Normal and Pathologic Functions of CTCF and Its Distinct Classes of DNA-targets

CTCF 的正常和病理功能及其不同类型的 DNA 靶标

• 批准号: 8336142
• 负责人: Victor Lobanenkov
• 金额: $ 86.88万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8336142
• 关键词: Aberrant DNA Methylation; Affect; Apoptosis; Architecture; Binding; Binding Sites; Biological; Biological Process; Biology; CCCTC-binding factor; Catalytic Domain; Cell Proliferation; Cell physiology; Cells; Centrosome; Chromatin; Chromatin Loop; Chromosomal Stability; Chromosome Pairing; Chromosome Segregation; Chromosome Structures; Complement; Complex; DNA; DNA Binding; DNA Binding Domain; DNA Polymerase II; DNA Repair; DNA biosynthesis; Data; Development; Dimerization; Enhancers; Epigenetic Process; Essential Genes; Exons; Fingers; Functional RNA; Gene Expression; Gene Expression Regulation; Gene Order; Gene Targeting; Genes; Genetic Transcription; Genome; Goals; Heterochromatin; Hormones; Housekeeping; Human; Human Genome; Immune response; Individual; Interphase; Introns; Knockout Mice; Link; Location; Malignant Neoplasms; Mediating; Meiosis; Mitosis; Mitotic Chromosome; Modification; Nuclear; Pathologic; Pathway interactions; Play; Point Mutation; Post-Translational Protein Processing; Reading; Regulation; Regulator Genes; Role; Site; Specificity; Telomerase; Time; Transcript; Transcription Initiation Site; Transcriptional Regulation; Tumor Suppressor Genes; Vertebrates; Work; X Inactivation; cancer therapy; cell transformation; cell type; early embryonic stage; gene repression; genome-wide; human disease; imprint; novel; promoter; tumor

CTCF is a highly conserved, multi-functional nuclear factor that involved both in global genome architecture and in many aspects of gene regulation, the latter ranging from the direct gene repression/activation to enhancer blocking and hormone-facilitated silencing. CTCF contains a highly conserved 11 Zn-finger DNA-binding domain that mediates sequence-specificity of DNA binding. As we discovered, CTCF functions through two major mechanisms: either direct regulation of a gene downstream of CTSes or indirect regulation via the formation of chromatin loops. The chromatin loops, stabilized by CTCF dimerization, affect relationships among promoters, enhancers and/or ICRs. Dimerization activity of DNA-bound CTCF may potentially be at the core of its activity as a versatile chromatin-bridging and chromatin-looping agent in most cell types, underlying its core biological functions. Furthermore, the loop-forming activity of CTCF can be naturally extended to formation of localized somatic inter-chromosome pairing sites that therefore acquire potential for epigenetic co-regulation such as transcription factories, DNA replication factories, and DNA repair foci. Many other chromatin-anchored functions, such as the establishment of imprinting marks and their reading, X-chromosome inactivation, and apoptosis are regulated by CTCF. CTCF has emerged as a key facilitator of 3D organization of interphase chromatin, as well as a major player in cell proliferation control. In some cases, the loop-forming activity of CTCF was found to be accompanied/complemented by the more direct regulation of a particular gene. This mixed mode regulation is likely the most appropriate representation of a native gene regulation framework. We also identified a novel CTCF activity that directly links CTCF to transcriptional machinery binding of CTCF to Pol II. This novel pathway provides either a mechanism for opening loop-independent transcription start sites downstream of the promoter-determined +1 site (at intron/exon sequences) or may have specifically evolved to induce non-coding transcripts throughout the genome. Mechanistically, the regulated recruitment and the subsequent release of Pol II from a DNA-bound CTCF complex indicates that the CTCF site itself could act as an attenuator and/or promoter in some locations in the genome. While CTCF is mostly known as a regulator of gene expression, our data on its potential functions in heterochromatin and centrosomes, as well as its roles in mitosis and meiosis, suggested a significant housekeeping role of CTCF in genome organization and chromosome segregation. CTCF was previously shown to undergo a variety of posttranslational modifications, and we expanded these studies to characterize novel modifications. Another pathological aspect of the deregulated CTCF occupancy of promoter targets sites is aberrant DNA methylation in cancers. Both of these novel biological roles of CTCF are subjects of ongoing studies in the MPS. We previously analyzed genome-wide CTCF targets for the first time (Cell 2007, vol. 128, pp1231-1245), and the fundamental roles of CTCF in cellular functions were validated by a strong correlation of CTSs with gene positions in human genome. By virtue of having so many vital functions CTCF became an essential gene in vertebrates, as CTCF-knockout mice are non-viable (lethality at the very early embryonic stages). With respect to human disease, CTCF is a candidate tumor suppressor gene (TSG); several functional point mutations in the 11ZF DBD of CTCF have been characterized in primary cancers, in combination with the LOH of the CTCF locus. In the past year, we studied several loci in order to understand contributions of CTCF CTSes to their regulation. They included genes important for immune responses, as well as genes with a potential for the development of approaches for cancer treatment. As a general rule, we have found that if a CTCF binding site is located upstream of the transcriptional start site, it tends to play an activator role, while CTSes located downstream of (+1) usually behave as repressors. The gene for catalytic subunit of human telomerase (hTERT) is one of the most prominent genes in this study.

