DESCRIPTION (provided by applicant): DNA segregation or partition is one of the most fundamental processes in biology and ensures that genetic information is accurately retained from one generation to the next. While the molecules involved in partition have largely been identified in prokaryotes and eukaryotes, the mechanisms that drive this process are not understood at the atomic level. Moreover, currently, very little is known about DNA segregation in archaea. The overarching goals of this proposal are to define the molecular mechanisms and principles involved in DNA segregation using model bacterial and archaeal partition (par) systems. Bacterial plasmid par systems represent excellent model systems to study segregation at an atomic level because they minimally require only 3 elements, a centromere, a centromere-binding protein (CBP) and an NTPase. The most common par system is the so-called type I, or Walker-box based system. This type of par system is involved in the partitioning of both prokaryotic chromosomes and plasmids, and includes CBPs called ParB and Walker-type NTPases called ParA. Our recent combination of structural and in vivo super-resolution studies has started to provide a detailed atomic as well as dynamic understanding of bacterial segregation. Eukaryotes appear to use machinery distinct from prokaryotes whereby their centromeres are marked by "centromeric nucleosomes", which contain CenpA in the place of histone H3 along with H4, H2A and H2B. Although these systems seem vastly disparate, our recent studies have revealed structural and, potentially, functional links between DNA segregation in the 3 domains of life. Specifically, our initial characterization of the first archaa par system, pNOB8, reveals that it employs a Walker-box NTPase similar to those used by prokaryotes while our preliminary structure of the pNOB8 ParB reveals that it contains a fold similar to the eukaryotic histone CenpA protein. Based on these findings, we hypothesize that shared structural and functional principles may exist in DNA segregation mechanisms across the domains of life. The goals of this proposal are to provide detailed structural and functional dissections of DNA segregation using two prototypical model systems; the bacterial TP228 par system and the archaeal pNOB8 par system.

PUBLIC HEALTH RELEVANCE: DNA segregation is necessary for all organisms. Understanding the molecular basis for this process in the three domains of life will reveal commonalities and differences that can be exploited for the treatment of multiple diseases and infections.