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，不仅参与全球基因组结构，还参与基因调控的许多方面，后者包括直接的基因抑制/激活、增强子阻断和激素促进的沉默。CTCF含有一个高度保守的11个锌指DNA结合域，它介导DNA结合的序列特异性。正如我们发现的那样，CTCF通过两种主要机制发挥作用：一种是直接调节CTS下游的基因，另一种是通过形成染色质环来间接调节。染色质环由CTCF二聚体稳定，影响启动子、增强子和/或ICR之间的关系。DNA结合的CTCF的二聚化活性可能是其在大多数细胞类型中作为一种多功能的染色质桥联和染色质环化试剂的核心活性，其核心生物学功能。此外，CTCF的环形成活性可以自然扩展到形成局部的体细胞染色体间配对位点，从而获得表观遗传共同调节的可能性，如转录工厂、DNA复制工厂和DNA修复焦点。其他许多染色质锚定的功能，如印迹标记的建立和读取，X染色体的失活，以及细胞的凋亡都受到CTCF的调控。CTCF已经成为间期染色质3D组织的关键促进者，也是细胞增殖控制的主要参与者。在某些情况下，CTCF的环形成活性被发现伴随/补充了对特定基因的更直接的调节。这种混合模式的调控可能是天然基因调控框架的最合适的代表。我们还发现了一种新的CTCF活性，它直接将CTCF与CTCF与POL II的转录机制结合在一起。这一新的途径提供了一种机制，可以在启动子决定的+1位点下游(内含子/外显子序列)打开环独立的转录起始点，或者可能已经特异地进化到诱导整个基因组中的非编码转录本。从机制上讲，从DNA结合的CTCF复合体中调节Pol II的募集和随后的释放表明CTCF位点本身可以在基因组的某些位置起到衰减器和/或启动子的作用。虽然CTCF主要被认为是基因表达的调节因子，但我们对其在异染色质和中心体中的潜在功能以及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之前被证明经历了各种翻译后修饰，我们扩大了这些研究，以表征新的修饰。去调控的CTCF占据启动子靶点的另一个病理方面是癌症中异常的DNA甲基化。CTCF的这两个新的生物学作用都是MPS正在进行的研究课题。 我们先前首次分析了全基因组的CTCF靶标(Cell 2007，Vol.128，pp1231-1245)，通过CTS与人类基因组中基因位置的强烈关联，验证了CTCF在细胞功能中的基础作用。由于具有如此多的重要功能，CTCF成为脊椎动物的基本基因，因为CTCF基因敲除的小鼠是不能存活的(在非常早期的胚胎阶段就是致命的)。 在人类疾病方面，CTCF是一个候选的肿瘤抑制基因(TSG)；CTCF的11ZF DBD上的几个功能点突变已在原发癌中被鉴定，并结合CTCF基因的杂合性缺失。在过去的一年里，我们研究了几个基因座，以了解CTCFCTS对其调控的贡献。它们包括对免疫反应重要的基因，以及有潜力开发癌症治疗方法的基因。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，它往往起到激活作用，而位于(+1)下游的CTS通常起到抑制作用。人类端粒酶催化亚单位(HTERT)基因是本研究中最显著的基因之一。