CTCF是一种高度保守的多功能核因子，涉及全球基因组结构和基因调控的许多方面，后者的范围从直接基因抑制/激活到增强子阻断和激素促进的沉默。 CTCF 包含高度保守的 11 锌指 DNA 结合结构域，可介导 DNA 结合的序列特异性。正如我们发现的，CTCF 通过两种主要机制发挥作用：直接调节 CTS 下游基因或通过形成染色质环进行间接调节。通过 CTCF 二聚化稳定的染色质环影响启动子、增强子和/或 ICR 之间的关系。 DNA 结合 CTCF 的二聚活性可能是其在大多数细胞类型中作为多功能染色质桥接剂和染色质环化剂的活性的核心，也是其核心生物学功能的基础。此外，CTCF的环形成活性可以自然地扩展到局部体细胞染色体间配对位点的形成，从而获得表观遗传共调节的潜力，例如转录工厂、DNA复制工厂和DNA修复灶。许多其他染色质锚定功能，例如印记标记的建立及其读取、X 染色体失活和细胞凋亡，均受 CTCF 调节。 CTCF 已成为间期染色质 3D 组织的关键促进者，以及细胞增殖控制的主要参与者。在某些情况下，发现 CTCF 的环形成活性伴随着/补充了对特定基因的更直接的调节。这种混合模式调节可能是天然基因调节框架的最合适的代表。我们还发现了一种新的 CTCF 活性，该活性直接将 CTCF 与 CTCF 与 Pol II 的转录机制结合起来。这种新颖的途径提供了一种在启动子确定的+1位点（内含子/外显子序列）下游打开环独立转录起始位点的机制，或者可能已经专门进化以诱导整个基因组中的非编码转录本。从机制上讲，Pol II 从 DNA 结合的 CTCF 复合物中受调控的募集和随后的释放表明 CTCF 位点本身可以​​在基因组中的某些位置充当衰减子和/或启动子。虽然 CTCF 主要被认为是基因表达的调节因子，但我们关于其在异染色质和中心体中的潜在功能以及其在有丝分裂和减数分裂中的作用的数据表明 CTCF 在基因组组织和染色体分离中具有重要的管家作用。 CTCF 之前被证明会经历各种翻译后修饰，我们扩展了这些研究以表征新的修饰。 CTCF 启动子靶位点占用失调的另一个病理学方面是癌症中的异常 DNA 甲基化。 CTCF 的这两种新的生物学作用都是 MPS 正在进行的研究主题。 我们之前首次分析了全基因组 CTCF 靶标（Cell 2007，第 128 卷，第 1231-1245 页），并且通过 CTS 与人类基因组中基因位置的强相关性验证了 CTCF 在细胞功能中的基本作用。由于 CTCF 具有如此多的重要功能，CTCF 成为脊椎动物的必需基因，因为 CTCF 敲除小鼠无法存活（在胚胎早期阶段即具有致死性）。 就人类疾病而言，CTCF是候选抑癌基因（TSG）； CTCF 的 11ZF DBD 中的几个功能性点突变与 CTCF 基因座的 LOH 相结合，已在原发性癌症中得到了表征。在过去的一年中，我们研究了几个位点，以了解 CTCF CTS 对其调控的贡献。它们包括对免疫反应重要的基因，以及具有开发癌症治疗方法潜力的基因。作为一般规则，我们发现如果 CTCF 结合位点位于转录起始位点的上游，它往往会发挥激活子的作用，而位于 (+1) 下游的 CTS 通常充当阻遏子。人端粒酶催化亚基（hTERT）基因是本研究中最重要的基因之一。