Project Summary/Abstract Eukaryotic chromosomes must be efficiently compacted such that the underlying DNA must fit into the nucleus yet be precisely organized to facilitate genome function. Essential nuclear processes, including gene expression, rely on a precise, non-stochastic organizational mechanisms for the proper regulation of chromosomal structures and the correct temporal and spatial control of mRNA synthesis. However, it is presently unclear if genome topology directly influences gene expression or is strictly required for the structural organization of the genome. In human neuroblastoma and pancreatic cancers that can arise following large genome rearrangements, extensive gene expression changes have been observed, implicating genome organization as an important factor for gene control. Further, many other genetic or epigenetic factors have not been examined for whether they are necessary for organizing genomic DNA. The long-term goal of this project is to elucidate how genome topology is established in eukaryotic nuclei and dissect the role of genome organization on transcriptional control. To this end, we will use the innovative fungal organism Neurospora crassa as a model for human systems, as its compaction properties and epigenetic regulation mirror that of metazoans, yet its smaller genome is amenable to genomic studies, and it is less complex and genetically tractable. The research proposed in this application will examine genome organization changes by chromosome conformation capture coupled with high-throughput sequencing (Hi-C) in different genetic backgrounds to understand the underlying, possibly mechanistic, factors establishing genome topology and the transcriptional outcomes following topological alterations. Given that compaction of silent regions within eukaryotic genomes drive its organization, genetic manipulations followed by bioinformatics will examine the regional topological control of underlying silent DNA and if transcriptional regulation can be achieved through long-range chromosomal interactions. Further, consequences of large genome rearrangements upon the genome topology and gene expression will help elucidate the role of genome organization on normal transcriptional regulation as well as how disorder of long-range interactions in oncogenic tissues in metazoans can disrupt gene control, as examined by molecular biology, genomics, and bioinformatic approaches. Finally, variations in the post-translational modifications of histones that epigenetically demarcate genomic regions may influence the genome architecture, either directly through changing the clustering of chromatin in the nucleus, or indirectly by altering the expression of a protein controlling genome topology; altering expression of the machinery catalyzing epigenetic marks may be a method cells use to induce plasticity to genome organization, a possibility we will explore using genomics and bioinformatics. Together, our multi- faceted approach will examine how genome topology is formed and how its alteration impacts genome function ? topics that directly influence the normal function of human genomes and can cause cancerous tumors to develop upon genome disorder.

R15GM140396

2020-09-10

2023-08-31