Cheutin Thierry, Gorski Stanislaw A, May Karen M, Singh Prim B, Misteli Tom
National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Mol Cell Biol. 2004 Apr;24(8):3157-67. doi: 10.1128/MCB.24.8.3157-3167.2004.
The mechanism for transcriptional silencing of pericentric heterochromatin is conserved from fission yeast to mammals. Silenced genome regions are marked by epigenetic methylation of histone H3, which serves as a binding site for structural heterochromatin proteins. In the fission yeast Schizosaccharomyces pombe, the major structural heterochromatin protein is Swi6. To gain insight into Swi6 function in vivo, we have studied its dynamics in the nucleus of living yeast. We demonstrate that, in contrast to mammalian cells, yeast heterochromatin domains undergo rapid, large-scale motions within the nucleus. Similar to the situation in mammalian cells, Swi6 does not permanently associate with these chromatin domains but binds only transiently to euchromatin and heterochromatin. Swi6 binding dynamics are dependent on growth status and on the silencing factors Clr4 and Rik1, but not Clr1, Clr2, or Clr3. By comparing the kinetics of mutant Swi6 proteins in swi6(-) and swi6(+) strains, we demonstrate that homotypic protein-protein interactions via the chromoshadow domain stabilize Swi6 binding to chromatin in vivo. Kinetic modeling allowed quantitative estimation of residence times and indicated the existence of at least two kinetically distinct populations of Swi6 in heterochromatin. The observed dynamics of Swi6 binding are consistent with a stochastic model of heterochromatin and indicate evolutionary conservation of heterochromatin protein binding properties from mammals to yeast.
从裂殖酵母到哺乳动物，着丝粒周围异染色质转录沉默的机制是保守的。沉默的基因组区域以组蛋白H3的表观遗传甲基化为标记，组蛋白H3作为结构异染色质蛋白的结合位点。在裂殖酵母粟酒裂殖酵母中，主要的结构异染色质蛋白是Swi6。为了深入了解Swi6在体内的功能，我们研究了其在活酵母细胞核中的动态变化。我们证明，与哺乳动物细胞不同，酵母异染色质结构域在细胞核内经历快速、大规模的运动。与哺乳动物细胞的情况类似，Swi6不会永久地与这些染色质结构域结合，而是仅短暂地结合常染色质和异染色质。Swi6的结合动态取决于生长状态以及沉默因子Clr4和Rik1，但不依赖于Clr1、Clr2或Clr3。通过比较swi6(-)和swi6(+)菌株中突变Swi6蛋白的动力学，我们证明通过染色体阴影结构域的同型蛋白质-蛋白质相互作用在体内稳定了Swi6与染色质的结合。动力学建模允许对停留时间进行定量估计，并表明在异染色质中至少存在两个动力学上不同的Swi6群体。观察到的Swi6结合动态与异染色质的随机模型一致，并表明从哺乳动物到酵母，异染色质蛋白结合特性具有进化保守性。
