We propose to study the structure of chromosomes from the yeasts S. cerevisiae and S. prombe, using a combination of biochemical and genetic approaches. The proposed experiments build on results that we have already obtained with yeast chromatin (see below) and complement our continuing studies of higher cell chromatin structure. Our first goal is to identify proteins (and their genes) that are involved in the higher order folding of the nucleosome filament at different points in the cell cycle. including stationary phase and points in interphase, mitosis and meiosis. Current models for the structure of mitotic and meiotic chromosomes (from higher cells) imply that key proteins necessary for the highest levels of folding are likely to be present in trace stoichiometries relative to histones; we therefore propose genetic screens that will allow us to identify these proteins. We have evidence that yeast have a histone H1-line activity; therefore, in addition to general screens for proteins important for chromosome folding, we have devised several different specific functional assays to find the yeast equivalent of H1. We will carry out experiments designed to elucidate the function of each of the proteins that we identify. We expect to gain insights into the three-dimensional structures that are formed at different levels of chromosomal folding, by structural studies of chromatin from mutant yeast strains that are deficient in own or another of these proteins. Finally, we will as far as possible extend the results of our studies on yeast chromatin to higher cells.
