The Signal Transduction of the Growth Hormone Receptor Is Regulated by the Ubiquitin/Proteasome System and Continues After Endocytosis

Abstract

The growth hormone receptor (GHR) intracellular domain contains all of the information required for signal transduction as well as for endocytosis. Previously, we showed that the proteasome mediates the clathrin-mediated endocytosis of the GHR. Here, we present evidence that the proteasomal inhibitor MG132 prolongs the GH-induced activity of both GHR and JAK2, presumably through stabilization of GHR and JAK2 tyrosine phosphorylation. If proteasomal inhibitor was combined with ligand in an endocytosis-deficient GHR mutant, the same phenomenon occurred indicating that proteasomal action on tyrosine dephosphorylation is independent of endocytosis. Experiments with a GHRtruncated tail mutant (GHR-(1-369)) led to a prolonged JAK2 phosphorylation caused by the loss of a phosphatase-binding site. This raised the question of what happens to the signal transduction of the GHR after its internalization. Co-immunoprecipitation of GH⅐GHR complexes before and after endocytosis showed that JAK2 as well as other activated proteins are bound to the GHR not only at the cell surface but also intracellularly, suggesting that the GHR signal transduction continues in endosomes. Additionally, these results provide evidence that GHR is present in endosomes both in its full-length and truncated form, indicating that the receptor is down-regulated by the proteasome.

Three Lysine Residues in the Common β Chain of the Interleukin-5 Receptor Are Required for Janus Kinase (JAK)-dependent Receptor Ubiquitination, Endocytosis, and Signaling

“…In conclusion, our data demonstrate that the Box 1 motif is only partially required for optimal ␤c ubiquitination, but fully required for JAK1/2 binding and ␤c tyrosine phosphorylation, as has been shown for other Type I receptors (Reviewed in 42,[43][44][45][46]. Furthermore, these findings support our earlier hypothesis that the presence of ␤c K1-3 (Lys 457 , Lys 461 , and Lys 467 ) is required for efficient ␤c ubiquitination and suggest that these residues possibly act cooperatively to promote this activity.…”

“…In general, it is well accepted that a canonical P-X-P motif in the Box 1 region of Type I and Type II cytokine receptors is necessary for JAK kinase binding because mutation of these proline residues leads to reduced binding and activation of the JAKs (37,(42)(43)(44)(45)(46). It was therefore quite unexpected to find that substitution of 3 lysine residues to arginine in the ␤c intracellular domain consistently resulted in severely impaired JAK kinase binding to the receptor even though the ␤c P-X-P motif was still present.…”

Crystal structure of the SOCS2–elongin C–elongin B complex defines a prototypical SOCS box ubiquitin ligase

“…Because of the key role of SOCS2 as a major negative regulator of GH signaling, the development of SOCS2 antagonists has been proposed as an alternative treatment regime to GH injections. The direct interaction between the SOCS box and elongin C constitutes such an intervention point, because the signal for GHR (as well as several other cytokine receptors) is prolonged in the presence of proteasome inhibitors (30). Indeed, we identify one potential site in proximity to the buried SOCS2 C terminus (Fig.…”

LEPROT and LEPROTL1 cooperatively decrease hepatic growth hormone action in mice

“…However, there is no evidence that GH-induced STAT signaling is only a direct consequence of cell-surface receptor abundance. Indeed, if we consider that (a) heterozygous Ghr-deficient mice (+/-) have normal plasma IGF1 levels despite a 50% decrease of Ghr mRNA expression and total GH binding in liver (19) and that (b) GH-induced STAT5 signaling lasts intracellularly after GHR endocytosis (28), then the duration of intracellular signaling may also be of crucial importance. For some receptors, endocytic organelles, as intracellular stations, may participate in signaling specificity and regulation (29).…”