Structure and Function of CTCF: Mouse Model Studies

CTCF的结构和功能：小鼠模型研究

• 批准号: 7253908
• 负责人: GALINA N FILIPPOVA
• 金额: $ 38.55万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-06-05 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7253908
• 关键词: 16q22; 16q22.1; Alleles; Binding; Breast; Cell Death; Cell Survival; Chromatin; Clinical; Congenital Myotonic Dystrophy; DNA Binding Domain; Data; Development; Embryo; Embryonic Development; Epigenetic Process; Exhibits; Fingers; Gene Expression Regulation; Gene Silencing; Gene Targeting; Genetic; Genomic Imprinting; Green Fluorescent Proteins; Hormone Responsive; Human; Human Chromosomes; Knock-in Mouse; Knock-out; Knockout Mice; Link; Loss of Heterozygosity; Malignant Neoplasms; Maps; Methylation; Missense Mutation; Molecular; Mus; Mutate; Mutation; Nephroblastoma; Normal tissue morphology; Numbers; Pattern; Phenotype; Point Mutation; Predisposition; Prostate; Proteins; Protocols documentation; Regulation; Repression; Role; Site; Structure; System; Testing; Therapeutic Intervention; Tissues; Transgenic Mice; Tumor Suppression; Tumor Suppressor Genes; X Chromosome; X Inactivation; base; cancer cell; cancer prevention; imprint; insight; malignant phenotype; mouse model; mutant; prevent; promoter; transcription factor; tumor; tumor initiation; tumorigenesis

DESCRIPTION (provided by applicant): CTCF is a highly conserved 11 Zn finger (ZF) transcription factor and insulator protein. CTCF is involved in multiple aspects of gene regulation including promoter activation and repression, hormone-responsive gene silencing, methylation-dependent chromatin insulation, and genomic imprinting. Moreover, our recent discovery of a methylationsensitive CTCF insulator at the DM1 locus that might be responsible for the severe phenotype of Congenital Myotonic Dystrophy (Filippova et al.,2001), and our preliminary data on the role of CTCF insulators in X chromosome inactivation strongly suggest a link between CTCF and epigenetic regulation in development. Several findings also point to a role for CTCF as a tumor suppressor gene in both cancer epigenetics and genetics. Aberrant methylation of certain CTCF target sites, that is predicted to prevent CTCF-binding, has been demonstrated in a number of tumors. CTCF maps to human chromosome 16q22, within a region that displays frequent cancer-associated deletions (Filippova et al.,1998). We have observed somatic missense mutations within the CTCF 11ZF DNA-binding domain in breast, prostate and Wilms' tumors (Filippova et al.,2002). CTCF +/- mice exhibit enhanced tumor development in multiple tissues. Homozygous deletion of the CTCF results in early embryonic lethality, whereas the CTCF heterozygous mice exhibit decreased embryonic survival. Our broad and longterm hypothesis is that certain functions of CTCF are essential in early development and maintaining cell viability, whereas its other functions, when lost, result in the malignant phenotype. To test this hypothesis our specific aims will: Extend our studies on the role of CTCF in tumorigenesis (Aim 1) and early embryonic development (Aim 2). We will determine what genetic and epigenetic mechanisms are involved in CTCF haploinsufficiency. And finally (Aim 3), we will use the Cre-loxP system to generate a conditional CTCF mutant allele, that in combination with various tissue-specific Cre transgenic mice will allow us to test the hypothesis that complete loss of CTCF function leads to cell death in both normal and/or malignant cells, whereas loss of one CTCF allele in somatic tissue predisposes to tumor development. We will also generate knock-in mice harboring CTCF Zn finger point mutations that we have observed in human tumors to determine the functional significance of such mutated alleles in tumor predisposition. The significance of this proposal is that identifying the mechanisms of the CTCF haploinsufficiency will provide new insight into the broad spectrum of clinical human cancer phenotypes associated with LOH at 16q22.1 where CTCF maps. The health-relatedness is that the identified mechanisms can be targeted for both cancer prevention and therapeutic intervention.

