Insight Into the Mechanism of Nucleosome Reorganization From Histone Mutants That Suppress Defects in the FACT Histone Chaperone
Insight Into the Mechanism of Nucleosome Reorganization From Histone Mutants That Suppress Defects in the FACT Histone Chaperone
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DOI：
10.1534/genetics.111.128769
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发表时间：
2011-08-01
期刊：
影响因子：
3.3
通讯作者：
Formosa, Tim
中科院分区：
文献类型：
--
作者：
McCullough, Laura;Rawlins, Robert;Formosa, Tim
FACT（FACilitates Chromatin Transcription/Transactions）通过建立和克服染色质的抑制特性在真核生物的转录和复制中起着核心作用。FACT通过将核小体在标准形式和更开放的重组形式之间相互转换来促进这些相反的目标。在正向，重组使核小体不稳定，而逆反应促进核小体组装。核小体去稳定化涉及破坏组蛋白H2 A-H2 B二聚体、（H3-H4）（2）四聚体和DNA之间的接触。在这里，我们表明，突变，削弱核小体中的二聚体：四聚体界面抑制缺陷所造成的FACT缺陷在体内的酵母酿酒酵母。突变的基因编码的Spt 16亚基的FACT的原因表型与缺陷的转录和复制，我们确定组蛋白突变体，选择性地抑制那些与复制。纯化组分的分析表明，FACT的缺陷版本是无法有效地维持重组的核小体状态，而突变组蛋白的核小体重组比正常更容易。因此，当FACT缺陷与组蛋白缺陷组合时观察到的遗传抑制揭示了核小体内接触的动态重组对FACT体内功能的重要性，特别是对FACT在促进DNA复制复合物进展中的明显作用。我们还表明，H2 B突变导致不同的表型，这取决于编码这种蛋白质的两个相似基因中的哪一个被改变，揭示了这些重复基因之间意想不到的功能差异，并质疑通过从单个质粒携带的等位基因表达来检查组蛋白突变体的影响的做法。
FACT (FAcilitates Chromatin Transcription/Transactions) plays a central role in transcription and replication in eukaryotes by both establishing and overcoming the repressive properties of chromatin. FACT promotes these opposing goals by interconverting nucleosomes between the canonical form and a more open reorganized form. In the forward direction, reorganization destabilizes nucleosomes, while the reverse reaction promotes nucleosome assembly. Nucleosome destabilization involves disrupting contacts among histone H2A-H2B dimers, (H3-H4)(2) tetramers, and DNA. Here we show that mutations that weaken the dimer: tetramer interface in nucleosomes suppress defects caused by FACT deficiency in vivo in the yeast Saccharomyces cerevisiae. Mutating the gene that encodes the Spt16 subunit of FACT causes phenotypes associated with defects in transcription and replication, and we identify histone mutants that selectively suppress those associated with replication. Analysis of purified components suggests that the defective version of FACT is unable to maintain the reorganized nucleosome state efficiently, whereas nucleosomes with mutant histones are reorganized more easily than normal. The genetic suppression observed when the FACT defect is combined with the histone defect therefore reveals the importance of the dynamic reorganization of contacts within nucleosomes to the function of FACT in vivo, especially to FACT's apparent role in promoting progression of DNA replication complexes. We also show that an H2B mutation causes different phenotypes, depending on which of the two similar genes that encode this protein are altered, revealing unexpected functional differences between these duplicated genes and calling into question the practice of examining the effects of histone mutants by expressing them from a single plasmid-borne allele.