Assessment of cell line competence for studies of pharmacological GPR30 modulation.
J Recept Signal Transduct Res
; 37(2): 181-188, 2017 Apr.
Article
de En
| MEDLINE
| ID: mdl-27401115
ABSTRACT
CONTEXT/OBJECTIVE:
Cell lines used to study the role of the G protein-coupled receptor 30 (GPR30) or G protein-coupled estrogen receptor (GPER) as a mediator of estrogen responses have yielded conflicting results. This work identified a simple assay to predict cell line competence for pharmacological studies of GPR30. MATERIALS ANDMETHODS:
The phosphorylation or expression levels of ERK1/2, Akt, c-Fos and eNOS were evaluated to assess GPR30 activation in response to known agonists (17ß-estradiol and G-1) in MCF-7 and T-47D breast cancer cell lines and in bovine aortic endothelial cells. GPR30 expression was analyzed by qRT-PCR and Western blot with two distinct antibodies directed at its carboxy and amino terminals.RESULTS:
None of the agonists, at any of the concentrations tested, activated any of those target proteins. Additional experiments excluded the disruption of the signaling pathway, interference of phenol red in the culture medium and constitutive proteasome degradation of GPR30 as possible causes for the lack of response of the three cell lines. Analysis of receptor expression showed the absence of clearly detectable GPR30 species of 44 and 50-55 kDa previously identified in cell lines that respond to 17ß-estradiol and G-1. DISCUSSION ANDCONCLUSION:
Cells that do not express the 44 and 50-55 kDa species do not respond to GPR30 agonists. Thus, the presence or absence of these GPR30 species is a simple and rapid manner to determine whether a given cell line is suitable for pharmacological or molecular studies of GPR30 modulation.Mots clés
Texte intégral:
1
Collection:
01-internacional
Base de données:
MEDLINE
Sujet principal:
Tumeurs du sein
/
Récepteurs des oestrogènes
/
Récepteurs couplés aux protéines G
/
Récepteur alpha des oestrogènes
/
Oestrogènes
Limites:
Animals
/
Female
/
Humans
Langue:
En
Journal:
J Recept Signal Transduct Res
Sujet du journal:
BIOQUIMICA
/
FISIOLOGIA
Année:
2017
Type de document:
Article
Pays d'affiliation:
Portugal