Functional analysis of the CYLD tumor suppressor

CYLD抑癌基因的功能分析

• 批准号: 7382527
• 负责人: Julide T. Celebi
• 金额: $ 12.4万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7382527
• 关键词: 16q12; Ablation; Affect; Alleles; Amino Acids; Animal Model; Animals; Apoptosis; Behavior; Binding; Binding Proteins; Biochemical; Biological; Biological Assay; Cell Cycle Regulation; Cell Nucleus; Cell Proliferation; Cells; Childhood; Chromosomes; Clinical; Clinical Research; Code; Correlation Studies; Cutaneous; Cytoplasm; Cytoplasmic Protein; Data; Defect; Deubiquitinating Enzyme; Development; Disease; Distant Metastasis; Dominant-Negative Mutation; Eccrine Spiradenoma; Embryo; Epidermis; Exhibits; Exons; Family; Generations; Genes; Genomics; Genotype; Germ-Line Mutation; Goals; Hair Follicle Neoplasm; Hair follicle structure; Hela Cells; Heterogeneity; Histology; Human; Hydrolase; Immunohistochemistry; Individual; Inherited; Invasive; Knock-out; Lead; Life; Loss of Heterozygosity; Malignant - descriptor; Mediating; Microtubules; Missense Mutation; Molecular; Mus; Mutant Strains Mice; Mutation; Neoplasms; Nonsense Codon; Nuclear; Nuclear Export; Numbers; Organogenesis; Pathway interactions; Patients; Personal Satisfaction; Phenotype; Phosphotransferases; Play; Principal Investigator; Proliferation Marker; Proteins; Proto-Oncogenes; Protocols documentation; Regulation; Reporting; Research Personnel; Role; Scalp structure; Signal Pathway; Signal Transduction; Signaling Molecule; Skin; Skin Appendage Neoplasm; Skin Neoplasms; Sweat Gland Neoplasms; Sweat Glands; Syndrome; Thinking; Tissues; Trichoepithelioma; Tumor Suppression; Tumor Suppressor Genes; Tumor Suppressor Proteins; Ubiquitin; appendage; base; clinical phenotype; embryonic stem cell; gain of function; homologous recombination; human disease; insertion/deletion mutation; insight; interest; leptomycin B; loss of function; mRNA Decay; member; mouse model; novel; p65; positional cloning; programs; size; tumor; tumorigenesis

DESCRIPTION (provided by applicant): CYLD is a novel tumor suppressor gene that was discovered by positional cloning of the linkage interval for Familial Cylindromatosis(FC) on chromosome 16q12. FC is an autosomal dominantly inherited syndrome characterized by disfiguring skin appendage tumors, such as cylindromas, trichoepitheliomas and spiradenomas. Typically these tumors are located on the scalp and face,appear in childhood or early adulthood, and gradually increase in size and number throughout life. Moreover, malignant transformation of these tumors with locally invasive behavior as well as distant metastasis can occur. Germline mutations in CYLD have been demonstrated in families with FC, and loss of heterozygosity at the CYLD locus has been found in these neoplasms, suggesting that CYLD functions as a tumor suppressor. The protein product of CYLD is 956 amino acids and contains sequence motifs found in deubiquitinating enzymes and microtubule binding proteins. It is expressed in a variety of tissues, and of interest, its expression in the skin is observed in the epidermis as well as in the skin appendages. Mutations leading to gain of function of proto-oncogenes or loss of function of tumor suppressor genes result in tumor development. Tumor suppressor genes either inhibit proliferation, promote apoptosis, or enhance differentiation, and maintain genomic integrity via regulation of distinct cellular pathways, one of which is the NF-KB signaling pathway. Recent data suggests that CYLD has enzymatic activity to deubiquitinate target proteins. It has been shown to interact with several members of the NF-KB signaling pathway, such as TRAF-2, TRIP, and IKKy/NEMO, and negatively regulate NF-KB activation. However, the molecular and cellular mechanism(s) of CYLD tumor suppression is largely unknown. NF-KB signaling is essential for ectodermal organogenesis. NF-KB suppression results in severe defects in the development of epidermal appendages including hair follicles and sweat glands. In addition, abnormalities in NF-KB signaling play a role in epidermal neoplasia. However, the mechanisms of tumor development related to NF-KB signaling, and in particular the role of CYLD-dependent tumorigenesis, are not well understood. Our overall goal is to define the functions of CYLD in cutaneous tumorigenesis. We have identified a variety of mutations in the CYLD gene in patients with FC. However, the mutational data on CYLD is currently limited. In Aim 1, we will evaluate genotype and phenotype correlation in FC that will lead to a molecular-based understanding of the skin appendage tumors. As a crucial step in defining the mechanisms of CYLD-mediated tumor suppression, we will establish a mouse model for FC in Aim 2. And lastly, our Preliminary Studies demonstrate that CYLD is present in both the nucleus and the cytoplasm of HeLa cells at steady state and that leptomycin B treatment increases its nuclear localization. This observation suggests that CYLD constitutively shuttles between cytoplasmic and nuclear compartments in a CRMl-dependent manner. However, its role in the nucleus has not been defined. In Aim 3, we first plan to identify a functional nuclear export signal responsible for nucleo-cytoplasmic shuttling of CYLD and evaluate localization of CYLD during NF-KB activation. Second, to provide insights into its nuclear role, we will attempt to identify its interaction partners in the nucleus. We anticipate that significant insights into the pathway of CYLD regulated tumor suppression will arise in the course of these studies, thereby extending our understanding of tumorigenesis.

