Jin M H, Sawamoto K, Ito M, Okano H
Division of Neuroanatomy, Department of Neuroscience, Biomedical Research Center, Osaka University Graduate School of Medicine, Osaka, Japan.
Mol Cell Biol. 2000 Mar;20(6):2098-107. doi: 10.1128/MCB.20.6.2098-2107.2000.
Drosophila Argos (Aos), a secreted protein with an epidermal growth factor (EGF)-like domain, has been shown to inhibit the activation of the Drosophila EGF receptor (DER). However, it has not been determined whether Aos binds directly to DER or whether regulation of the DER activation occurs through some other mechanism. Using DER-expressing cells (DER/S2) and a recombinant DER extracellular domain-Fc fusion protein (DER-Fc), we have shown that Aos binds directly to the extracellular domain of DER with its carboxyl-terminal region, including the EGF-like domain. Furthermore, Aos can block the binding of secreted Spitz (sSpi), a transforming growth factor alpha-like ligand of DER, to the extracellular domain of DER. We observed that sSpi stimulates the dimerization of both the soluble DER extracellular domain (sDER) and the intact DER in the DER/S2 cells and that Aos can block the sSpi-induced dimerization of both sDER and intact DER. Moreover, we have shown that, by directly interacting with DER, Aos and SpiAos (a chimeric protein that is composed of the N-terminal region of Spi and the C-terminal region of Aos) inhibit the dimerization and phosphorylation of DER that are induced by DER's overexpression in the absence of sSpi. These results indicate that Aos exerts its inhibitory function through dual molecular mechanisms: by blocking both the receptor dimerization and the binding of activating ligand to the receptor. This is the first description of this novel inhibitory mechanism for receptor tyrosine kinases.
果蝇的Argos（Aos）是一种具有表皮生长因子（EGF）样结构域的分泌蛋白，已被证明可抑制果蝇EGF受体（DER）的激活。然而，尚未确定Aos是否直接与DER结合，或者DER激活的调节是否通过其他机制发生。利用表达DER的细胞（DER/S2）和重组DER细胞外结构域-Fc融合蛋白（DER-Fc），我们发现Aos通过其羧基末端区域，包括EGF样结构域，直接与DER的细胞外结构域结合。此外，Aos可阻断分泌型Spitz（sSpi）与DER细胞外结构域的结合，sSpi是DER的一种转化生长因子α样配体。我们观察到sSpi刺激了DER/S2细胞中可溶性DER细胞外结构域（sDER）和完整DER的二聚化，并且Aos可阻断sSpi诱导的sDER和完整DER的二聚化。此外，我们还表明，通过直接与DER相互作用，Aos和SpiAos（一种由Spi的N末端区域和Aos的C末端区域组成的嵌合蛋白）抑制了在没有sSpi的情况下DER过表达诱导的DER二聚化和磷酸化。这些结果表明，Aos通过双重分子机制发挥其抑制功能：通过阻断受体二聚化和激活配体与受体的结合。这是对受体酪氨酸激酶这种新型抑制机制的首次描述。
