In vivo effects of histone H3 depletion on nucleosome occupancy and position in Saccharomyces cerevisiae.
In vivo effects of histone H3 depletion on nucleosome occupancy and position in Saccharomyces cerevisiae.
复制标题
组蛋白H3耗竭对酿酒酵母中核小体占用和位置的体内影响。
DOI：
10.1371/journal.pgen.1002771
复制
发表时间：
2012
期刊：
影响因子：
4.5
通讯作者：
Lieb JD
中科院分区：
文献类型：
--
作者：
Gossett AJ;Lieb JD
先前对酿酒酵母的研究证实，组蛋白H4的耗尽会导致数百个基因的全基因组转录去抑制。为了探索这种转录去抑制的机制，我们通过组蛋白H3转录的条件性抑制在体内耗尽核小体。然后，我们通过RNase-seq测量了转录的变化，并通过MNase-seq测量了染色质组织的变化。该实验还涉及反式作用因子和DNA编码元件在体内影响核小体位置和占据的程度。我们在全基因组范围内鉴定了60，000个核小体，并且我们将60，000个分类为在H3耗尽后优先减少占用，并且将60，000个分类为优先保留。我们发现，在体内的DNA序列的影响，有利于或不利于核小体占用增加组蛋白H3耗尽后，核小体密度有助于缓和的DNA序列对核小体形成在体内的影响。为了确定影响核小体占有率和位置的重要因素，我们将我们的数据与40个现有的全基因组数据集进行了比较。与启动子相关的因子，如组蛋白乙酰化和H2A.z掺入，在核小体丢失的位点富集。核小体保留与稳定标记如H3 K36 me 2有关。值得注意的是，染色质重塑Isw 2与保留的占用和改变的定位唯一相关，与Isw 2稳定组蛋白-DNA接触和使核小体在体内位于可用DNA的中心一致。RNA-seq揭示了比以前的研究更多数量的去抑制基因（约2，500个），并且这些基因在其启动子中表现出减少的核小体占用。总之，我们确定了在正常生长条件下可能影响核小体稳定性的因素以及它们作用的特定基因组位置。我们发现，DNA编码的核小体稳定性和染色质组成决定哪些核小体将在限制组蛋白的条件下丢失，这反过来又决定了哪些基因容易失去调控保真度。染色质是通过将146 bp的DNA包裹在一种称为组蛋白的盘状蛋白质复合物上形成的。这些蛋白质-DNA结构被称为核小体。核小体有助于调节基因转录，因为核小体与转录因子竞争DNA。已知核小体占据的精确定位和水平对于转录调控是至关重要的，但是调节核小体的位置和占据的机制尚未完全理解。近年来，DNA序列和染色质重塑蛋白在细胞凋亡中的作用成为研究热点。在这里，我们操纵细胞中组蛋白的浓度，以确定哪些核小体最容易受到占位和位置变化的影响。我们发现，染色质相关蛋白Sir 2和Tup 1，和染色质重塑Isw 2和Rsc 8，与稳定的核小体。组蛋白乙酰化和组蛋白变体H2A.z的掺入是与不稳定的核小体最高度相关的因素。某些DNA序列特性也有助于稳定性。这些数据确定了可能影响核小体稳定性的因素，并显示了染色质变化与组蛋白耗竭后转录变化之间的直接联系。
Previous studies in Saccharomyces cerevisiae established that depletion of histone H4 results in the genome-wide transcriptional de-repression of hundreds of genes. To probe the mechanism of this transcriptional de-repression, we depleted nucleosomes in vivo by conditional repression of histone H3 transcription. We then measured the resulting changes in transcription by RNA–seq and in chromatin organization by MNase–seq. This experiment also bears on the degree to which trans-acting factors and DNA–encoded elements affect nucleosome position and occupancy in vivo. We identified ∼60,000 nucleosomes genome wide, and we classified ∼2,000 as having preferentially reduced occupancy following H3 depletion and ∼350 as being preferentially retained. We found that the in vivo influence of DNA sequences that favor or disfavor nucleosome occupancy increases following histone H3 depletion, demonstrating that nucleosome density contributes to moderating the influence of DNA sequence on nucleosome formation in vivo. To identify factors important for influencing nucleosome occupancy and position, we compared our data to 40 existing whole-genome data sets. Factors associated with promoters, such as histone acetylation and H2A.z incorporation, were enriched at sites of nucleosome loss. Nucleosome retention was linked to stabilizing marks such as H3K36me2. Notably, the chromatin remodeler Isw2 was uniquely associated with retained occupancy and altered positioning, consistent with Isw2 stabilizing histone–DNA contacts and centering nucleosomes on available DNA in vivo. RNA–seq revealed a greater number of de-repressed genes (∼2,500) than previous studies, and these genes exhibited reduced nucleosome occupancy in their promoters. In summary, we identify factors likely to influence nucleosome stability under normal growth conditions and the specific genomic locations at which they act. We find that DNA–encoded nucleosome stability and chromatin composition dictate which nucleosomes will be lost under conditions of limiting histone protein and that this, in turn, governs which genes are susceptible to a loss of regulatory fidelity. Chromatin is formed by wrapping 146 bp of DNA around a disc-shaped complex of proteins called histones. These protein–DNA structures are known as nucleosomes. Nucleosomes help to regulate gene transcription, because nucleosomes compete with transcription factors for access to DNA. The precise positioning and level of nucleosome occupancy are known to be vital for transcriptional regulation, but the mechanisms that regulate the position and occupancy of nucleosomes are not fully understood. Recently, many studies have focused on the role of DNA sequence and chromatin remodeling proteins. Here, we manipulate the concentration of histone proteins in the cell to determine which nucleosomes are most susceptible to changes in occupancy and position. We find that the chromatin-associated proteins Sir2 and Tup1, and the chromatin remodelers Isw2 and Rsc8, are associated with stabilized nucleosomes. Histone acetylation and incorporation of the histone variant H2A.z are the factors most highly associated with destabilized nucleosomes. Certain DNA sequence properties also contribute to stability. The data identify factors likely to influence nucleosome stability and show a direct link between changes in chromatin and changes in transcription upon histone depletion.
登录
查看更多内容
影响因子：
12.3
作者：
Kaplan N;Hughes TR;Lieb JD;Widom J;Segal E
通讯作者：
Segal E
影响因子：
16
作者：
Badis, Gwenael;Chan, Esther T.;van Bakel, Harm;Pena-Castillo, Lourdes;Tillo, Desiree;Tsui, Kyle;Carlson, Clayton D.;Gossett, Andrea J.;Hasinoff, Michael J.;Warren, Christopher L.;Gebbia, Marinella;Talukder, Shaheynoor;Yang, Ally;Mnaimneh, Sanie;Terterov, Dimitri;Coburn, David;Yeo, Ai Li;Yeo, Zhen Xuan;Clarke, Neil D.;Lieb, Jason D.;Ansari, Aseem Z.;Nislow, Corey;Hughes, Timothy R.
通讯作者：
Hughes, Timothy R.
影响因子：
5.3
作者：
Gelbart, ME;Rechsteiner, T;Tsukiyama, T
通讯作者：
Tsukiyama, T
影响因子：
30.8
作者：
Buck, Michael J.;Lieb, Jason D.
通讯作者：
Lieb, Jason D.
影响因子：
11.4
作者：
HAN, M;KIM, UJ;GRUNSTEIN, M
通讯作者：
GRUNSTEIN, M