DNA Deformations and Interactions with Proteins

DNA 变形和与蛋白质的相互作用

基本信息

项目摘要

We report on several specific projects that are related to the DNA sequence-dependent structural characteristics, important for interactions with proteins (including bacterial gal-repressor and human p53), higher order self-organization of genomic DNA, and gene regulation. 1. DNA looping in prokaryotes Stabilization of the multi-subunit protein-DNA complexes is facilitated by DNA looping. One of the best characterized is the gal loop in E. coli, involved in regulation of the gal operon. To determine the optimal trajectory of the DNA loop in such a complex structure, we used the knowledge-based elastic model suitable for large-scale DNA simulations (developed by us earlier). As a result, we found that the "antiparallel" gal loop is energetically more favorable than the "parallel" one. The same trend was found for the DNA loop formed upon binding of the lac repressors to DNA. Based on these computations, we designed detailed experiments to visualize the 3D organization of the DNA loops in bacteria. The atomic force microscopy supports the "antiparallel" DNA looping, both with the gal and lac repressors. These results imply that the "antiparallel" DNA looping may be a general feature of the condensed bacterial nucleoid, as opposed to the parallel DNA "wrapping" around histones in eukaryotic chromatin. Importantly, the regular DNA folding in prokaryotes is consistent with the periodic distribution of the curved A-tracts in bacterial genomes, described below. 2. Distribution of A- and G-tracts and DNA packaging in pro- and eukaryotes Periodic positioning of the A- and G-tracts in DNA causes DNA curvature in solution and facilitates its bending in the complexes with proteins. Here, we analyzed distribution of these sequences in the pro- and eukaryotic genomes. We found that distribution of the strongly bent A-tracts (4-7 bp) in the prokaryotic genomes demonstrates a remarkable periodicity of 10-11 bp. Such a periodicity may reflect the intrinsic propensity of prokaryotic DNA to form the loop-shaped structures. Based on these data and by analogy with the "gene repression" gal- and lac-loops in E. coli, we hypothesize that the loop folds with the structural period of 100 bp may be elementary units of the prokaryotic nucleoid packaging. This hypothesis was tested by the micrococcal nuclease digestion of bacterial nucleoids (in collaboration with S. Adhya, NCI). The results show that the 100 bp DNA fragments are highly overrepresented in digestion products, thereby implying a highly specific nucleoid packaging, with the DNA structural period of 100 bp. On the other hand, the G-tracts are underrepresented in prokaryotic genomes. In contrast, both the A- and G-tracts of all lengths are highly overrepresented in eukaryotic genomes. However, the "optimal" A-tracts (4-7 bp) do not reveal the 10-11 bp periodicity. Apparently, the intrinsic curvature of DNA, caused by the A-tracts, is not a necessary prerequisite for the formation of nucleosomes. Rather, the overabundant long purine (A and G) runs observed in eukaryotic genomes may serve as the "chromatin organizers," decreasing the DNA propensity for the formation of nucleosomes, especially in the promoter regions. We are currently studying the G-tract clustering in the CpG islands, in particular in the vicinity of the cancer-related genes' promoters. 