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.

地下蛋白层粘连蛋白5（LN5）是层粘连蛋白α3，β3和γ2链的异构体，可能会促进细胞粘合剂和运动。在这项研究中，我们研究了LN5及其在肿瘤侵袭中的病理作用的结构和功能关系。 1）LN5的α3链在其羧基端包含五个全球域（G1至G5）。为了阐明每个G结构域的功能，我们将全长的cDNA和α3链的五个缺失衍生物转换为人类HT1080细胞，这些细胞仅表达层粘连蛋白β3和γ2链，并制备了重组LN5，缺乏每个G域。 G5或G4-5的缺失不会影响LN5的生物学活性，但是没有G3-5的LN5显示细胞粘合剂和迁移的活性很小。还发现G3包含两个不同的区域来刺激细胞粘合剂和迁移。此外，我们发现分泌后的G3和G4之间裂解了α3链，并确定了裂解位点。 2）LN5的105kDaγ2链在氨基末端区域切割，以产生105 kDa形式。我们发现MT1-MMP催化了这种加工，并且这种处理增加了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，γ2链的氨基末端区域对LN5的生物活性具有重要影响。 3）免疫组织化学分析表明，层粘连蛋白γ2链单体在入侵人类癌细胞中过表达。当将γ2链的氨基末端区域的cDNA转换为人类癌细胞系时，该细胞系在体内的侵入性潜力大大增强。这些结果表明，γ2链本身在肿瘤侵袭中起着一些重要作用。