Functional analysis of DAN and DA41 proteins in early embryogenesis

DAN 和 DA41 蛋白在早期胚胎发生中的功能分析

• 批准号: 10680658
• 负责人: OZAKI Toshinori
• 金额: $ 2.18万
• 依托单位: Chiba Cancer Center Research Institute
• 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-10680658/
• 关键词: BMP4; DA41; DAN; Growth suppression; DA41; BMP2; 分泌蛋白質

(A) Functional analysis of DANNorthern blot analysis revealed that the expression of DAN was significantly down-regulated in human osteosarcoma cell line SAOS-2. Overexpression of DAN in SAOS-2 resulted in a massive growth suppression as demonstrated by colony formation assays. N-terminal truncated form of DAN did not exhibit the growth suppressive activity, indicating that the secreted form of DAN is active. Reporter gene assay demonstrated that DAN inhibited the BMP4 signaling in mouse embryonic carcinoma P19 cells. These observations suggest that DAN, as well as Cerberus and Gremlin, may function as an antagonist against BMP in fog embryos and mammalian cells.(B) Characterization of DA41Ectopic overexpression of rat DA41 in V-Ha-ras-transformed 3Y1 cells (ras-3Y1) resulted in a significant growth suppression, which was associated with a remarkable reduction of CDK2 activity. Fluorescence in situ hybridization revealed that DA41 was mapped to human chromosome 9q21.2-q21.3, a position overlapping the candidate tumor suppressor locus for bladder cancer. Data base search for DA41-related proteins(s) identified mouse PLIC-1, PLIC-2, frog XDRP1, and yeast DSK2. It has been shown that XDRP1 inhibited the degradation of cyclin A and blocked the cell division. These observations suggest that DA41 and XDRP1 could share the similar function in the cell cycle regulation.

(A) DAN功能分析northern blot分析显示，在人骨肉瘤细胞系SAOS-2中，DAN的表达明显下调。在SAOS-2中，DAN的过表达导致了大量的生长抑制，这一点在集落形成实验中得到了证实。n端截断形式的DAN没有表现出生长抑制活性，说明分泌形式的DAN是有活性的。报告基因实验表明，DAN抑制小鼠胚胎癌P19细胞BMP4信号通路。这些观察结果表明，DAN以及Cerberus和Gremlin可能在雾胚和哺乳动物细胞中作为BMP的拮抗剂。大鼠DA41在v - ha -ras-转化的3Y1细胞（ras-3Y1）中的异位过表达导致显著的生长抑制，这与CDK2活性的显著降低有关。荧光原位杂交显示，DA41定位于人类染色体9q21.2-q21.3，与膀胱癌候选肿瘤抑制基因位点重叠。对da41相关蛋白进行数据库检索，鉴定出小鼠PLIC-1、PLIC-2、青蛙XDRP1和酵母DSK2。研究表明，XDRP1能够抑制细胞周期蛋白A的降解，阻断细胞分裂。这些观察结果表明，DA41和XDRP1在细胞周期调控中可能具有相似的功能。

项目成果

Takada, N., Ozaki, T. et al.: "Identification of a transactivation activity in the COOH-terminal region of p73 which is impaired in the naturally occurring mutants found in human neuroblastomas"Cancer Res.. 59. 2810-2814 (1999)

Takada, N., Ozaki, T. 等人：“p73 COOH 末端区域反式激活活性的鉴定，该活性在人神经母细胞瘤中发现的天然突变体中受损”Cancer Res.. 59. 2810-2814（

Hanaoka E.,Ozaki T.et al.: "Molecular cloning and expression analysis of the human DA41 gene and its mapping to chromosome 9q21.2-q21.3"J.Hum.Genet.. (in press).

Hanaoka E.、Ozaki T.et al.：“人类 DA41 基因的分子克隆和表达分析及其与染色体 9q21.2-q21.3 的映射”J.Hum.Genet..（出版中）。

Takada N.,Ozaki T.et al.: "Identification of a transactivation activity in the COOH-terminal region of p73 which is impaired in the naturally occurring mutants found in human neuroblastomas"Cancer Res.. 59. 2810-2814 (1999)

Takada N.、Ozaki T.等人：“p73 COOH 末端区域反式激活活性的鉴定，该活性在人神经母细胞瘤中发现的天然突变体中受损”Cancer Res.. 59. 2810-2814 (1999)

Hishiki,T.,Ozaki,T.et al.: "GDNF/NTN-induced differentiation and its enhancement by retinoic acid in primary human neuroblastomas expressing c-Ret,GFRα-1 and GFRα-2." Cancer Res.58. 2158-2165 (1998)

Hishiki, T., Ozaki, T. 等人：“表达 c-Ret、GFRα-1 和 GFRα-2 的原发性人神经母细胞瘤中 GDNF/NTN 诱导的分化及其增强作用。”Cancer Res.58。 -2165 (1998)

Hanaoka, E., Ozaki T. et al.: "Molecular cloning and expression analysis of the human DA41 gene and its mapping to chromosome 9q21.2-q21.3"J.hum. Genet.. (in press).

Hanaoka, E., Ozaki T. 等人：“人类 DA41 基因的分子克隆和表达分析及其与染色体 9q21.2-q21.3 的映射”J.hum。