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Oxytocin Induces Dephosphorylation of Eukaryotic Elongation Factor 2 in Human Myometrial Cells

Departments of Medicine (D.D., M.G., M.-E.C., C.R., H.H.Z.), Pharmacology and Therapeutics (D.D., M.-E.C., H.H.Z.), and Obstetrics and Gynecology (H.H.Z.), McGill University, Montreal, Quebec, Canada H3A 1A1

Address all correspondence and requests for reprints to: Hans H. Zingg, M.D., Ph.D., Laboratory of Molecular Endocrinology, Royal Victoria Hospital, Room H7.63, 687 Pine Avenue West, Montreal, Quebec, Canada H3A 1A1. E-mail: hans.zingg{at}mcgill.ca.

The oxytocin (OT) receptor (OTR) mediates a wide spectrum of biological actions and is expressed in a large number of different tissues, including uterine, breast, and lung tumors. To define more completely the intracellular signaling mechanisms linked to OTR activation, we have used a phosphoproteomics approach and have characterized changes in the phosphorylation states of intracellular proteins in response to OTR activation in OTR-expressing cell lines. Using a specific antiphosphothreonine antibody, we observed several distinct changes in the threonine phosphorylation patterns. The most prominent change involved dephosphorylation of a 95-kDa moiety. Purification by ion exchange chromatography combined with one- and two-dimensional polyacrylamide gel electrophoresis followed by N-terminal micro-sequence analysis revealed that the 95-kDa moiety corresponded to eukaryotic elongation factor 2. This protein is a key regulator of cellular protein synthesis and mediates, upon dephosphorylation, the translocation step of peptide chain elongation. Dose-response curves in myometrial cells expressing the endogenous OTR indicated a significant effect of OT on eukaryotic elongation factor 2 dephosphorylation at 1 nM, a concentration close to the dissociation constant (K_d) of OT. Time course analysis indicates that the effect is rapid with a significant effect occurring at 5 min. To determine directly the effect of OT on protein synthesis, the incorporation of [³⁵S]Met into total protein was assessed. In myometrial cells, OTR activation led to significant 29% increase in total protein synthesis over a 2-h period. These findings establish a novel link between OTR activation and cellular protein synthesis and thus define a mechanism by which OT assumes a so far unrecognized, physiologically relevant trophic function.

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				D. Devost, M.-E. Carrier, and H. H. Zingg Oxytocin-Induced Activation of Eukaryotic Elongation Factor 2 in Myometrial Cells Is Mediated by Protein Kinase C Endocrinology, January 1, 2008; 149(1): 131 - 138. [Abstract] [Full Text] [PDF]