Functional analysis of the CYLD tumor suppressor

CYLD抑癌基因的功能分析

• 批准号: 6870730
• 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/6870730
• 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（申请人提供）：CYLD是一种新型抑癌基因，是通过对染色体16q12上的家族性圆柱瘤病（FC）连锁间隔进行定位克隆而发现的。 FC 是一种常染色体显性遗传综合征，其特征是皮肤附属器肿瘤毁容，例如圆柱瘤、毛发上皮瘤和螺旋腺瘤。通常，这些肿瘤位于头皮和面部，出现在儿童期或成年早期，并在一生中逐渐增大大小和数量。此外，这些肿瘤可能发生恶性转化，具有局部侵袭行为以及远处转移。 CYLD 的种系突变已在 FC 家族中得到证实，并且在这些肿瘤中发现了 CYLD 基因座杂合性的丧失，表明 CYLD 具有肿瘤抑制因子的功能。 CYLD 的蛋白质产物由 956 个氨基酸组成，包含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，有趣的是，它在皮肤中的表达在表皮以及皮肤附属器中观察到。导致原癌基因功能获得或抑癌基因功能丧失的突变会导致肿瘤的发生。肿瘤抑制基因要么抑制增殖，促进细胞凋亡，要么增强分化，并通过调节不同的细胞通路（其中之一是 NF-KB 信号通路）维持基因组完整性。最近的数据表明，CYLD 具有使靶蛋白去泛素化的酶活性。它已被证明与 NF-KB 信号通路的多个成员（例如 TRAF-2、TRIP 和 IKKy/NEMO）相互作用，并负向调节 NF-KB 激活。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）发育严重缺陷。此外，NF-KB 信号传导异常在表皮瘤形成中也发挥着作用。然而，与 NF-KB 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，尚不清楚。我们的总体目标是确定 CYLD 在皮肤肿瘤发生中的功能。我们已经在 FC 患者的 CYLD 基因中发现了多种突变。然而，目前有关 CYLD 的突变数据有限。在目标 1 中，我们将评估 FC 的基因型和表型相关性，这将导致对皮肤附属器肿瘤的分子了解。作为定义 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 小鼠模型。最后，我们的初步研究表明 CYLD 稳定存在于 HeLa 细胞的细胞核和细胞质中。状态并且瘦霉素 B 治疗增加了其核定位。该观察结果表明CYLD以CRM1依赖性方式在细胞质和核区室之间组成型穿梭。然而，它在细胞核中的作用尚未确定。在目标 3 中，我们首先计划识别负责 CYLD 核质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，为了深入了解其核作用，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产生对 CYLD 调节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。