Analysis of cellular function of Neurofibromatosis Type 2 tumor suppressor gene product (Merlin)

神经纤维瘤病2型抑癌基因产物（Merlin）的细胞功能分析

• 批准号: 10671308
• 负责人: ARAKI Norie
• 金额: $ 2.43万
• 依托单位: Kumamoto University School of Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671308/
• 关键词: Neurofibromatosis Type 2; NF2; Merlin; Tumor suppressor gene; Poly (ADP-ribose) polymerase (PARP); Ku-antigen; Cellular signal transduction; Nuclear export signal (NES); NF2、; 神経繊維腫症2型

The neurofibromatosis type 2 (NF2) is an autosomal dominantly inherited disorder, and strongly associated with development of benign intracranial tumors including bilateral vestibular schwannomas and meningiomas. NF2 gene was recently cloned, and the protein it encodes (termed as merlin/schwannomin) was found to be striking similarity to the moesin-ezrin-radixin (MER) family of cytoskeleton-associated proteins. To elucidate the biological function of NF2 protein (merlin), we analyzed the mutation pattern of merlin, its cellular localization, and the cellular signals via merlin. 1) NF2 gene mutations in NF2 patients were analyzed by protein truncation test. The majority of NF2 mutations are nonsense, frame shift, or exon-missing mutations. More than 70%of the mutations are clustered in the region encoding the N-terminal half of merlin. In addition, post-transrational proteolytic cleavage of N-terminal merlin by the ubiquitous protease calpain was found in some schwannomas and meningioma … More s. These results suggest that the N-terminal region of merlin is the most important for the tumor suppressive function, 2) Cellular expression systems of mutants or wild NF2 were established, and their cellular localization was analyzed by confocal lazer scanning Microscopy (CLSM). The wild merlin showed cytoplasmic and submembranous localization that was found to be directed by its nuclear export signal (NES) dependent transport mechanism. The N-terminal mutation resulted in nuclear accumulation of merlin. 3) Cellular binding proteins of merlin were purified from bovine brain extracts and identified as poly ADP-ribose polymerase, DNA-PK subunits Ku-antigen 85 and 70 by the analysis of their internal amino-acid sequences. The N-terminal(19-339) region of merlin is essential for their interaction. The immuno-precipitation, western blotting, and CLMS study confirmed their cellular co-localization. NF2 mutations impair the merlin-related complex formation and their cellular signal transport. This loss of cellular function of merlin may link to the intracranial tumor generation. Less

神经纤维瘤病2型（NF 2）是一种常染色体显性遗传疾病，与良性颅内肿瘤包括双侧前庭神经鞘瘤和脑膜瘤的发生密切相关。NF 2基因是最近克隆的，它编码的蛋白质（称为merlin/schwannomin）被发现与细胞凋亡相关蛋白的moesin-ezrin-radixin（MER）家族具有惊人的相似性。为了阐明NF 2蛋白（merlin）的生物学功能，我们分析了merlin的突变模式、细胞定位以及通过merlin传递的细胞信号。1)采用蛋白质截短试验分析NF 2患者NF 2基因突变情况。大多数NF 2突变是无义突变、移码突变或外显子缺失突变。超过70%的突变聚集在编码merlin的N-末端一半的区域。此外，在一些神经鞘瘤和脑膜瘤中发现了普遍存在的蛋白酶钙蛋白酶对N-末端merlin的翻译后蛋白水解切割 ...更多信息 S. 2）建立了突变体和野生型NF 2的细胞表达系统，并利用激光共聚焦显微镜（CLSM）对突变体和野生型NF 2的细胞定位进行了分析。野生型merlin蛋白在细胞质和膜下的定位与其依赖于核输出信号（内斯）的转运机制有关。N-末端突变导致merlin在核中积累。3)从牛脑提取物中纯化了merlin的细胞结合蛋白，经氨基酸序列分析鉴定为聚ADP-核糖聚合酶、DNA-PK亚基Ku-抗原85和70。Merlin蛋白的N端（19-339）区域是它们相互作用的关键。免疫沉淀、蛋白质印迹和CLMS研究证实了它们的细胞共定位。NF 2突变损害merlin相关复合物的形成及其细胞信号传递。这种梅林细胞功能的丧失可能与颅内肿瘤的产生有关。少

项目成果

Kuwahara H, Araki N, Makino K, Masuko N, Honda S, Kaibuchi K, Fukunaga K Miyamoto E, Ogawa M, and Saya H: "A novel NE-dlg/SAP102-associated protein, p51-nedasin, related to the aminohydrolyse superfamily, interferes with the association between NE-dlg/SAP

Kuwahara H、Araki N、Makino K、Masuko N、Honda S、Kaibuchi K、Fukunaga K Miyamoto E、Okawa M 和 Saya H：“一种新型 NE-dlg/SAP102 相关蛋白 p51-nedasin，与氨基水解有关

masuko N, et al.: "Interaction of NE-dlg/SAP102, a neuron and endocrine tissue-specific membrane-associated guanylate kinase protein, with calmodulin and PSD-95/SAP90. A possible reguulatory role in molecular clustering at synaptic sites"J. Biol. Chem.. 2

Masuko N 等人：“NE-dlg/SAP102（一种神经元和内分泌组织特异性膜相关鸟苷酸激酶蛋白）与钙调蛋白和 PSD-95/SAP90 的相互作用。突触位点分子聚集中可能的调节作用”

Araki,N.,Saya,H.: "Cellular signal transduction via the neurofibromatosis type 2 tumor supressor gene product;Merlin."Seikagaku. 71(2). 43-49 (1999)

Araki,N.,Saya,H.：“通过神经纤维瘤病 2 型肿瘤抑制基因产物进行细胞信号转导；Merlin。”Seikagaku。

Araki,N.: "Cellular signal transduction via the neurofibromatosis type 2 tumor supressor gene product;Merlin." Seikagaku. 71(2). 43-49 (1999)

Araki,N.：“通过神经纤维瘤病 2 型肿瘤抑制基因产物进行细胞信号转导；Merlin。”

Shibayama R. et al.: "Autoantibody against N(epsilon)-(carboxymethyl)lysine: an advanced glycation end product of the Maillard reaction"Diabetes.. 48(9). 1842-1849 (1999)

Shibayama R.等人：“针对N（ε）-（羧甲基）赖氨酸的自身抗体：美拉德反应的高级糖基化终产物”糖尿病.. 48（9）。