Parathyroid hormone receptor trafficking contributes to the activation of extracellular signal-regulated kinases but is not required for regulation of cAMP signaling.
Agonist-mediated activation of the type 1 parathyroid hormone receptor (PTH1R) results in several signaling events and receptor endocytosis. It is well documented that arrestins contribute to desensitization of both G(s)- and G(q)-mediated signaling and mediate PTH1R internalization. However, whether PTH1R trafficking directly contributes to signaling remains unclear. To address this question, we investigated the role of PTH1R trafficking in cAMP signaling and activation of extracellular signal-regulated kinases ERK1/2 in HEK-293 cells. Dominant negative forms of dynamin (K44A-dynamin) and beta-arrestin1 (beta-arrestin1-(319-418)) abrogated PTH1R internalization but had no effect on cAMP signaling; neither acute cAMP production by PTH nor desensitization and resensitization of cAMP signaling were affected. Therefore, PTH1R trafficking is not necessary for regulation of cAMP signaling. PTH-(1-34) induced rapid and robust activation of ERK1/2. A PTHrP-based analog ([p-benzoylphenylalanine1, Ile5,Arg(11,13),Tyr36]PTHrP-(1-36)NH2), which selectively activates the G(s)/cAMP pathway without inducing PTH1R endocytosis, failed to stimulate ERK1/2 activity. Inhibition of PTH1R endocytosis by K44A-dynamin dampened ERK1/2 activation in response to PTH-(1-34) by 69%. Incubation with the epidermal growth factor receptor inhibitor AG1478 reduced ERK1/2 phosphorylation further. In addition, ERK1/2 phosphorylation occurred following internalization of a PTH1R mutant induced by PTH-(7-34) in the absence of G protein signaling. Collectively, these data indicate that PTH1R trafficking and G(q) (but not G(s)) signaling independently contribute to ERK1/2 activation, predominantly via transactivation of the epidermal growth factor receptor.
Syme
Colin A
CA
Division of Endocrinology and Metabolism, Department of Medicine, Univesity of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA.
Friedman
Peter A
PA
Bisello
Alessandro
A
D019390:Arrestins / Q000502:physiology; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism*; D004705:Endocytosis; D004789:Enzyme Activation; D066246:ErbB Receptors / Q000502:physiology; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000502:physiology; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D010766:Phosphorylation; D044168:Receptor, Parathyroid Hormone, Type 1 / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*; D015533:Transcriptional Activation; D000071557:beta-Arrestins
D019390:Arrestins; D044168:Receptor, Parathyroid Hormone, Type 1; D000071557:beta-Arrestins; D000242:Cyclic AMP; D066246:ErbB Receptors; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D043802:GTP-Binding Protein alpha Subunits, Gq-G11