EXPERIMENTAL FIGURE 9-36 Antibody and DNA probes colocalize SIR3 protein with telomeric heterochromatin in yeast nuclei. (a) Confocal micrograph 0.3 mm thick through three diploid yeast cells, each containing 68 telomeres. Telomeres were labeled by hybridization to a fluorescent telomere-specific probe (yellow). DNA was stained red to reveal the nuclei. The 68 telomeres coalesce into a much smaller number of regions near the nuclear periphery. (b, c) Confocal micrographs of yeast cells labeled with a telomere-specific hybridization probe (b) and a fluorescent-labeled antibody specific for SIR3 (c). Note that SIR3 is localized in the repressed telomeric heterochromatin. Similar experiments with RAP1, SIR2, and SIR4 have shown that these proteins also colocalize with the repressed telomeric heterochromatin. (d) Schematic model of the silencing mechanism at yeast telomeres. (Top left) Multiple copies of RAP1 bind to a simple repeated sequence at each telomere region that lacks nucleosomes. SIR3 and SIR4 bind to RAP1, and SIR2 binds to SIR4. SIR2 is a histone deacetylase that deacetylates the tails on the histones neighboring the repeated RAP1-binding site. (Middle) The hypoacetylated histone tails are also binding sites for SIR3 and SIR4, which in turn bind additional SIR2, deacetylating neighboring histones. Repetition of this process results in spreading of the region of hypoacetylated histones with associated SIR2, SIR3, and SIR4. (Bottom) Interactions between complexes of SIR2, SIR3, and SIR4 cause the chromatin to condense and several telomeres to associate, as shown in a–c. The higher-order chromatin structure generated sterically blocks other proteins from interacting with the underlying DNA. See M. Grunstein, 1997, Curr. Opin. Cell Biol. 9:383.