Genomic study of replication initiation in human chromosomes reveals the influence of transcription regulation and chromatin structure on origin selection.
Genomic study of replication initiation in human chromosomes reveals the influence of transcription regulation and chromatin structure on origin selection.
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DOI：
10.1091/mbc.e09-08-0707
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发表时间：
2010-02-01
影响因子：
3.3
通讯作者：
Dutta A
中科院分区：
文献类型：
--
作者：
Karnani N;Taylor CM;Malhotra A;Dutta A
后生动物的DNA复制始于称为起源的多个染色体位点。本研究确定了150个新的复制起源，通过两种新生链纯化方法得到证实。我们辨别转录起始和调控的作用，以及染色质特征在确定人类基因组的起源选择。后生动物的DNA复制始于称为起源的多个染色体位点。目前，纯化以起始点为中心的新生链有两种方法：lambda外切酶酶切法和抗溴脱氧尿苷免疫沉淀法。由于这两种方法都有其独特的优势和局限性，我们用这两种方法纯化了新生链，将它们独立杂交到平铺阵列（1%基因组），并比较了数据，以获得全基因组起源分布的准确视图。根据这一标准，我们确定了150个新的起源，这些起源在不同的方法中是可重复的。通过染色质免疫沉淀法检测这些起源的一个亚基，针对起源识别复合体（ORC）亚基2和3，发现93%的起始峰定位在ORC结合位点1 kb以内。起源与基因组功能元件的相关性显示，起源活性在转录起始位点（tss）周围显著富集。与接近tss一致，我们发现三分之一的起始事件发生在RNA聚合酶II结合位点或附近。有趣的是，约50%的早期起源活性位于转录调节因子结合区簇的5kb内。起源周围的染色质特征富集于组蛋白上的H3K4-（二甲基化和三甲基化）和H3乙酰化修饰。起源对开放染色质的亲和力也通过它们靠近dna酶i超敏感位点而得到重申。复制起始峰具有丰富的AT，并且有50%的起源定位于基因组的进化保守区域。综上所述，这些发现表明复制起始受到转录起始和调控以及染色质结构的影响。
DNA replication in metazoans initiates from multiple chromosomal loci called origins. This study identifies 150 new origins of replication that were confirmed by two methods of nascent strand purification. We discern the role of transcription initiation and regulation, as well as chromatin signatures in determining origin selection in human genome. DNA replication in metazoans initiates from multiple chromosomal loci called origins. Currently, there are two methods to purify origin-centered nascent strands: lambda exonuclease digestion and anti-bromodeoxyuridine immunoprecipitation. Because both methods have unique strengths and limitations, we purified nascent strands by both methods, hybridized them independently to tiling arrays (1% genome) and compared the data to have an accurate view of genome-wide origin distribution. By this criterion, we identified 150 new origins that were reproducible across the methods. Examination of a subset of these origins by chromatin immunoprecipitation against origin recognition complex (ORC) subunits 2 and 3 showed 93% of initiation peaks to localize at/within 1 kb of ORC binding sites. Correlation of origins with functional elements of the genome revealed origin activity to be significantly enriched around transcription start sites (TSSs). Consistent with proximity to TSSs, we found a third of initiation events to occur at or near the RNA polymerase II binding sites. Interestingly, ∼50% of the early origin activity was localized within 5 kb of transcription regulatory factor binding region clusters. The chromatin signatures around the origins were enriched in H3K4-(di- and tri)-methylation and H3 acetylation modifications on histones. Affinity of origins for open chromatin was also reiterated by their proximity to DNAse I-hypersensitive sites. Replication initiation peaks were AT rich, and >50% of the origins mapped to evolutionarily conserved regions of the genome. In summary, these findings indicate that replication initiation is influenced by transcription initiation and regulation as well as chromatin structure.
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影响因子：
11.4
作者：
Abdurashidova, G;Riva, S;Falaschi, A
通讯作者：
Falaschi, A
影响因子：
1.6
作者：
BIAMONTI, G;PERINI, G;FALASCHI, A
通讯作者：
FALASCHI, A
DOI：
10.1073/pnas.93.4.1498
发表时间：
1996-02-20
影响因子：
11.1
作者：
Dimitrova, DS;Giacca, M;Falaschi, A
通讯作者：
Falaschi, A
DOI：
10.1006/meth.1997.0529
发表时间：
1997-11-01
期刊：
METHODS-A COMPANION TO METHODS IN ENZYMOLOGY
影响因子：
--
作者：
Giacca, M;Pelizon, C;Falaschi, A
通讯作者：
Falaschi, A
影响因子：
4.8
作者：
Dhar, SK;Delmolino, L;Dutta, A
通讯作者：
Dutta, A