Molecular basic research for obtaining genes specifying characteristics of pancreatic α and β cells.

获得指定胰腺α和β细胞特征的基因的分子基础研究。

• 批准号: 12671115
• 负责人: YOSHIMOTO Katsuhiko
• 金额: $ 2.3万
• 依托单位: The University of Tokushima
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671115/
• 关键词: pancreatic islet; large cDNA sequencing; β-cell specific expression; DNA microarray; large scale cDNA squereing; セリンプロテアーゼ; 膵β細胞特異的遺伝子発現

To characterize expressed sequence tag (EST) profiling in pancreatic islet β cells, a large-scale cDNA sequencing analysis was carried out for 3,429 ESTs derived from MIN6 cells, a cell line of murine pancreatic islet β cells. Homology comparison with the GenBank database revealed 369 unknown ESTs, which corresponded to those in the EST database (dbEST). Among these, 3 individual ESTs were found to code one novel protein which we termed isletasin. Isletasin gene encoded two variant mRNAs of a short and long form, from which two forms of isletasin with 482 and 552 amino acids were generated, respectively. Isletasin contained repeated catalytic triads for S1 family of serine proteases, and its expression was limited to MIN6 cells or mouse pancreatic islets. Immunoblotting of extract from COS-7 cells expressing epitope-tagged isletasin showed that isletasin is not an integral but a peripheral protein. Isletasin is strongly associated with the luminal surface of the microsomal membrane, where it undergoes signal peptide cleavage and N-glycosylation. The fluorescence detection of each organelle marker protein and green fluorescence protein (GFP)-tagged isletasin in MIN6 cells revealed that isletasin is an early endosomal protein. In conclusion, novel protein of isletasin of which cDNA was efficiently obtained by screening 3,429 ESTs derived from MIN6 cells, is specifically expressed in pancreatic islet β cells, as an early endosomal protein.

为了研究胰岛β细胞表达序列标签的特征，对小鼠胰岛β细胞系MIN6细胞中的3,429条EST进行了大规模的序列分析。与GenBank数据库进行同源性比较，发现369条未知EST序列，与EST数据库(DBEST)中的EST序列相对应。其中，3个单独的EST编码一种新的蛋白质，我们将其命名为Isetasin。Isletasin基因编码两种不同形式的mRNA，分别含有482和552个氨基酸。Isletasin含有对S1家族丝氨酸蛋白酶的重复催化三联体，其表达仅限于MIN6细胞或小鼠胰岛。免疫印迹分析表明，表达表位标记的异莱菌素的COS-7细胞提取物不是一个完整的蛋白质，而是一种外周蛋白。Isletasin与微体膜的管腔表面密切相关，在那里它经历信号肽的裂解和N-糖基化。对MIN6细胞中各细胞器标志蛋白和绿色荧光蛋白(GFP)标记的异莱菌素的荧光检测表明，异莱他辛是一种早期的内体蛋白。综上所述，从MIN6细胞中筛选出3,429条EST，获得了一种在胰岛β细胞中特异表达的新蛋白，该蛋白是一种早期的内体蛋白。

项目成果

Y Imanishi, Y Hosokawa, K Yoshimoto, E Schipani, S Mallya, A Papanikolaou, O Kifor, T Tokura, M Sablosky, F Ledgard, G Grnowicz, TC Wang, EV Schmidt, C Hall, E M Brown, R Bronson, A Arnold: "Primary Hyperparathyroidism Caused by Parathyroid-Targeted Overe

Y Imanishi、Y Hosokawa、K Yoshimoto、E Schipani、S Mallya、A Papanikolaou、O Kifor、T Tokura、M Sablosky、F Ledgard、G Grnowicz、TC Wang、EV Schmidt、C Hall、E M Brown、R Bronson、A Arnold

ZR Qian, CC Li, H Yamasaki, N Mizusawa, K Yoshimoto, S Yamada, T Tashiro, H Horiguchi, S Wakatsuki, M Hirokawa, T Sano: "Role of E-cadherin, α-, β-, and γ-Catenins, and p120 (Cell Adhesion Molecules) in Prolactinoma Behavior."Mod Pathol. 15(12). 1357-1365

ZR Qian、CC Li、H Yamasaki、N Mizusawa、K Yoshimoto、S Yamada、T Tashiro、H Horiguchi、S Wakatsuki、M Hirokawa、T Sano：“E-钙粘蛋白、α-、β-和 γ-连环蛋白的作用和 p120（细胞粘附分子）在催乳素瘤行为中的作用。”Mod Pathol. 15(12). 1357-1365

Bing Xu et al.: "A Predominant Increase of APC Gene Isoform with Exon 9a in a Case of Attenuated Familial Adenomatous Polyposis"Clin Genet. 63(1). 71-72 (2003)

Bing Xu 等人：“A PC 基因亚型与外显子 9a 的显着增加在一例减毒型家族性腺瘤性息肉病中”Clin Genet。

T Yamaoka, K Yoshino, T Yamada, C Idehara, MO Hoque, M Moritani, K Yoshimoto, J Hata, M Itakura: "Diabetes and tumor formation in transgenic mice expressing reg 1."Biochem Biophys Res Commun. 278. 368-376 (2000)

T Yamaoka、K Yoshino、T Yamada、C Idehara、MO Hoque、M Moritani、K Yoshimoto、J Hata、M Itakura：“表达 reg 1 的转基因小鼠中的糖尿病和肿瘤形成。”Biochem Biophys Res Commun。

Zhi Rong Qian et al.: "Role of E-cadherin, α-, β-, and γ-Catenins, and p120 (Cell Adhesion Molecules) in Prolactinoma Behavior"Mod Pathol. 15(12). 1357-1365 (2002)

钱志荣等人：“E-钙粘蛋白、α-、β-和 γ-连环蛋白以及 p120（细胞粘附分子）在催乳素瘤行为中的作用”Mod Pathol 15(12) 1357-1365。