3. Genome-wide distribution of p53 sites in human DNA The tetrameric p53 binding to DNA plays a key role in tumor suppression. In response to DNA damage and other types of cellular stress, the p53 protein becomes activated and binds DNA sequence-specifically, functioning as a transcriptional factor or cell cycle regulator. p53 is unique in regulating a wide spectrum of genes: thousands of human genes are either activated or repressed by p53. Normally, the p53 tetramer binds to DNA response elements, consisting of two decamers RRRCWWGYYY (half-sites) separated by a spacer. (The length of the spacer, S, varies from 0 to 14 bp in the known functional binding sites, but in most cases S=0 or 1.) How many putative p53 binding sites, consistent with this scheme, are there in the human genome? What is the distribution of the spacer lengths? With the human genome sequence we can directly answer these questions. The distribution of spacers proves to be extremely nonuniform in all human chromosomes, with strong peaks in the profile, exceeding the average background 3-4 fold. The peaks at S=0 and 10 bp, and the gap at 4-5 bp are consistent with our earlier computer modeling and electrophoresis measurements, indicating the lateral positioning of the p53 core domains on the outer side of the DNA loop. In general, these data agree with the idea that the p53 tetramer can bind DNA specifically without unwrapping nucleosomes in the course of transcriptional activation of the chromatin-assembled genes. Currently, we are exploring localization of the putative p53 sites with respect to the starts of transcription. Our data indicate strong difference between the up- and down-regulated genes in terms of distribution of the p53 sites in the vicinity of genes. The up-regulated genes are characterized by a twofold higher occurrence of the p53 sites within 1 Kbp from the start of transcription, compared to the down-regulated genes. In addition, the down-regulated genes reveal a higher fraction of the p53 sites with "unusual" spacer S=3, shown earlier to repress transcription of the corresponding genes. These are extremely important observations, as they can be used for prediction of the p53-activated and repressed genes. Summarizing, distribution of p53 sites in the human genome reflects the versatility of p53 binding and its tumor suppressor functions.
我们报告了几个与DNA序列依赖的结构特征有关的特定项目,这些特征对于与蛋白质(包括细菌GAL抑制子和人类P53)的相互作用、基因组DNA的高阶自组织和基因调控非常重要。 1.原核生物中的DNA环 DNA环有助于多亚基蛋白质-DNA复合体的稳定。其中最具特点的是大肠杆菌中的GAL环,参与GAL操纵子的调节。为了确定在这样一个复杂的结构中DNA环的最佳轨迹,我们使用了适合于大规模DNA模拟的基于知识的弹性模型(我们之前开发的)。结果,我们发现“反平行”的GAL环在能量上比“平行”的GAL环更有利。同样的趋势也被发现在乳胶抑制物与DNA结合时形成的DNA环。 在这些计算的基础上,我们设计了详细的实验来可视化细菌中DNA环的3D组织。原子力显微镜支持带有半乳糖和乳胶抑制物的“反平行”DNA环路。 这些结果表明,“反平行”的DNA环可能是浓缩细菌类核的一个普遍特征,而不是真核细胞染色质中平行的DNA“包裹”在组蛋白周围。重要的是,原核生物中规则的DNA折叠与细菌基因组中弯曲的A区的周期分布一致,如下所述。 2.原核生物和真核生物中A-和G-链的分布和DNA包装 DNA中A-和G-链的周期性定位导致了DNA在溶液中的弯曲,并促进了它在与蛋白质的复合体中的弯曲。在这里,我们分析了这些序列在原核基因组和真核基因组中的分布。 我们发现,在原核生物基因组中,强弯曲的A-链(4-7个碱基)的分布呈现出10-11个碱基的显著周期性。这种周期性可能反映了原核生物DNA形成环状结构的内在倾向。根据这些数据,并通过与大肠杆菌中的“基因抑制”半乳糖环的类比,我们推测该环的结构周期为100bp,可能是原核类核包装的基本单位。这一假说通过微球菌核酸酶消化细菌类核得到了验证(与S.Adhya,NCI合作)。结果表明,100bp的DNA片段在消化产物中高度表达,这意味着具有高度特异性的类核包装,其DNA结构周期为100bp。另一方面,G-链在原核生物基因组中的表达不足。 相反,所有长度的A-和G-道在真核基因组中都有高度的过度表达。然而,“最佳”A波道(4-7BP)并不显示10-11BP的周期性。显然,由A束引起的DNA的固有曲率并不是形成核小体的必要条件。相反,在真核基因组中观察到的过多的长嘌呤(A和G)可能是“染色质组织者”,降低了DNA形成核小体的倾向,特别是在启动子区域。我们目前正在研究CpG岛上的G-Track聚集性,特别是在癌症相关基因启动子附近。 3.人类DNA中P53位点的全基因组分布 四聚体P53与DNA的结合在肿瘤抑制中起着关键作用。在DNA损伤和其他类型的细胞应激反应中,P53蛋白被激活并与DNA序列结合--具体地说,作为转录因子或细胞周期调节因子发挥作用。P53在调控广泛的基因方面是独一无二的:成千上万的人类基因要么被P53激活,要么被抑制。正常情况下,P53四聚体与DNA反应元件结合,由由间隔物隔开的两个十聚体RRRCWWGYYY(半个位点)组成。(在已知的功能结合位点上,间隔区S的长度从0到14bp不等,但在大多数情况下,S=0或1。)人类基因组中有多少个与这一方案一致的假定的P53结合位点?间隔物长度的分布是怎样的? 利用人类基因组序列,我们可以直接回答这些问题。间隔区在所有人类染色体中的分布极不均匀,图谱中有强烈的峰,超过平均本底3-4倍。S=0和10bp处的峰以及4-5bp处的间隙与我们早期的计算机模拟和电泳法测量结果一致,表明P53核心区在DNA环的外侧定位。总体而言,这些数据与P53四聚体可以在染色质组装基因转录激活过程中不解开核小体而特异性地结合DNA的观点一致。 目前,我们正在探索与转录起始相关的假定的P53位点的定位。我们的数据表明,在基因附近的P53位点的分布方面,上调和下调基因之间存在着显著的差异。与表达下调的基因相比,上调的基因的特征是从转录开始的1kBP内,p53位点的出现频率要高出两倍。此外,下调的基因揭示了较高比例的P53位点,其“不寻常”的间隔区S=3,前面显示了抑制相应基因转录的作用。这些是极其重要的观察,因为它们可以用来预测P53激活和抑制的基因。 综上所述,人类基因组中P53位点的分布反映了P53结合及其肿瘤抑制功能的多功能性。

