Subcellular localization, stoichiometry, and protein levels of 26 S proteasome subunits in yeast

The 26 S proteasome of eukaryotes is responsible for the degradation of pro teins targeted for proteolysis by the ubiquitin system. Yeast has been an i mportant model organism for understanding eukaryotic proteasome structure a nd function, Toward a quantitative characterization of the proteasome, we h ave determined the localization, cellular levels, and stoichiometry of prot easome subunits, The subcellular localization of two ATPase components of t he regulatory complex of the proteasome, Suga/Rpt4 and Sug1/Rpt6, and a sub unit of the 20 S proteasome, Pre1, were determined by immunofluorescence, I n contrast to findings in multicellular organisms, these proteins are local ized almost exclusively to the nucleus throughout the cell cycle. We have a lso determined the cellular abundance and stoichiometry of these proteasome subunits. Sug1/Rpt6, Sug2/Rpt4, and Pre1 are present in roughly equal stoi chiometry with an abundance of 15,000-30,000 molecules/cell, corresponding to a concentration of 13-26 mu M in the nucleus. Also, in contrast to mamma lian cells, we find no evidence of a p27-containing "modulator" of the prot easome in yeast. This information will be useful in comparing and contrasti ng the yeast and mammalian proteasomes and should contribute to a mechanist ic understanding of how this complex functions.