A highly enriched spindle pole preparation was prepared from budding yeast and fractionated by SDS gel electrophoresis. Forty-five of the gel bands that appeared enriched in this fraction were analyzed by high-mass accuracy matrix-assisted laser desorption/ ionization (MALDI) peptide mass mapping combined with sequence database searching. This identified twelve of the known spindle pole components and an additional eleven gene products that had not previously been localized to the spindle pole. Immunoelectron microscopy localized eight of these components to different parts of the spindle. One of the gene products, Ndc80p, shows homology to human HEC protein (Chen, Y., D.J. Riley, P-L. Chen, and W-H. Lee. 1997. Mol. Cell Biol. 17:6049–6056) and temperature-sensitive mutants show defects in chromosome segregation. This is the first report of the identification of the components of a large cellular organelle by MALDI peptide mapping alone.

Tub4p is a novel tubulin found in Saccharomyces cerevisiae. It most resembles gamma‐tubulin and, like it, is localized to the yeast microtubule organizing centre, the spindle pole body (SPB). In this paper we report the identification of SPC98 as a dosage‐dependent suppressor of the conditional lethal tub4–1 allele. SPC98 encodes an SPB component of 98 kDa which is identical to the previously described 90 kDa SPB protein. Strong overexpression of SPC98 is toxic, causing cells to arrest with a large bud, defective microtubule structures, undivided nucleus and replicated DNA. The toxicity of SPC98 overexpression was relieved by co‐overexpression of TUB4. Further evidence for an interaction between Tub4p and Spc98p came from the synthetic toxicity of tub4–1 and spc98–1 alleles, the dosage‐dependent suppression of spc98–4 by TUB4, the binding of Tub4p to Spc98p in the two‐hybrid system and the co‐immunoprecipitation of Tub4p and Spc98p. In addition, Spc98–1p is defective in its interaction with Tub4p in the two‐hybrid system. We suggest a model in which Tub4p and Spc98p form a complex involved in microtubule organization by the SPB.

Pericentromeric heterochromatin (PCH) gives rise to highly dense chromatin sub-structures rich in the epigenetic mark corresponding to the trimethylated form of lysine 9 of histone H3 (H3K9me3) and in heterochromatin protein 1α (HP1α), which regulate genome expression and stability. We demonstrate that Tau, a protein involved in a number of neurodegenerative diseases including Alzheimer’s disease (AD), binds to and localizes within or next to neuronal PCH in primary neuronal cultures from wild-type mice. Concomitantly, we show that the clustered distribution of H3K9me3 and HP1α, two hallmarks of PCH, is disrupted in neurons from Tau-deficient mice (KOTau). Such altered distribution of H3K9me3 that could be rescued by overexpressing nuclear Tau protein was also observed in neurons from AD brains. Moreover, the expression of PCH non-coding RNAs, involved in PCH organization, was disrupted in KOTau neurons that displayed an abnormal accumulation of stress-induced PCH DNA breaks. Altogether, our results demonstrate a new physiological function of Tau in directly regulating neuronal PCH integrity that appears disrupted in AD neurons.