Functional analysis of the CYLD tumor suppressor

CYLD抑癌基因的功能分析

• 批准号: 7046015
• 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/7046015
• 关键词: autosomal dominant trait; bioassay; biotechnology; blood tests; clinical research; disease /disorder model; family genetics; gene mutation; genetic disorder; genetically modified animals; histology; human genetic material tag; human subject; immunocytochemistry; laboratory mouse; model design /development; neoplasm /cancer genetics; neoplastic process; nuclear factor kappa beta; nuclear transport; protein structure function; skin neoplasms; tissue /cell culture; tumor suppressor genes; tumor suppressor proteins

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是一种新型的肿瘤抑制基因，是通过在16q12染色体上的家族性胞染色体症（FC）的位置克隆发现的。 FC是一种常染色体，主要遗传综合征，其特征是毁容皮肤附属肿瘤，例如圆柱瘤，毛thoepitheliomas和spiradenomas。通常，这些肿瘤位于头皮和面部，出现在儿童时期或成年初期，并在整个生命中逐渐增加。此外，可能会发生这些具有局部侵入性行为以及远处转移的肿瘤的恶性转化。在FC家族中已经证明了CYLD中的种系突变，并且在这些肿瘤中发现了CYLD基因座的杂合性丧失，这表明CYLD充当肿瘤抑制器。 CYLD的蛋白质产物是956个氨基酸，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，并且感兴趣，在表皮和皮肤附属物中观察到其在皮肤中的表达。突变导致原始基因的功能或肿瘤抑制基因功能的丧失导致肿瘤发育。肿瘤抑制基因可以通过调节不同的细胞途径来抑制增殖，促进凋亡或增强分化，并维持基因组完整性，其中之一是NF-KB信号通路。最近的数据表明，CYLD具有去泛素化靶蛋白的酶促活性。已显示它与NF-KB信号通路的多个成员相互作用，例如TRAF-2，TRIP和IKKY/NEMO，并对NF-KB激活进行负面调节。然而，CYLD肿瘤抑制的分子和细胞机制在很大程度上尚不清楚。 NF-KB信号传导对于外胚层器官发生至关重要。 NF-KB抑制会导致表皮附属物（包括毛囊和汗腺）的出现严重缺陷。此外，NF-KB信号传导的异常在表皮肿瘤中起作用。然而，尚不清楚与NF-KB信号传导相关的肿瘤发育机制，尤其是CYLD依赖性肿瘤发生的作用。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基因中的各种突变。但是，目前，CYLD的突变数据受到限制。在AIM 1中，我们将评估FC中的基因型和表型相关性，这将导致基于分子的皮肤附属肿瘤的理解。作为定义CYLD介导的肿瘤抑制机制的关键步骤，我们将在AIM 2中建立小鼠模型。最后，我们的初步研究表明，CYLD存在于稳态状态下的Hela细胞中的细胞核和稳态细胞的细胞质中，并且Leptomycin B治疗会增加其核定位。该观察结果表明，CYLD以CRML依赖性方式在细胞质和核区室之间进行了组成型穿梭。但是，尚未定义其在细胞核中的作用。在AIM 3中，我们首先计划确定负责CYLD核总质梭子穿梭的功能性核输出信号，并评估NF-KB激活期间CYLD的定位。其次，为了洞悉其核作用，我们将尝试确定其在核中的相互作用伙伴。我们预计，在这些研究过程中，将出现对CYLD调节肿瘤抑制途径的大量见解，从而扩展我们对肿瘤发生的理解。