项目成果

期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)

数据更新时间:{{ journalArticles.updateTime }}

{{ item.title }}
{{ item.translation_title }}
  • DOI:
    {{ item.doi }}
  • 发表时间:
    {{ item.publish_year }}
  • 期刊:
  • 影响因子:
    {{ item.factor }}
  • 作者:
    {{ item.authors }}
  • 通讯作者:
    {{ item.author }}

数据更新时间:{{ journalArticles.updateTime }}

{{ item.title }}
  • 作者:
    {{ item.author }}

数据更新时间:{{ monograph.updateTime }}

{{ item.title }}
  • 作者:
    {{ item.author }}

数据更新时间:{{ sciAawards.updateTime }}

{{ item.title }}
  • 作者:
    {{ item.author }}

数据更新时间:{{ conferencePapers.updateTime }}

{{ item.title }}
  • 作者:
    {{ item.author }}

数据更新时间:{{ patent.updateTime }}

ROBERT L JERNIGAN其他文献

ROBERT L JERNIGAN的其他文献

{{ item.title }}
{{ item.translation_title }}
  • DOI:
    {{ item.doi }}
  • 发表时间:
    {{ item.publish_year }}
  • 期刊:
  • 影响因子:
    {{ item.factor }}
  • 作者:
    {{ item.authors }}
  • 通讯作者:
    {{ item.author }}

{{ truncateString('ROBERT L JERNIGAN', 18)}}的其他基金

Novel Use of Genome Information to Understand Mutations
利用基因组信息来理解突变的新方法
  • 批准号:
    10488281
  • 财政年份:
    2021
  • 资助金额:
    --
  • 项目类别:
Novel Use of Genome Information to Understand Mutations
利用基因组信息来理解突变的新方法
  • 批准号:
    10303852
  • 财政年份:
    2021
  • 资助金额:
    --
  • 项目类别:
Novel Use of Genome Information to Understand Mutations
利用基因组信息来理解突变的新方法
  • 批准号:
    10661834
  • 财政年份:
    2021
  • 资助金额:
    --
  • 项目类别:
Modeling Ribosomal Control, Function and Assembly
核糖体控制、功能和组装建模
  • 批准号:
    7290378
  • 财政年份:
    2006
  • 资助金额:
    --
  • 项目类别:
Modeling Ribosomal Control, Function and Assembly
核糖体控制、功能和组装建模
  • 批准号:
    7486144
  • 财政年份:
    2006
  • 资助金额:
    --
  • 项目类别:
Modeling Ribosomal Control, Function and Assembly
核糖体控制、功能和组装建模
  • 批准号:
    7681539
  • 财政年份:
    2006
  • 资助金额:
    --
  • 项目类别:
Modeling Ribosomal Control, Function and Assembly
核糖体控制、功能和组装建模
  • 批准号:
    7149659
  • 财政年份:
    2006
  • 资助金额:
    --
  • 项目类别:
Coarse-Grained Models of Proteins
蛋白质的粗粒度模型
  • 批准号:
    6914431
  • 财政年份:
    2004
  • 资助金额:
    --
  • 项目类别:
Coarse-Grained Models of Proteins
蛋白质的粗粒度模型
  • 批准号:
    6829176
  • 财政年份:
    2004
  • 资助金额:
    --
  • 项目类别:
Coarse-Grained Models of Proteins
蛋白质的粗粒度模型
  • 批准号:
    8209105
  • 财政年份:
    2004
  • 资助金额:
    --
  • 项目类别:

