The genetic material present in chromosomes is organized in a non-random fashion within the nucleus. In most eukaryotes, the ends of chromosomes (telomeres) are associated with the nuclear membrane, and often associate with other telomeres. Whole chromosomes, and specific parts of chromosomes, can occupy unique domains within a nucleus, and in some organisms homologous chromosomes are paired in somatic cells. There are some indications that nuclear organization is required for normal cellular functions. For example, defects in telomere structure and function appear during aging, and in cancer cells., and alteration of domain structures can negatively impact gene expression and, thus, cell and organismal viability. However, in multicellular eukaryotes there are few proteins known to be involved in the organization of chromosomes within the nucleus, and very few examples of systems where cytologically-visible changes in nuclear and chromosome organization can be directly linked to altered biological functions. The powerful genetic, molecular and cell biological approaches available in the Drosophila (fruit fly) model system combines many of the molecular-genetic approaches available in yeasts with the cytological resolution, multicellularity and chromosome structure of mammals. In addition, there is a well-defined, manipulable Drosophila minichromosome (Dp1187) that has been successfully utilized in the study of higher eukaryotic centromeres and other aspects of chromosome structure and function. The experiments described in this proposal use Drosophila cytology, genetics and molecular biology to identify proteins and mechanisms involved in nuclear organization in metazoans, and to understand their regulation, properties and in vivo functions. We will 1) identify candidate Drosophila genes involved in nuclear organization and function with a genetic screen, clone the protein products, and determine their biological functions, and 2) investigate the function of candidate centromere protein genes and determine their roles in the cell using cell biological, genetic and molecular methodologies. The results of these studies will provide new information about the trans-acting components responsible for chromosome and nuclear organization in Drosophila, and will serve as a model system for further investigations in other eukaryotes. Elucidating basic information about nuclear organization components and mechanisms in multicellular eukaryotes has intrinsic interest, but is also likely to have applications to the diagnosis and treatment of aging and aneuploidy in human populations.
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