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Figure 7 | BMC Cell Biology

Figure 7

From: A novel link between the proteasome pathway and the signal transduction pathway of the Bone Morphogenetic Proteins (BMPs)

Figure 7

A novel Smad1 Nuclear Interacting Protein (SNIP1) is recruited to interact with Az in a BMP receptor activation-dependent fashion. A. SNIP1 is a nuclear protein. COS1 cells were transfected with T7-tagged SNIP1. Anti-T7 monoclonal antibody and FITC conjugated anti-mouse secondary antibody was used to detect SNIP1. B. SNIP1 specifically interacts with Smad1, Az and HsN3 but not with ODC in the yeast two-hybrid system. The panels labeled as "1" and "2" represent yeast transformants on U-H-W- galactose or glucose plates, respectively. B42 fusion proteins were only induced in yeast grown on galactose plates. C. Domain mapping of Smad1 interaction with SNIP1 in yeast. "Smad1GS" represents the mutant Smad1 (G419S) [40]. Smad1NL contains the MH1 and the linker region; Smad1L contains only the linker region. D. SNIP1 interacts with the pro-sequence-containing form of HsN3 but not with Az in COS cells upon coexpression. COS cells were transfected with the indicated plasmids. Cells were metabolically labeled with 35S-methionine for the last 4 hours before lysates were made for immunoprecipitation. Az is marked by an asterisk in lane 6, while the two forms of HsN3 are marked as in previous Figures. E. SNIP1 interacts with Az upon its co-expression with Smad1, Smad4 and the BMP type I receptor activation enhances the interaction. COS cells were transfected with 2 μg each of the indicated plasmids. The anti-T7 immunoprecipitates were analyzed by Western blot with monoclonal anti-Flag (top panel). The co-precipitated F-Az is indicated (lane 3). Protein expression was monitored by Western blot analyses of total lysates as shown in bottom panels.

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