相似海外基金

Targeting pathogenic TAR DNA-binding protein 43 to treat frontotemporal dementia and motor neuron disease
靶向致病性 TAR DNA 结合蛋白 43 治疗额颞叶痴呆和运动神经元疾病
  • 批准号:
    nhmrc : 2001572
  • 财政年份:
    2021
  • 资助金额:
    --
  • 项目类别:
    Ideas Grants
Electron microscopic analysis of a G4 DNA-binding protein Rif1, a key organizer of chromosomal domains
G4 DNA 结合蛋白 Rif1(染色体结构域的关键组织者)的电子显微镜分析
  • 批准号:
    18K06102
  • 财政年份:
    2018
  • 资助金额:
    --
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Functional analysis of methylated DNA-binding protein CIBZ in mouse embryogenesis
甲基化DNA结合蛋白CIBZ在小鼠胚胎发生中的功能分析
  • 批准号:
    16K08587
  • 财政年份:
    2016
  • 资助金额:
    --
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Continuous directed evolution of a light-controlled DNA-binding protein
光控DNA结合蛋白的连续定向进化
  • 批准号:
    437922-2013
  • 财政年份:
    2015
  • 资助金额:
    --
  • 项目类别:
    Postgraduate Scholarships - Doctoral
Function and evolution of mitochondrial DNA-binding protein in the fission yeast
裂殖酵母线粒体DNA结合蛋白的功能和进化
  • 批准号:
    15K07168
  • 财政年份:
    2015
  • 资助金额:
    --
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Development of a photo-controlled DNA-binding protein
光控 DNA 结合蛋白的开发
  • 批准号:
    459937-2014
  • 财政年份:
    2015
  • 资助金额:
    --
  • 项目类别:
    Alexander Graham Bell Canada Graduate Scholarships - Doctoral
Functional analysis of the single-stranded DNA-binding protein FUBP1 as a transcriptional regulator of hematopoietic stem cell self-renewal
单链DNA结合蛋白FUBP1作为造血干细胞自我更新转录调节因子的功能分析
  • 批准号:
    276833671
  • 财政年份:
    2015
  • 资助金额:
    --
  • 项目类别:
    Research Grants
Continuous directed evolution of a light-controlled DNA-binding protein
光控DNA结合蛋白的连续定向进化
  • 批准号:
    437922-2013
  • 财政年份:
    2014
  • 资助金额:
    --
  • 项目类别:
    Postgraduate Scholarships - Doctoral
Structural ans functional analysis of single-stranded DNA-binding protein DdrA
单链 DNA 结合蛋白 DdrA 的结构和功能分析
  • 批准号:
    26506030
  • 财政年份:
    2014
  • 资助金额:
    --
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Development of a photo-controlled DNA-binding protein
光控 DNA 结合蛋白的开发
  • 批准号:
    459937-2014
  • 财政年份:
    2014
  • 资助金额:
    --
  • 项目类别:
    Alexander Graham Bell Canada Graduate Scholarships - Doctoral
{{ showInfoDetail.title }}

作者:{{ showInfoDetail.author }}

知道了