Cloning and characterization of a possible adhesion molecule expressed by hepatic non-parenchymal cells

肝非实质细胞表达的可能粘附分子的克隆和表征

• 批准号: 06670523
• 负责人: OGATA Itsuro
• 金额: $ 1.41万
• 依托单位: University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06670523/
• 关键词: Hepatic stellate cell; protein-kinase; Serine; threonine protein-kinase; Endothelial cell; Kupffer cell; Non-parenchymal cell; Molecular biology; フッパー細胞; スレオニンプロティンキナーゼ

Cells change their phenotype and functions in response to the specificity of the extracellular matrix. We now report the cloning and characterization of a cDNA encoding a new member of the protein-kinase family, which can be involved in the cell-matrix interaction. The expression cDNA library obtained from a fat-storing cell (FSC) line was screened with an antibody prepared against a plasma membrane fraction of rat hepatocytes, which was supposed to contain the matrix receptor. The antibody reacted with a protein localized on the sinusoidal domain of hepatocytes. It also recognized various proteins synthesized by FSC.From 1.5x10^5 phages, 11 clones were isolated. Ten of the 11 clones derived from the same mRNA corresponding an unkown protein. Finally, we dtermined the nucleotide sequence of 6456bp. The predicted protein of this clone had two copies of serine/threonine protein-kinase domains in N-terminal end, which showed less than 40% homology with catalytic domains of previously reported protein kinases. In the C-terminus, it contained a leucine zipper domain. The cDNA recognized a 7kb mRNA in freshly isolated and cultured FSC,endothelial and Kupffer cells. In contrast, this mRNA was not detected in hepatocytes. The expression of this mRNA appeared to decrease in cultured FSC.The function of the cloned cDNA is currently under investigation.

细胞响应细胞外基质的特异性而改变其表型和功能。我们现在报告的cDNA编码的蛋白激酶家族，它可以参与细胞基质相互作用的新成员的克隆和表征。从脂肪储存细胞（FSC）系获得的表达cDNA文库进行了筛选与针对大鼠肝细胞，这是应该包含基质受体的质膜部分制备的抗体。抗体与肝细胞窦状结构域上的蛋白质反应。从1.5 × 10^5个克隆中分离到11个克隆。11个克隆中有10个克隆来源于相同的mRNA，对应一种未知的蛋白质。最后，我们测定了6456 bp的核苷酸序列。该克隆预测的蛋白质在N端含有两个丝氨酸/苏氨酸蛋白激酶结构域，与已报道的蛋白激酶催化结构域的同源性低于40%。在C-末端，其含有亮氨酸拉链结构域。该cDNA在新鲜分离和培养的FSC、内皮细胞和枯否细胞中识别7 kb的mRNA。相反，在肝细胞中未检测到该mRNA。该mRNA在培养的FSC中的表达出现下降。克隆的cDNA的功能目前正在研究中。

项目成果

尾形逸郎: "Cloning and characterization of a new member of the protein-Kinase family from a rat fat-storing cell cDNA library." Hepatology. 18. 169A (1993)

Ituro Ogata：“从大鼠脂肪储存细胞 cDNA 文库中克隆和表征蛋白质激酶家族新成员”，《肝病学》18. 169A (1993)。

松井淳(分担): "国際医書出版" 肝類洞壁細胞研究の進歩〔第7巻〕. 261 (1995)

Jun Matsui（撰稿人）：《国际医学图书出版》肝窦壁细胞研究进展[第7卷]（1995年）。

松井 淳: "Inhibition of protein synthesis in liver stellate cells (LSC) by an antisense oligonucleotide from an RNA-binding protein of activated LSC." Hepatology. 22. 281A (1995)

Jun Matsui：“来自激活的 LSC 的 RNA 结合蛋白的反义寡核苷酸对肝星状细胞 (LSC) 中蛋白质合成的抑制。” 22. 281A (1995)。

尾形逸郎: "Cloning and characterization of a new member of the protein-kinase family from a rat fat-storing cell cDNA library." Hepatology. 18. 169A- (1993)

Ituro Ogata：“从大鼠脂肪储存细胞 cDNA 文库中克隆和表征蛋白质激酶家族新成员”，18. 169A- (1993)。

尾形逸郎: "Rat liver fat-storing cell lines express sarcomeric myosin heavy chain mRNA and protein." Cell Motility and the Cytoskeleton. 26. 125-132 (1993)

Ituro Ogata：“大鼠肝脏脂肪储存细胞系表达肌节肌球蛋白重链 mRNA 和蛋白质。”细胞运动性和细胞骨架（1993 年）。