Rap1-Sir4 binding independent of other Sir, yKu, or histone interactions initiates the assembly of telomeric heterochromatin in yeast.

In Saccharomyces cerevisiae, heterochromatin-like regions are found near telomeres and at the silent mating-type loci, where they can repress genes in an epigenetic manner. Several proteins are involved in telomeric heterochromatin structure including Rap1, Sir2, Sir3, Sir4, yKu70 (Hdf1), yKu80 (Hdf2), and the N termini of histones H3 and H4. By recognizing cis-acting DNA-binding sites, Rap1 is believed to recruit Sir and other silencing proteins and determine where heterochromatin forms. The integrity of heterochromatin also requires the binding of Sir proteins to histones that may form a scaffold for Sir protein interactions with chromatin. In this study we describe how the heterochromatin complex may form initially and how it differs from the complex that spreads along the chromosome. We found that close to the telomere end, Sir4 can bind Rap1 independently of Sir2, Sir3, yKu70/yKu80, and the intact H4 N terminus. In contrast, Sir4 binding requires all of the silencing factors further along telomeric heterochromatin. These data indicate that Sir4 binding to Rap1 initiates the sequential association of Sir and other proteins, allowing the subsequent spreading of the heterochromatin proteins along the chromosome.