REPRODUCTIVE CYCLE OF SPINDLE POLE BODIES IN FISSION YEAST

University Of Colorado At Boulder, Boulder CO

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Abstract

The fine structure of Schizosaccharomyces pombe has been preserved with unprecedented quality, using a high pressure freezer and freeze substitution fixation. The resulting EM specimens have allowed us both to analyze the 3-D fine structure of the mitotic spindles in S. pombe (Ding et al. 1993) and to investigate the structure of specific intracellular organelles, such as the spindle pole bodies (SPBs). We have now characterized the process of SPB duplication in wild-type cells. During interphase, the SPB resides in the cytoplasm, immediately associated with the nuclear envelope. It duplicates late in G2, but the two SPBs remain attached as they settle into an invagination that forms in the nuclear envelope. This membrane specialization then opens to form a special fenestra in the nuclear envelope. The duplicated SPB attaches to the edges of the fenestra, then MTs grow from the inner surface of the SPBs as they separate and move to become the poles of the forming spindle. During anaphase, the SPBs are extruded from the fenestrae at each end of the spindle and re-enter the cytoplasm, where they reside until they duplicate and repeat the cycle (Ding et al., Mol Biol. Cell). We have also fixed various strains of yeast that are mutant in genes required for normal mitosis in order to use 3-D fine structural analysis to characterize mutant phenotype and to help us understand the function of the corresponding wild type gene products in the normal cell division process. Seven cells carrying the the mitotic mutation, cut11-2 have been reconstructed in 3-D, allowing us to learn that these cells have aberrant spindle pole function. The Cut11 protein product appears to be required for normal attachment of the SPB to the nuclear envelope during mitosis (West et al., in preparation). [unreadable]