Hepatocyte growth factor activator inhibitor-2 prevents shedding of matriptase.
Hepatocyte growth factor activator inhibitor-2 prevents shedding of matriptase.
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DOI：
10.1016/j.yexcr.2013.01.008
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发表时间：
2013-04-01
影响因子：
3.7
通讯作者：
Vogel LK
中科院分区：
文献类型：
--
作者：
Larsen BR;Steffensen SD;Nielsen NV;Friis S;Godiksen S;Bornholdt J;Soendergaard C;Nonboe AW;Andersen MN;Poulsen SS;Szabo R;Bugge TH;Lin CY;Skovbjerg H;Jensen JK;Vogel LK
肝细胞生长因子激活因子抑制剂-2 （HAI-2）在体外是多种蛋白酶的抑制剂，包括膜结合丝氨酸蛋白酶、基质酶。对敲除小鼠的研究表明，HAI-2仅在表达基质酶的小鼠中对胎盘发育至关重要，这表明HAI-2对基质酶的调节很重要。先前的研究表明，除非与另一种HAI HAI-1共表达，否则基质酶的重组表达不成功。在本研究中，我们发现在犬细胞系MDCK中单独重组表达人基质酶时，可以检测到人基质酶mRNA，并将人基质酶外畴脱落到培养基中，表明单独表达的基质酶通过分泌途径快速转运并脱落。而基质酶与HAI-1或HAI-2一起表达，在激活的质膜上积累，通过Arg614的裂解和细胞提取物的肽解活性增加来判断。Kunitz结构域1的突变而不是Kunitz结构域2的突变使HAI-2的这一功能消失。HAI-2似乎在细胞内发挥其功能，因为绝大多数HAI-2位于细胞内，而且在基质酶所在的基侧质膜上无法检测到HAI-2。然而，在顶端质膜上可以检测到少量未发生内吞作用的HAI-2。我们的研究结果表明，HAI-2的Kunitz结构域1导致基质酶以膜结合形式积聚在基底外侧质膜上。
Hepatocyte growth factor activator inhibitor-2 (HAI-2) is an inhibitor of many proteases in vitro, including the membrane-bound serine protease, matriptase. Studies of knock-out mice have shown that HAI-2 is essential for placental development only in mice expressing matriptase, suggesting that HAI-2 is important for regulation of matriptase. Previous studies have shown that recombinant expression of matriptase was unsuccessful unless co-expressed with another HAI, HAI-1. In the present study we show that when human matriptase is recombinantly expressed alone in the canine cell line MDCK, then human matriptase mRNA can be detected and the human matriptase ectodomain is shed to the media, suggesting that matriptase expressed alone is rapidly transported through the secretory pathway and shed. Whereas matriptase expressed together with HAI-1 or HAI-2 accumulates on the plasma membrane where it is activated, as judged by cleavage at Arg614 and increased peptidolytic activity of the cell extracts. Mutagenesis of Kunitz domain 1 but not Kunitz domain 2 abolished this function of HAI-2. HAI-2 seems to carry out its function intracellularly as this is where the vast majority of HAI-2 is located and since HAI-2 could not be detected on the basolateral plasma membrane where matriptase resides. However, minor amounts of HAI-2 not undergoing endocytosis could be detected on the apical plasma membrane. Our results suggest that Kunitz domain 1 of HAI-2 cause matriptase to accumulate in a membrane-bound form on the basolateral plasma membrane.