Growth Hormone-promoted Tyrosyl Phosphorylation of SHC Proteins and SHC Association with Grb2

Growth hormone (GH) promotes signaling by causing activation of the non-receptor tyrosine kinase, JAK2, which associates with the GH receptor. GH causes phosphorylation of epidermal growth factor receptor (EGFR; ErbB-1) and its family member, ErbB-2. For EGFR, JAK2-mediated GH-induced tyrosine phosphorylation may allow EGFR to serve as a scaffold for GH signaling. For ErbB-2, GH induces serine/threonine phosphorylation that dampens basal and EGF-induced ErbB-2 kinase activation. We now further explore GH-induced EGFR phosphorylation in 3T3-F442A, a preadipocytic fibroblast cell line that expresses endogenous GH receptor, EGFR, and ErbB-2. Using a monoclonal antibody that recognizes ERK consensus site phosphorylation (PTP101), we found that GH caused PTP101-reactive phosphorylation of EGFR. This GH-induced EGFR phosphorylation was prevented by MEK1 inhibitors but not by a protein kinase C inhibitor. Although GH did not discernibly affect EGF-induced EGFR tyrosine phosphorylation, we observed by immunoblotting a substantial decrease of EGF-induced EGFR degradation in the presence of GH. Fluorescence microscopy studies indicated that EGF-induced intracellular redistribution of an EGFR-cyan fluorescent protein chimera was markedly reduced by GH cotreatment, in support of the immunoblotting results. Notably, protection from EGF-induced degradation and inhibition of EGF-induced intracellular redistribution afforded by GH were both prevented by a MEK1 inhibitor, suggesting a role for GH-induced ERK activation in regulating the trafficking itinerary of the EGF-stimulated EGFR. Finally, we observed augmentation of early aspects of EGF signaling (EGF-induced ERK2 activation and EGF-induced Cbl tyrosine phosphorylation) by GH cotreatment; the GH effect on EGF-induced Cbl tyrosine phosphorylation was also prevented by MEK1 inhibition. These data indicate that GH, by activating ERKs, can modulate EGF-induced EGFR trafficking and signaling and expand our understanding of mechanisms of cross-talk between the GH and EGF signaling systems.

Section: Growth Hormone (Gh)mentioning

confidence: 95%

Growth Hormone-induced Phosphorylation of Epidermal Growth Factor (EGF) Receptor in 3T3-F442A Cells

Huang

Kim

Jiang

et al. 2003

“…SIRPα1 also contains a proline-rich motif that conforms to the proline-rich box 1 region of cytokine receptors. The proline-rich region in the cytokine receptors mediates the association between the JAK kinases and the cytokine receptors (23,24,28). Deletion of the proline-rich region of SIRPα1 or proline to alanine mutation of conserved prolines in this region had no effect upon the association of JAK2 with SIRPα1.…”

Section: Discussionmentioning

confidence: 99%

Negative Regulation of Growth Hormone Receptor/JAK2 Signaling by Signal Regulatory Protein α

Stofega

Argetsinger²,

Wang³

et al. 2000

Self Cite

SUMMARYSIRPs (signal regulatory proteins) are receptor-like transmembrane proteins, the majority of which contain a cytoplasmic proline-rich region and four cytoplasmic tyrosines that, when phosphorylated, bind SH2 domain-containing protein tyrosine phosphatases (SHP). We Consistent with reduced JAK2 activity, overexpression of wild-type SIRPα1 but not SIRP 4YF reduces GH-induced phosphorylation of ERKs 1 and 2, STAT3 and STAT5B. These results suggest that SIRPα1 is a negative regulator of GH signaling and that the ability of SIRPα1

“…Thus, a single tyrosine module is sufficient for STAT activation, as already described by Behrmann et al (52), even if multiple tyrosine residues in the cytosolic domains can also promote Jak/STAT activation (49 -51,76). This Tyr 157 is present in an original motif, EKV-VYVGVW, different from the two main structural motifs, YXXQ and GXGYM, previously identified (48,(77)(78)(79)(80)(81)(82)(83)(84) but which is a predicted tyrosine phosphorylation site (score 0.822) using the NetPhos 2.0 server. It is worth noting that Tyr 157 is the only Tyr residue in CXCR4 located toward the cell cytoplasm present in a consensus motif of phosphorylation.…”

Section: Discussionmentioning

confidence: 99%

Identification of the Cytoplasmic Domains of CXCR4 Involved in Jak2 and STAT3 Phosphorylation

Ahr

Denizot

Robert-Hebmann

et al. 2005

The chemokine SDF-1␣ transduces G i -dependent and -independent signals through CXCR4. Activation of Jak2/STAT3, a G i -independent signaling pathway, which plays a major role in survival signals, is known to be activated after SDF-1␣ binding to CXCR4 but the domains of CXCR4 involved in this signaling remain unexplored. Using human embryonic kidney HEK-293 cells stably expressing wild-type or mutated forms of CXCR4, we demonstrated that STAT3 phosphorylation requires the N-terminal part of the third intracellular loop (ICL3) and the tyrosine 157 present at the end of the second intracellular loop (ICL2) of CXCR4. In contrast, neither the conserved Tyr 135 in the DRY motif at the N terminus of ICL2 nor the Tyr 65 and Tyr 76 in the first intracellular loop (ICL1) are involved in this activation. ICL3, which does not contain any tyrosine residues, is needed to activate Jak2. These results demonstrate that two separate domains of CXCR4 are involved in Jak2/ STAT3 signaling. The N-terminal part of ICL3 is needed to activate Jak2 after SDF-1␣ binding to CXCR4, leading to phosphorylation of only one cytoplasmic Tyr, present at the C terminus of ICL2, which triggers STAT3 activation. This work has profound implications for the understanding of CXCR4-transduced signaling.