Conservation of chromatin recruitment mechanisms in metazoan DNA replication licensing factors

University Of Texas Southwestern Medical Center, Dallas TX

Investigators

Abstract

Life requires the faithful transmission of genetic information from parents to offspring through replication of genomic DNA. All animals must replicate their DNA, yet the proteins required for replication differ substantially between animals. This project will investigate similarities and differences in DNA replication across the animal kingdom by studying the process in evolutionarily ancient and more modern animals. The studies will extend from sea sponge to humans and should clarify how DNA replication is initiated and how evolution might result in differences in replication proteins while still maintaining their function. The project will also provide summer research opportunities to community college students who would not otherwise have the opportunity to train in an academic laboratory at an R1 institution. This broader impacts program is specifically aimed at increasing participation in science and encouraging student transition to four-year STEM degree and Ph.D. programs. The first step in DNA replication is loading of the replicative helicase, Mcm2-7 onto chromatin. This licensing process requires three proteins: Orc1-6, Cdc6, and Cdt1. Fly Orc1 possesses an intrinsically disordered region (IDR) that is necessary and sufficient for chromatin recruitment in vivo and mediates DNA-dependent phase separation in vitro. These activities are negatively regulated by phosphorylation. Interestingly, while all metazoan Orc1 orthologs possess IDRs, these regions are highly variable in length and lack linear sequence similarity, raising questions about functional conservation. Orc1 IDR orthologs do share similar amino acid composition, leading to the hypothesis that sequence composition may define the functions of this IDR class. This hypothesis will be tested by studies that investigate in vitro DNA binding and in vivo chromatin recruitment activity of different Orc1 IDR orthologs. The outcomes are expected to advance understanding of the conservation of replication licensing mechanisms and will also broadly inform the study of intrinsically disordered proteins by developing the concept of compositional homology, which may be applicable to many IDR classes. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.