Grb10 Identified as a Potential Regulator of Growth Hormone (GH) Signaling by Cloning of GH Receptor Target Proteins

Abstract

The cloning of receptor targets procedure, used so far to identify proteins associated with tyrosine kinase receptors was modified to clone SH2 proteins able to bind to the growth hormone receptor (GHR). The cytoplasmic region of GHR, a member of the cytokine receptor superfamily does not contain tyrosine kinase activity. It was thus phosphorylated in bacteria by the Elk tyrosine kinase and radiolabeled to screen a mouse expression library. With this probe, we identified Shc and the p85 subunit of phosphatidylinositol 3-kinase as direct targets of the receptor. The other proteins identified, Csk, Shb, Grb4, and Grb10 are new potential transducers for cytokine receptors. We show in Huh-7 hepatoma cells that Grb10 and GHR associate under GH stimulation. Co-transfections in 293 cells further show that Grb10 interacts with both the GHR and Jak2. Functional tests demonstrate that Grb10 inhibits transcription of two reporter genes containing, respectively, the serum response element of c-fos and the GH response element 2 of the Spi2.1 gene, whereas it has no effect on a reporter gene containing only Stat5 binding elements. Our results suggest that Grb10 is a new target for a member of the cytokine receptor family that down-regulates some GH signaling pathways downstream of Jak2 and independently of Stat5.

Evidence for an interaction between the insulin receptor and Grb7. A role for two of its binding domains, PIR and SH2

“…In addition, Grb10 and Grb7 can interact with other families of proteins. Cytokines receptors like the GH receptor, the tyrosine kinases Jak2 and Tec, and the kinases MEK and Raf were found in association with Grb10 (Mano et al, 1998;Moutoussamy et al, 1998;Nantel et al, 1998). After EGF or PDGF stimulation Grb7 is associated with Shc, another molecular adapter, or with the tyrosine phosphatase SHP2 (Keegan et al, 1994;Stein et al, 1994;Yokote et al, 1996).…”

The Grb7 family proteins: structure, interactions with other signaling molecules and potential cellular functions

“…Grb10 has also been shown to associate with several proteins including the EGF receptor (Frantz et al, 1997;He et al, 1998;Ooi et al, 1995), IR (Hansen et al, 1996;Liu and Roth, 1995;Dong et al, 1997;Frantz et al, 1997;O'Neill et al, 1996), Ret (Pandey et al, 1995), IGFR (Dey et al, 1996;He et al, 1998;Morrione et al, 1996;O'Neill et al, 1996), Elk/EphB1 (Stein et al, 1996), PDGF receptor (Frantz et al, 1997), Tek/Tie (Mano et al, 1998), Raf/MEK1 (Nantel et al, 1998), Nedd4 (Morrione et al, 1999), BCR-ABL (Bai et al, 1998), and GH receptor (Moutoussamy et al, 1998). In contrast, Grb14 has been reported to have fewer binding partners including EGFR/PDGFR (Daly et al, 1996), IR (Kasus-Jacobi et al, 1998), Tek/ Tie2 (Jones et al, 1999), FGF receptor (Reilly et al, 2000).…”

The adapter protein, Grb10, is a positive regulator of vascular endothelial growth factor signaling

“…Although Grb10 interacts with tyrosine kinase receptors such as PDGF, EGF, insulin and IGF-I receptors, its function as an adapter molecule remains to be discovered (He et al, 1998;Stein et al, 1996). Indeed, Grb10 binds to signaling molecules such as Raf1, MEK1, BCR-Abl, Jak2, NEDD4 (Nantel et al, 1998;Bai et al, 1998;Moutoussamy et al, 1998;Morrione et al, 1999), but the consequences of these associations have not been elucidated. The role of Grb10 in cellular signaling is controversial.…”