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Endocrine Abstracts (2020) 70 AEP781 | DOI: 10.1530/endoabs.70.AEP781

ECE2020 Audio ePoster Presentations Reproductive and Developmental Endocrinology (79 abstracts)

Luteinizing hormone/choriogonadotropin receptor (LHCGR) mediates different kinetics of G proteins, β-arrestins and cAMP activation

Elia Paradiso 1,2 , Riccardo Benevelli 1 , Clara Lazzaretti 1,2 , Samantha Sperduti 1,3 , Giulia Brigante 1,4 , Manuela Simoni 1,3,4,5 & Livio Casarini 1,3

1University of Modena and Reggio Emilia, Unit of Endocrinology, Department of Biomedical, Metabolic and Neural Sciences, Modena, Italy; 2University of Modena and Reggio Emilia, International PhD School in Clinical and Experimental Medicine (CEM), Modena, Italy; 3University of Modena and Reggio Emilia, Center for Genomic Research , Modena, Italy; 4Azienda Ospedaliero-Universitaria di Modena. Baggiovara Hospital, Department of Medical Specialties, Modena, Italy; 5Institut National de la Recherche Agronomique, Unitè Physiologie de la Reproduction et des Comportements, Nouzilly, France

Pituitary luteinizing hormone (LH) and placental human choriogonadotropin (hCG) are two heterodimeric glycoprotein hormones regulating reproduction. They bind the same receptor (LHCGR) expressed in gonadal cells, activating hormone-specific G protein- and β-arrestins-dependent signaling cascades, before LHCGR internalization. LH induces preferential proliferative signals, while hCG activates mainly the steroidogenic pathway, reflecting their physiological roles. In this study,we compared the kinetics of LH- versus hCG-mediatedG proteins and β-arrestin 2 activation in vitro. The HEK293 cell line was transiently transfected with the LHCGR-encoding plasmid, together with that encoding G proteins- or β-arrestin 2-tagged bioluminescence resonance energy transfer (BRET) biosensors. Cells were treated by various hCG and LH concentrations (pM-nM range) and the activation of Gαs, Gαq, Gαi protein and β-arrestins was evaluated by BRET, under native conditions or after inhibition of LHCGR internalization by Dynasore. We demonstrated that LH/hCG binding to LHCGR mediates the activation of G proteins and recruitment of β-arrestin 2 in a hormone-specific manner. LHCGR-Gαs protein interaction increased upon 10 nM hCG, but not LH treatment (one-way ANOVA; P < 0.05; n = 8), reflecting the previously demonstrated preferential activation of intracellular cAMP signaling by the placental hormone [doi:10.1210/er.2018-00065]. These data are confirmed by kinetics analysis intracellular cAMP increase over 120-min, which achieved higher levels upon treatment by hCG than LH when hormones are administrated to cells at the 50% effective concentration (EC50; hCG = 100 pM; LH = 500 pM; Kruskal-Wallis test, P < 0.05; n = 4). On the other hand, LH was more potent than hCG in promoting LHCGR-Gαi protein and LHCGR-β-arrestin 2 interactions, thus confirming preferential activation of proliferative signals (means ± SEM one-way ANOVA; P < 0.05; n = 8) [doi:10.1210/er.2018-00065]. Interestingly, LH and hCG mediated similar kinetics of intracellular cAMP increase between 30–120 min, when LHCGR internalization is blocked by Dynasore, while short-term (0-30) min hCG-induced cAMP levels are anyway higher than those LH-mediated (Kruskal-Wallis test, P < 0.05; n = 4). These results suggest that endosomal compartmentalization of LHCGR is involved in the differentiation of the long-term LH/hCG-induced cAMP signaling.

We have demonstrated that LH and hCG drive LHCGR in hormone-specific interaction with Gαs and i proteins, as well as β-arrestin 2 and endosomal compartments, and resulting in different intracellular signaling patterns.

Volume 70

22nd European Congress of Endocrinology

05 Sep 2020 - 09 Sep 2020

European Society of Endocrinology 

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