakar Computational Health Sciences Institute and Department of Epidemiology and Biostatistics, UCSF, San Francisco, CA
Keywords: l. DNA replication; h. nuclear pore complex
The regulation of DNA replication is critical to ensure the accurate duplication of genetic information and maintain genome stability. Replication initiates at thousands of replication start sites throughout the genome. The origin recognition complex (ORC) binds throughout the genome to initiate DNA replication. In metazoans, however, it remains largely unknown how ORC is targeted to replication origins to facilitate helicase loading and replication initiation. We hypothesized that ORC’s genomic binding was driven through protein-protein interactions. To address this, we performed immunoprecipitations coupled with mass spectrometry for ORC2, a subunit of ORC, in Drosophila embryonic extract. Surprisingly, we found that ORC2 associates with several subunits of the Nup107-160 subcomplex of the nuclear pore. We determined that this interaction is developmentally regulated, occurring most strongly in the first six hours of embryogenesis. Bioinformatic analysis revealed that, relative to all modENCODE factors, nucleoporins are the most enriched factors at ORC2 binding sites. Critically, ORC2 binding to chromatin is dependent on ELYS, a member of the Nup107-160 complex. Consistent with a function in ORC2 loading, knock down of ELYS delays S phase entry and impairs DNA synthesis. Our work reveals a new connection between ORC, replication initiation, and the nuclear pore. We propose that specific nucleoporins have a previously unrecognized function in metazoan replication initiation.