Insulator function and CTCF

我们一直对蛋白质CTCF感兴趣，几年前我们首次确定它具有绝缘体的特性，当放在增强剂和启动子之间时，它可以阻止增强剂和启动子之间的作用。我们证明了这种活性在调节原发源亲本的等位基因特异性基因在Igf2/H19印迹基因座上的表达中起着重要作用。近年来其他实验室的工作表明，CTCF的一个主要作用模式是招募粘附素，粘附素反过来稳定DNA上CTCF结合位点之间的相互作用，导致环状结构域的形成。根据相互作用的几何形状，这种环可以排除导致绝缘的增强子，或者使增强子和启动子更紧密地结合在一起，导致激活。 细胞核内的DNA被包装成染色质，并进一步组织成分隔活跃和不活跃基因组区域的拓扑相关区域(TADS)。TADS的建立和维持需要蛋白质CTCF，我们有兴趣识别和研究CTCF与被招募用于绝缘体功能的蛋白质和核酸伙伴的相互作用。我们已经证明，CTCF与DNA结合的11个锌指两侧的N-末端和C-末端结构域似乎本质上是无序的，这在一定程度上解释了其他研究中发现的大量CTCF结合伙伴。目前的工作集中在进一步表征这些结构域的物理性质，识别与高亲和力结合的伙伴，并研究形成的复合体。 我们参与了一项合作，研究SNP在与重大精神疾病相关的基因表达中影响CTCF结合强度的可能作用。我们还开始了对核仁内非核糖体基因组织的研究，这项研究既涉及基因组内容，也涉及核仁基因组的组织。 作为我们对大规模基因组组织感兴趣的一部分，我们对转录因子MAZ进行了研究，该转录因子经常结合在CTCF位点旁边。我们观察到这两种蛋白质之间的相互作用以及MAZ对CTCF性质的影响。 作为我们对基因组组织感兴趣的一部分，我们对多个细胞系中核仁的基因组含量进行了比较研究。这解决了核仁中存在哪些非核糖体基因的问题，目的是确定决定这种占有率的属性。结果表明，不同细胞类型之间的基因同一性存在很大差异。 所有这些结果都与染色质结构、组蛋白修饰和基因组的长程组织在细胞功能中的作用有关，并反过来与正常和异常的细胞代谢和细胞分裂问题有关。

We have been interested in the protein CTCF, which we first identified some years ago as having properties of an insulator, blocking interaction between enhancers and promoters when placed between them. We demonstrated that this activity plays an important role in regulating parent of origin allele-specific gene expression at the Igf2/H19 imprinted locus. Work in other laboratories in recent years has shown that a principal mode of action of CTCF is to recruit cohesin which in turn stabilizes interactions between CTCF binding sites on DNA, leading to formation of loop domains. Depending on the geometry of the interactions such loops can either exclude an enhancer leading to insulation, or bring enhancer and promoter closer together, leading to activation. DNA within the cell nucleus is packaged into chromatin, and further organized into topologically associated domains (TADs) separating active and inactive genomic regions. The establishment and maintenance of TADs requires the protein CTCF, and we are interested in identifying and studying the interactions of CTCF with the protein and nucleic acid partners recruited for insulator function. We have shown that the N- and C-terminal domains that flank the DNA binding 11 zinc fingers of CTCF appear to be intrinsically disordered explaining, in part, the large number of CTCF binding partners identified in other studies. Current work focuses on further characterizing the physical nature of these domains, identifying partners that bind with high affinity, and studying the complexes formed. We have participated in a collaboration investigating a possible role for SNPs affecting CTCF binding strength in expression of a gene associated with major mental illnesses. We have also begun a study of the organization of non-ribosomal genes within the nucleolus which is addressing both the genomic content and organization of the nucleolar genome. As part of our interest in large scale genome organization we have undertaken a study of the transcription factor MAZ, which is often bound at next to CTCF sites. We have observed interactions between the two proteins and an effect of MAZ on CTCF properties. As part of our interest in genome organization we have carried out a comparative study of the genomic content of nucleoli in multiple cell lines. This addresses the question of what non-ribosomal genes are present in nucleoli, with the goal of identifying the properties that determine that occupancy. Results show that there is a wide divergence of gene identity among cell types. All of these results relate to the role of chromatin structure, histone modifications, and long range organization of the genome in cell function, and are in turn related to questions of normal and abnormal cell metabolism and cell division.