Mekhail Karim, Seebacher Jan, Gygi Steven P, Moazed Danesh
Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Nature. 2008 Dec 4;456(7222):667-70. doi: 10.1038/nature07460. Epub 2008 Nov 9.
Repetitive DNA sequences, which constitute half the genome in some organisms, often undergo homologous recombination. This can instigate genomic instability resulting from a gain or loss of DNA. Assembly of DNA into silent chromatin is generally thought to serve as a mechanism ensuring repeat stability by limiting access to the recombination machinery. Consistent with this notion is the observation, in the budding yeast Saccharomyces cerevisiae, that stability of the highly repetitive ribosomal DNA (rDNA) sequences requires a Sir2-containing chromatin silencing complex that also inhibits transcription from foreign promoters and transposons inserted within the repeats by a process called rDNA silencing. Here we describe a protein network that stabilizes rDNA repeats of budding yeast by means of interactions between rDNA-associated silencing proteins and two proteins of the inner nuclear membrane (INM). Deletion of either the INM or silencing proteins reduces perinuclear rDNA positioning, disrupts the nucleolus-nucleoplasm boundary, induces the formation of recombination foci, and destabilizes the repeats. In addition, artificial targeting of rDNA repeats to the INM suppresses the instability observed in cells lacking an rDNA-associated silencing protein that is typically required for peripheral tethering of the repeats. Moreover, in contrast to Sir2 and its associated nucleolar factors, the INM proteins are not required for rDNA silencing, indicating that Sir2-dependent silencing is not sufficient to inhibit recombination within the rDNA locus. These findings demonstrate a role for INM proteins in the perinuclear localization of chromosomes and show that tethering to the nuclear periphery is required for the stability of rDNA repeats. The INM proteins studied here are conserved and have been implicated in chromosome organization in metazoans. Our results therefore reveal an ancient mechanism in which interactions between INM proteins and chromosomal proteins ensure genome stability.
重复DNA序列在某些生物体中占基因组的一半，常常发生同源重组。这会引发因DNA增减导致的基因组不稳定。DNA组装成沉默染色质通常被认为是一种通过限制重组机制的作用来确保重复序列稳定性的机制。与这一观点一致的是，在芽殖酵母酿酒酵母中观察到，高度重复的核糖体DNA（rDNA）序列的稳定性需要一个含Sir2的染色质沉默复合物，该复合物还通过一种称为rDNA沉默的过程抑制来自插入重复序列内的外源启动子和转座子的转录。在这里，我们描述了一个蛋白质网络，它通过rDNA相关沉默蛋白与内核膜（INM）的两种蛋白质之间的相互作用来稳定芽殖酵母的rDNA重复序列。删除INM或沉默蛋白会减少核周rDNA定位，破坏核仁 - 核质边界，诱导重组焦点的形成，并使重复序列不稳定。此外，将rDNA重复序列人工靶向到INM可抑制在缺乏通常用于重复序列外周拴系的rDNA相关沉默蛋白的细胞中观察到的不稳定性。而且，与Sir2及其相关的核仁因子不同，rDNA沉默不需要INM蛋白，这表明依赖Sir2的沉默不足以抑制rDNA基因座内的重组。这些发现证明了INM蛋白在染色体核周定位中的作用，并表明拴系到核周对于rDNA重复序列的稳定性是必需的。这里研究的INM蛋白是保守的，并且在多细胞动物的染色体组织中发挥作用。因此，我们的结果揭示了一种古老的机制，其中INM蛋白与染色体蛋白之间的相互作用确保了基因组稳定性。