描述（由申请人提供）：CYLD是一种新的肿瘤抑制基因，通过对染色体16 q12上家族性乳腺癌（FC）连锁区间的定位克隆发现。FC是一种常染色体显性遗传综合征，以皮肤附件肿瘤（如圆柱瘤、螺旋上皮瘤和螺旋腺瘤）为特征。通常这些肿瘤位于头皮和面部，出现在儿童或成年早期，并在一生中逐渐增加大小和数量。此外，这些肿瘤的恶性转化具有局部侵袭性行为以及远处转移可能发生。CYLD的生殖系突变已被证明在家庭与FC，并在CYLD基因座的杂合性丢失已被发现在这些肿瘤，表明CYLD功能作为一种肿瘤抑制剂。CYLD的蛋白质产物是956个氨基酸，并且包含在去泛素化酶和微管结合蛋白中发现的序列基序。它在多种组织中表达，并且令人感兴趣的是，在表皮以及皮肤附属物中观察到它在皮肤中的表达。导致原癌基因功能获得或肿瘤抑制基因功能丧失的突变导致肿瘤发展。肿瘤抑制基因通过调节不同的细胞通路（其中之一是NF-κ B信号通路）抑制增殖、促进凋亡或增强分化并维持基因组完整性。最近的数据表明，CYLD具有去泛素化靶蛋白的酶活性。它已被证明与NF-κ B信号通路的几个成员，如TRAF-2，TRIP和IKKy/NEMO相互作用，并负调节NF-κ B活化。然而，CYLD肿瘤抑制的分子和细胞机制在很大程度上是未知的。NF-κ B信号传导对于外胚层器官发生是必需的。NF-κ B抑制导致包括毛囊和汗腺在内的表皮附属物发育的严重缺陷。此外，NF-κ B信号传导的异常在表皮瘤形成中起作用。然而，与NF-κ B信号传导相关的肿瘤发展机制，特别是CYLD依赖性肿瘤发生的作用，还没有很好地理解。我们的总体目标是确定CYLD在皮肤肿瘤发生中的功能。我们已经确定了FC患者CYLD基因的各种突变。然而，CYLD的突变数据目前有限。在目标1中，我们将评估FC的基因型和表型相关性，这将导致对皮肤附件肿瘤的分子基础的理解。作为确定CYLD介导的肿瘤抑制机制的关键一步，我们将在目标2中建立FC的小鼠模型。最后，我们的初步研究表明，CYLD是目前在细胞核和细胞质中的HeLa细胞在稳定状态和来普霉素B处理增加其核定位。该观察结果表明CYLD以CRM 1依赖性方式在细胞质和细胞核区室之间组成性穿梭。然而，其在核心中的作用尚未确定。在目标3中，我们首先计划鉴定负责CYLD的核质穿梭的功能性核输出信号，并评估NF-κ B活化期间CYLD的定位。第二，为了深入了解它的核作用，我们将试图确定它在核中的相互作用伙伴。我们预计，在这些研究的过程中，将出现对CYLD调节的肿瘤抑制途径的重要见解，从而扩展我们对肿瘤发生的理解。