Wang Dakun, Li Zaibo, Messing Edward M, Wu Guan
Department of Urology, Department of Pathology and Laboratory Medicine, and The James P. Wilmot Cancer Center, University of Rochester Medical Center, Rochester, New York 14642, USA.
J Biol Chem. 2005 Apr 22;280(16):16393-401. doi: 10.1074/jbc.M413897200. Epub 2005 Feb 15.
The suppressor of cytokine signaling (SOCS) protein family includes a SPRY (repeats in splA and RyR) domain-containing SOCS box protein (SSB) subfamily, which consists of four members, SSB-1, SSB-2, SSB-3, and SSB-4. These proteins contain a central SPRY domain and a C-terminal SOCS box. Although some of the SOCS protein subfamilies function as adaptors for a large family of ubiquitin-protein isopeptide ligases to regulate certain signaling pathways, the function of the SSB subfamily remains to be determined. In our previous studies, we have found that two SPRY domain-containing proteins, RanBP9 and RanBP10, interact with MET through the SPRY domain. In the present study, we explored the function of SSB proteins in the regulation of the hepatocyte growth factor (HGF)-MET signaling. Our results showed that all four SSB proteins also interacted with the MET. The MET interaction with SSB-1 was further investigated. We demonstrated that SSB-1 bound to MET tyrosine kinase domain through its SPRY domain. MET interacted with SSB-1 in both the absence and the presence of HGF, but HGF treatment resulted in the recruitment of more SSB-1 by MET. We showed that overexpression of SSB-1 but not other SSB proteins enhanced the HGF-induced serum response element (SRE)-luciferase activity. Overexpression of SSB-1 exhibited no effect on the basal level or epidermal growth factor-induced SRE-luciferase activity. SSB-1 also enhanced HGF-induced Erk phosphorylation. Suppression of SSB-1 by the RNA interference method down-regulated HGF-induced SRE-luciferase activity and decreased Elk-1 activation. These results suggest that SSB-1 may play an important role in enhancing the HGF-induced Erk-Elk-1-SRE pathway. Furthermore, we demonstrated that in response to HGF stimulation, the SSB-1 protein became phosphorylated at tyrosine residue 31. The phosphorylated SSB-1 protein bound to p120Ras-GTPase-activating protein (GAP) but did not promote the degradation of p120RasGAP, indicating that enhanced HGF-MET signaling by overexpression of SSB-1 was not dependent on p120RasGAP degradation.
细胞因子信号转导抑制蛋白（SOCS）家族包括一个含SPRY（splA和RyR重复序列）结构域的SOCS盒蛋白（SSB）亚家族，该亚家族由四个成员组成，即SSB-1、SSB-2、SSB-3和SSB-4。这些蛋白含有一个中央SPRY结构域和一个C端SOCS盒。尽管一些SOCS蛋白亚家族作为一大类泛素-蛋白异肽连接酶的衔接子发挥作用，以调节某些信号通路，但SSB亚家族的功能仍有待确定。在我们之前的研究中，我们发现两种含SPRY结构域的蛋白RanBP9和RanBP10通过SPRY结构域与MET相互作用。在本研究中，我们探究了SSB蛋白在肝细胞生长因子（HGF）-MET信号转导调节中的功能。我们的结果表明，所有四种SSB蛋白也与MET相互作用。我们进一步研究了MET与SSB-1的相互作用。我们证明SSB-1通过其SPRY结构域与MET酪氨酸激酶结构域结合。无论有无HGF，MET都与SSB-1相互作用，但HGF处理导致MET募集更多的SSB-1。我们发现，过表达SSB-1而非其他SSB蛋白可增强HGF诱导的血清反应元件（SRE）-荧光素酶活性。过表达SSB-1对基础水平或表皮生长因子诱导的SRE-荧光素酶活性无影响。SSB-1还增强了HGF诱导的Erk磷酸化。通过RNA干扰方法抑制SSB-1可下调HGF诱导的SRE-荧光素酶活性并降低Elk-1的激活。这些结果表明，SSB-1可能在增强HGF诱导的Erk-Elk-1-SRE途径中发挥重要作用。此外，我们证明，在HGF刺激下，SSB-1蛋白在酪氨酸残基31处发生磷酸化。磷酸化的SSB-1蛋白与p120Ras-鸟苷酸酶激活蛋白（GAP）结合，但不促进p120RasGAP的降解，这表明过表达SSB-1增强HGF-MET信号转导不依赖于p120RasGAP的降解。