描述（申请人提供）：CTCF是一种高度保守的11锌指（ZF）转录因子和绝缘子蛋白。CTCF参与基因调控的多个方面，包括启动子激活和抑制、激素应答性基因沉默、甲基化依赖性染色质绝缘和基因组印迹。此外，我们最近在DM1位点发现了甲基化敏感的CTCF绝缘子，这可能是导致先天性肌强张性营养不良严重表型的原因（Filippova等，2001），我们关于CTCF绝缘子在X染色体失活中的作用的初步数据强烈表明CTCF与发育中的表观遗传调控之间存在联系。一些研究结果还指出CTCF作为肿瘤抑制基因在癌症表观遗传学和遗传学中的作用。某些CTCF靶位点的异常甲基化，预计会阻止CTCF结合，已经在许多肿瘤中得到证实。CTCF映射到人类染色体16q22，在一个区域显示频繁的癌症相关缺失（Filippova et al.,1998）。我们已经在乳腺、前列腺和Wilms肿瘤中观察到CTCF 11ZF dna结合域内的体细胞错义突变（Filippova et al.,2002）。CTCF +/-小鼠在多个组织中表现出增强的肿瘤发展。纯合子缺失CTCF导致早期胚胎死亡，而CTCF杂合子小鼠表现出胚胎存活率降低。我们广泛和长期的假设是，CTCF的某些功能在早期发育和维持细胞活力中是必不可少的，而它的其他功能，当失去时，会导致恶性表型。为了验证这一假设，我们的具体目标是：扩展我们对CTCF在肿瘤发生（目标1）和早期胚胎发育（目标2）中的作用的研究。我们将确定CTCF单倍性不足的遗传和表观遗传机制。最后（目标3），我们将使用Cre- loxp系统生成一个条件CTCF突变等位基因，该等位基因与各种组织特异性Cre转基因小鼠相结合，将使我们能够验证CTCF功能的完全丧失导致正常和/或恶性细胞死亡的假设，而体细胞组织中CTCF等位基因的丧失易导致肿瘤发展。我们还将产生含有CTCF锌指点突变的敲入小鼠，我们已经在人类肿瘤中观察到这种突变等位基因在肿瘤易感性中的功能意义。该建议的意义在于，确定CTCF单倍不足的机制将为临床人类癌症表型的广谱谱提供新的见解，这些表型与CTCF所在的16q22.1位点的LOH相关。与健康相关的是，所确定的机制可以作为癌症预防和治疗干预的目标。

项目成果

Loss of maternal CTCF is associated with peri-implantation lethality of Ctcf null embryos.

• DOI: 10.1371/journal.pone.0034915
• 发表时间: 2012
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Moore JM;Rabaia NA;Smith LE;Fagerlie S;Gurley K;Loukinov D;Disteche CM;Collins SJ;Kemp CJ;Lobanenkov VV;Filippova GN
• 通讯作者: Filippova GN

CTCF cis-regulates trinucleotide repeat instability in an epigenetic manner: a novel basis for mutational hot spot determination.

• DOI: 10.1371/journal.pgen.1000257
• 发表时间: 2008-11
• 期刊: PLoS genetics
• 影响因子: 4.5
• 作者: Libby RT;Hagerman KA;Pineda VV;Lau R;Cho DH;Baccam SL;Axford MM;Cleary JD;Moore JM;Sopher BL;Tapscott SJ;Filippova GN;Pearson CE;La Spada AR
• 通讯作者: La Spada AR

Cell growth inhibition by the multifunctional multivalent zinc-finger factor CTCF.

• 发表时间: 2001-08
• 期刊: Cancer research
• 影响因子: 11.2
• 作者: J. Rasko;E. Klenova;J. Leon;G. Filippova;D. Loukinov;S. Vatolin;A. Robinson;Y. Hu;J. Ulmer;M. Ward;E. Pugacheva;P. Neiman;H. Morse;S. Collins;Victor V Lobanenkov