Abstract
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.
Highlights
Ras is localized at the cytoplasmic surface of the cell membrane and has two interconvertible forms: GDP-bound inactive and GTP-bound active forms (4 – 6)
We have demonstrated that the SPRY domain of RanBP9 and RanBP10 is involved in the interaction between these two proteins and the MET tyrosine kinase domain [16, 17]
We have demonstrated in our previous studies that the SPRY domains in RanBP9 and RanBP10 are responsible for their interactions with the tyrosine kinase domain of MET [16, 17]
Summary
Ras is localized at the cytoplasmic surface of the cell membrane and has two interconvertible forms: GDP-bound inactive and GTP-bound active forms (4 – 6). In all of these proteins, the SOCS box is located in their C termini, and other different domain structures are located in their middle or N-terminal regions [14] Some of these subfamilies are components of or function as adaptors for a large family of E3 ligases to regulate certain signaling pathways [15], the function of certain members, such as the SSB subfamily, remains to be determined. Because the SPRY domains of the SSB subfamily show significant amino acid sequence similarity with RanBP9 and RanBP10, we investigated whether these SPRY domain-containing SOCS box proteins played a role in the regulation of MET signaling pathways. These results suggest that SSB-1 is an important regulator in the HGF-MET signaling
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