ANALYZING CHROMATIN STRUCTURE AND TRANSCRIPTION FACTOR-BINDING IN YEAST

The study of chromatin, once thought to be a purely structural matrix serving to compact the DNA of the genome into the nucleus, is of incre asing value for our understanding of how DNA functions in the cell. Th is article provides two basic procedures for the study of chromatin in vivo. The first is a DNase I-based method for the treatment of isolat ed nuclei to resolve the chromatin structure of a particular region; t he second employs dimethyl sulfate footprinting of whole cells in vivo to determine the binding of factors to cis elements in the locus of i nterest. Specific examples illustrating the techniques described are g iven from our work on the regulation of the yeast PHO8 gene, but have also been successfully and reliably applied to the study of many other yeast loci. These procedures make it possible to correlate the bindin g of a transactivator with an altered or perturbed chromatin organizat ion at a specific locus. (C) 1998 Academic Press.