Functional Analysis of the von Hippel-Lindau Disease Tumor Suppressor Gene

冯·希佩尔-林道病肿瘤抑制基因的功能分析

• 批准号: 10671488
• 负责人: YAO Masahiro
• 金额: $ 2.05万
• 依托单位: Yokohama City University
• 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-10671488/
• 关键词: VHL gene; tumor suppressor gene; renal cell carcinoma; 腎細胞癌; DHL遺伝子

We found that, in normal human renal proximal tubule epithelial cells, the steady-state amount of VHL protein is strictly regulated by cell density. The cellular VHL content is more than 100-fold higher in dense cultures than in sparse cultures. The growth rates of renal cell cacinoma cells lacking an intact VHL gene and their derivatives with wild-type or mutant VHL expression vector do not differ significantly when they are growing in log-phase. Importantly, however, there is a difference when they reach confluency : cells lacking wild-type VHL grew continuously, while cells expressing exogenous VHL protein showed relatively limited cell growth. VHL protein functions as a growth suppressor at high cell density, and this might be the basis of the tumor suppressor function of VHL. We investigated the role of the VHL gene in CNS development using rodent CNS progenitor cells. We show that expression of the VHL protein is correlated with neuronal differentiation but not with glial differe … More ntiation in CNS progenitor cells, and we also show that VHL gene transduction induces neuronal differentiation. In addition, a VHL mRNA antisense oligonucleotide inhibits differentiation of CNS progenitor cells and up-regulates their cell cycle. The VHL protein contains two domains, alpha and beta. We report that the beta-domain interacts directly with atypical PKC isotypes, PKC zeta and PKC lambda. Further, the regulatory domain of aPKC is sufficient for this direct protein-protein interaction. To investigate the potential role of the PTEN gene in renal tumorigenesis, we searched for abnormalities of the gene in 68 primary renal-cell carcinomas (RCCs) as well as in 17 renal carcinoma-derived cell lines. Five of 68 (7.5 %) primary RCCs exhibited intragenic mutations, and 1 of 17 (5.9 %) cell lines had an insertion mutation. Four of the 5 primary tumors with PTEN mutation were high-grade, advanced clear-cell RCCs with distant metastases or renal vein tumor invasions, resulting in poor prognostic courses. PTEN mutation is observed m a subset of RCCs and that, especially in clear-cell RCCs, it occurs as a late-stage event and may contribute to the invasive and/or metastatic tumor phenotype. Less

我们发现，在正常的人肾近端小管上皮细胞中，VHL蛋白的稳态量受到细胞密度的严格调控。密集培养中的细胞VHL含量是稀疏培养中的100多倍。缺乏完整VHL基因的肾癌细胞及其与野生型和突变型VHL表达载体的衍生体在对数生长期的生长速度没有显著差异。然而，重要的是，当它们达到融合时，有一个不同：缺乏野生型VHL的细胞持续生长，而表达外源VHL蛋白的细胞显示出相对有限的细胞生长。VHL蛋白在细胞密度较高时具有生长抑制作用，这可能是VHL肿瘤抑制功能的基础。我们利用啮齿类CNS祖细胞研究了VHL基因在中枢神经系统发育中的作用。我们发现vhl蛋白的表达与神经元分化有关，但与胶质细胞分化…无关。在中枢神经前体细胞中有更多的分化，我们还发现VHL基因转导可以诱导神经元分化。此外，VHL mRNA反义寡核苷酸可抑制CNS祖细胞的分化并上调其细胞周期。VHL蛋白包含两个结构域，即α和β。我们报道了β结构域与非典型的PKC亚型PKC Zeta和PKC lambda直接相互作用。此外，aPKC的调节域足以实现这种直接的蛋白质-蛋白质相互作用。为了研究PTEN基因在肾脏肿瘤发生中的潜在作用，我们研究了68例原发肾细胞癌(RCC)以及17个肾癌来源的细胞系中PTEN基因的异常。68例原发肾癌中有5例(7.5%)存在基因内突变，17株原发肾癌中有1株(5.9%)存在插入突变。PTEN基因突变的5个原发肿瘤中有4个是高级别、晚期透明细胞肾癌，伴有远处转移或肾静脉肿瘤侵犯，导致预后不良。PTEN突变见于RCC的一个亚群，尤其是在透明细胞RCC中，它是一种晚期事件，可能与侵袭和/或转移的肿瘤表型有关。较少

项目成果

Kondo K, Yao M, Kobayashi K, Yoshida M, Kaneko S, Baba M, Sakai N, Kishida T, Kawakami S, Nagashima Y, Nakatani Y, Hosaka M: "PTEN/MMAC1/TEP1 mutations in human primary renal cell carcinomas and renal carcinoma cell lines"Int J Cancer. 91-2. 219-224 (2001

Kondo K、Yao M、Kobayashi K、Yoshida M、Kaneko S、Baba M、Sakai N、Kishida T、Kawakami S、Nagashima Y、Nakatani Y、Hosaka M：“人原发性肾细胞癌中的 PTEN/MMAC1/TEP1 突变和

Okuda H, Yao M et al.: "Direct interaction of the β-domain of the VHL tumor suppressor protein with the regulatorv domain of atypical PKC isotypes"Biochem Biophys Res Commun. 263・2. 491-497 (1999)

Okuda H、Yao M 等：“VHL 肿瘤抑制蛋白的 β 结构域与非典型 PKC 同种型的调节结构域的直接相互作用”Biochem Biophys Res Commun. 263·2。

Yoshida M, Ashida S, Kondo K, Kobayashi K, Kanno H, Shinohara N, Shitara N, Kishida T, Kawakami S, Baba M, Yamamoto I, Hosaka M, Shuin T, Yao M: "Germ-line Mutation Analysis in Patients with Von Hippel-Lindau Disease in Japan : an Extended Study of 77 Fam

Yoshida M、Ashida S、Kondo K、Kobayashi K、Kanno H、Shinohara N、Shitara N、Kishida T、Kawakami S、Baba M、Yamamoto I、Hosaka M、Shuin T、Yao M：“患者种系突变分析