9304512 Akey Comparative studies of yeast and mammalian nuclei are providing important insights into complex cellular processes such as control of the cell cycle, mitosis, meiosis and nucleocytoplasmic transport. Moreover, the well established area of yeast genetics allows rapid correlation between structural alterations at the genetic level with changes in cell behavior and functions of supra-molecular assemblies. In this project, the structure and function of a supra-molecular assembly which resides within pores in the yeast nuclear envelope will be studied: the spindle pole body (SPB). Electron microscopy of partially enriched cell fractions will be employed, together with three-dimensional (3D) image analyses, to reveal structural principles which govern the assembly and functions of these complex supra molecular structures. In the case of SPBs, it has recently been shown that these are highly ordered and amenable to structural studies. Moreover, there are two lateral size classes present in in vitro isolates. These will be studied using 3D tomography of both thin sectioned and frozen-hydrated specimens. These studies should reveal the architectural principles which govern the assembly and fusion pathways traversed by the SPB during normal cell growth and mating, and should also reveal molecular details of how the yeast microtubule organizing center anchors and nucleates microtubules. The work will complement ongoing biochemical and genetic analyses being carried out in other laboratories, and may broaden our understanding of centrosomal functions in other species. %%% In eukaryotic cells, the genetic material (DNA) is sequestered from the bulk cytoplasm, in a structure called the nucleus, by a double-membrane boundary, the nuclear envelope. In Saccharomyces yeast, the nuclear envelope does not break down at the time of mitosis (cell division); instead, the mitotic spindle (the microtubular structure responsible for the separation of daughter chromosomes during the mitotic p rocess) forms inside the nucleus. In these yeast cells, the spindle pole bodies (microtubule organizing centers), which are essential structures for the assembly of the mitotic spindle and the mitotic process itself, are associated with the nuclear pores. In this project, advanced methods for studying the morphology of yeast spindle pole bodies at high resolution will be used to study the structure of this cellular component. Yeast are amenable to genetic analysis, and both genetic and biochemical analyses of spindle pole body assembly are ongoing in other laboratories. The powerful combination of structural and genetic analyses in yeast will lead to an increased understanding of the essential process of cell division, and may broaden our understanding of centrosomal functions during mitosis in other eukaryotes. ***
