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Structure and Function of Laminin-5, a Cell Adhesion Protein of Basement Membrane

基底膜细胞粘附蛋白Laminin-5的结构与功能

基本信息

批准号：
11490030
负责人：
MIYAZAKI Kaoru
金额：
$ 3.33万
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 (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11490030/
关键词：
laminin-4 cell adhesion cell motility globular domain processing MT-MMP tumor invasion basement membrane

项目摘要

The basement protein laminin-5(LN5), a heterotrimer of laminin α3, β3 and γ2 chains, potently promotes cellular adhesion and motility. In this study, we investigated the structure and function relationship of LN5 and its pathological roles in tumor invasion. 1) The α3 chain of LN5 contains five globular domains (G1 to G5) in its carboxyl end. To clarify the function of each G domain, we transfected cDNAs for the full length and five deletion derivatives of the α3 chain into human HT1080 cells, which express only the laminin β3 andγ2 chains, and prepared recombinant LN5s lacking each G domain. The deletion of G5 or G4-5 did not affect the biological activities of LN5, but the LN5 without G3-5 showed little activities of cell adhesion and migration. It was also found that G3 contained two different regions to stimulate cell adhesion and migration. Furthermore, we found that the α3 chain was cleaved between G3 and G4 after secretion and determined the cleavage site. 2) The 105-kDa γ2 chain of LN5 is cleaved at an amino-terminal region to produce the 105-kDa form. We found that MT1-MMP catalyzed this processing, and that this processing increased the cell motility activity of LN5 but decreased its cell adhesion activity. These results demonstrate that the amino-terminal region of γ2 chain has an important effect on the biological activity of LN5. 3) Immunohistochemical analysis showed that the laminin γ2 chain monomer was overexpressed in invading human carcinoma cells. When a cDNA for the amino-terminal regions of the γ2 chain was transfected into a human carcinoma cell line, the invasive potential of this cell line in vivo was greatly enhanced. These results suggest that the γ2 chain itself plays some important roles in tumor invasion.

层粘连蛋白-5（laminin-5, LN5）是层粘连蛋白α3、β3和γ2链的异源三聚体，能有效促进细胞粘附和运动。在本研究中，我们探讨了LN5的结构和功能关系及其在肿瘤侵袭中的病理作用。1) LN5 α3链羧基端含有G1 ~ G5 5个球状结构域。为了明确每个G结构域的功能，我们将α3链全长cdna和α3链的5个缺失衍生物转染到人HT1080细胞中，只表达层粘连蛋白β3和γ2链，并制备了缺失每个G结构域的重组LN5s。G5或G4-5的缺失不影响LN5的生物活性，但缺失G3-5的LN5细胞粘附和迁移活性不明显。还发现G3含有两个不同的区域来刺激细胞粘附和迁移。此外，我们发现分泌后α3链在G3和G4之间发生了断裂，并确定了断裂位点。2) LN5的105-kDa γ - 2链在氨基末端被裂解生成105-kDa形式。我们发现MT1-MMP催化了这一过程，并且这一过程增加了LN5的细胞运动活性，但降低了其细胞粘附活性。这些结果表明，γ - 2链氨基末端区域对LN5的生物活性有重要影响。3)免疫组化分析显示，层粘连蛋白γ - 2链单体在侵袭的人癌细胞中过表达。将γ - 2链氨基末端的cDNA转染到人癌细胞中，可显著增强癌细胞在体内的侵袭能力。这些结果表明γ - 2链本身在肿瘤侵袭过程中起着重要作用。