Absolute nucleosome occupancy map for the Saccharomyces cerevisiae genome [ODM-seq]

Mapping of nucleosomes, the basic DNA packaging unit in eukaryotes, is fundamental for understanding genome regulation as nucleosomes modulate DNA access by their positioning along the genome. A cell population nucleosome map requires two observables: nucleosome positions along the DNA (“Where?”) and nucleosome occupancies across the population (“In how many cells?”). All available genome-wide nucleosome mapping techniques are yield methods as they score either nucleosomal (e.g., MNase-seq, chemical cleavage-seq) or non-nucleosomal (e.g., ATAC-seq) DNA but lose track of the total DNA population for each genomic region. Therefore, they only provide nucleosome positions and maybe compare relative occupancies between positions but cannot measure absolute nucleosome occupancy, which is the fraction of all DNA molecules occupied at a given position and time by a nucleosome. Here, we established two orthogonal and thereby crossvalidating approaches to measure absolute nucleosome occupancy across the Saccharomyces cerevisiae genome via restriction enzymes and DNA methyltransferases. The resulting high-resolution (9 bp) map shows uniform absolute occupancies. Most nucleosome positions are occupied in most cells: 97% of all nucleosomes called by chemical cleavage-seq have a mean absolute occupancy of 90 ± 6% (± SD). Depending on nucleosome position calling procedures, there are 57-60,000 nucleosomes per yeast cell. The few low absolute occupancy nucleosomes do not correlate with highly transcribed gene bodies, but with increased presence of the nucleosome-evicting RSC chromatin remodeling complex there and are enriched upstream of highly transcribed or regulated genes. Our work provides a quantitative method and reference frame in absolute terms for future chromatin studies. Chromatin occupancy measurements by ODM-seq (Occupancy via DNA methylation) and ORE-seq (Occupancy via Restriction Enzymes) of several S. cerevisiae chromatin replicates

核小体作为真核生物的核心DNA包装单元，其基因组定位图谱是解析基因组调控机制的基础：核小体通过沿基因组的位置排布，直接调控DNA的可及性。基于细胞群体的核小体图谱需要两类核心观测指标：一是核小体沿DNA分子的具体位置（即“定位在哪里？”），二是核小体在细胞群体中的占据比例（即“在多少比例的细胞中存在？”）。现有全基因组范围的核小体定位技术均属于产量检测法：它们仅能检测核小体结合DNA（例如微球菌核酸酶测序（MNase-seq）、化学切割测序（chemical cleavage-seq））或非核小体结合DNA（例如转座酶可及性测序（ATAC-seq）），但无法追踪每个基因组区域的总DNA分子群体。因此，此类技术仅能获取核小体位置信息，或仅可比较不同位置间的相对占据比例，却无法测量绝对核小体占据率——即特定基因组位置与时刻下，被核小体占据的DNA分子占该区域总DNA分子的比例。本研究建立了两种正交且可相互验证的方法，通过限制性内切酶与DNA甲基转移酶，实现对酿酒酵母（Saccharomyces cerevisiae）全基因组范围内绝对核小体占据率的定量检测。最终获得的高分辨率（9碱基对，bp）核小体定位图谱显示，核小体绝对占据率整体较为均一。绝大多数核小体位置在多数细胞中均被占据：通过化学切割测序（chemical cleavage-seq）鉴定的全部核小体中，97%的平均绝对占据率为90%±6%（±标准差，SD）。根据不同的核小体位置鉴定流程，每个酵母细胞的核小体数量约为57000~60000个。少数绝对占据率较低的核小体，并未与高转录活性的基因编码区呈现相关性，反而与该区域中核小体驱逐复合物RSC染色质重塑复合物的富集程度呈正相关，且在高转录活性或受调控基因的上游区域显著富集。本研究为未来的染色质研究提供了定量检测方法与绝对定量参考框架。本研究通过基于DNA甲基化的占据率测序（ODM-seq，Occupancy via DNA methylation）与基于限制性内切酶的占据率测序（ORE-seq，Occupancy via Restriction Enzymes），对多组酿酒酵母染色质重复样本进行了占据率检测。