FUNCTIONAL STUDY OF NATIVE AND SYNTHETIC NUCLEI

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A eukaryotic cell is compartmentalized into numerous membrane-enclosed organelles. Of fundamental interest to cell biologists are the mechanisms by which these organelles achieve unique compositions. The goal of this proposal is an understanding of the selective transport of macromolecules into the nucleus. Such transport occurs through nuclear pores, which perforate the nuclear envelope and comprise the sites at which selective entry is controlled. Two approaches are proposed to address the mechanism of transport through the pores: 1) analysis of a particular pore protein, a 63 kd glycoprotein, and 2) analysis of the requirements for nuclear transport itself. Using a system which allows the rapid in vitro assay of nuclear transport, we have identified a nuclear pore protein required for pore function. A molecular and functional analysis of this protein is porposed. The protein will be purified and antibodies raised against it. With these tools, fundamental questions of pore function will be asked, including how the protein is involved, the role of the carbohydrate moiety of the protein in function, and whether this protein is integral to the structure of the nucleus. In a second approach to understanding nuclear transport, experiments are proposed to determine the minimal cellular components involved in transport. It should be possible to determine whether a carrier protein is required for nuclear transport, as it is for import into other organelles. Lastly, the role of ATP in either the recognition step or the translocation step of nuclear transport will be addressed.