TY - JOUR
T1 - Oxytocin induces dephosphorylation of eukaryotic elongation factor 2 in human myometrial cells
AU - Devost, Dominic
AU - Girotti, Milena
AU - Carrier, Marie Eve
AU - Russo, Caterina
AU - Zingg, Hans H.
PY - 2005/5
Y1 - 2005/5
N2 - 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-kBa 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-kBa 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 (Kd) 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.
AB - 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-kBa 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-kBa 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 (Kd) 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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U2 - 10.1210/en.2004-1428
DO - 10.1210/en.2004-1428
M3 - Article
C2 - 15661857
AN - SCOPUS:17744380025
SN - 0013-7227
VL - 146
SP - 2265
EP - 2270
JO - Endocrinology
JF - Endocrinology
IS - 5
ER -