Schwall R H, Chang L Y, Godowski P J, Kahn D W, Hillan K J, Bauer K D, Zioncheck T F
Genentech, Inc., South San Francisco, California 94080-4990, USA.
J Cell Biol. 1996 May;133(3):709-18. doi: 10.1083/jcb.133.3.709.
Hepatocyte growth factor (HGF) is a potent epithelial mitogen whose actions are mediated through its receptor, the proto-oncogene c-Met. Two truncated variants of HGF known as NK1 and NK2 have been reported to be competitive inhibitors of HGF binding to c-Met, and to function as HGF antagonists (Lokker, N.A., and P.J. Godowski. 1993. J. Biol. Chem. 268: 17145-17150; Chan, A.M., J.S. Rubin, D.P. Bottaro, D.W. Hirschfield, M. Chedid, and S.A. Aaronson. 1991. Science (Wash. DC). 254:1382-1387). We show here, however, that NK1 acts as a partial agonist in mink lung cells. Interestingly, NK1, which is an HGF antagonist in hepatocytes in normal conditions, was converted to a partial agonist by adding heparin to the culture medium. The interaction of NK1 and heparin was further studied in BaF3 cells, which express little or no cell surface heparan sulfate proteoglycans. In BaF3 cells transfected with a plasmid encoding human c-Met, heparin and NK1 synergized to stimulate DNA synthesis and cell proliferation. There was no effect of heparin on the IL-3 sensitivity of BaF3-hMet cells, and no effect of NK1 plus heparin in control BaF3 cells, indicating that the response was specific and mediated through c-Met. The naturally occurring HGF splice variant NK2 also stimulated DNA synthesis in mink lung cells and exerted a heparin-dependent effect on BaF3-hMet cells, but not on BaF3-neo cells. The activating effect of heparin was mimicked by a variety of sulfated glycosaminoglycans. Mechanistic studies revealed that heparin increased the binding of NK1 to BaF3-hMet cells, stabilized NK1, and induced dimerization of NK1. Based on these studies, we propose that the normal agonist activity of NK1 and NK2 in mink lung cells is due to an activating interaction with an endogenous glycosaminoglycan. Consistent with that model, a large portion of the NK1 binding to mink lung cells could be blocked by heparin. Moreover, a preparation of glycosaminoglycans from the surface of mink lung cells induced dimerization of NK1. These data show that the activity of NK1 and NK2 can be modulated by heparin and other related glycosaminoglycans to induce proliferation in cells expressing c-Met.
肝细胞生长因子（HGF）是一种强效的上皮细胞有丝分裂原，其作用通过其受体原癌基因c-Met介导。据报道，两种被称为NK1和NK2的HGF截短变体是HGF与c-Met结合的竞争性抑制剂，并作为HGF拮抗剂发挥作用（洛克，N.A.，和P.J.戈多夫斯基。1993。《生物化学杂志》268:17145 - 17150；陈，A.M.，J.S.鲁宾，D.P.博塔罗，D.W.赫希菲尔德，M.切迪，和S.A.阿伦森。1991。《科学》（华盛顿特区）。254:1382 - 1387）。然而，我们在此表明，NK1在貂肺细胞中作为部分激动剂起作用。有趣的是，在正常条件下作为肝细胞中HGF拮抗剂的NK1，通过向培养基中添加肝素而转变为部分激动剂。在几乎不表达或不表达细胞表面硫酸乙酰肝素蛋白聚糖的BaF3细胞中，对NK1与肝素的相互作用进行了进一步研究。在用编码人c-Met的质粒转染的BaF3细胞中，肝素和NK1协同刺激DNA合成和细胞增殖。肝素对BaF3-hMet细胞的IL-3敏感性没有影响，NK1加肝素对对照BaF3细胞也没有影响，这表明该反应是特异性的，并且通过c-Met介导。天然存在的HGF剪接变体NK2也刺激貂肺细胞中的DNA合成，并对BaF3-hMet细胞产生肝素依赖性作用，但对BaF3-neo细胞没有作用。多种硫酸化糖胺聚糖模拟了肝素的激活作用。机制研究表明，肝素增加了NK1与BaF3-hMet细胞的结合，稳定了NK1，并诱导了NK1的二聚化。基于这些研究，我们提出NK1和NK2在貂肺细胞中的正常激动剂活性是由于与内源性糖胺聚糖的激活相互作用。与该模型一致，与貂肺细胞结合的大部分NK1可被肝素阻断。此外，从貂肺细胞表面制备的糖胺聚糖诱导了NK1的二聚化。这些数据表明，NK1和NK2的活性可被肝素和其他相关糖胺聚糖调节，以诱导表达c-Met的细胞增殖。
