TY - JOUR
T1 - Estrogen- and xenoestrogen-induced ERK signaling in pituitary tumor cells involves estrogen receptor-α interactions with G protein-αi and caveolin I
AU - Watson, Cheryl S.
AU - Jeng, Yow Jiun
AU - Hu, Guangzhen
AU - Wozniak, Ann
AU - Bulayeva, Nataliya
AU - Guptarak, Jutatip
PY - 2012/4
Y1 - 2012/4
N2 - Multiple physiologic estrogens (estradiol, estriol, and estrone), as well as xenoestrogenic compounds (including alkylphenols and bisphenol A), can act via nongenomic signaling initiated by liganding of the plasma membrane estrogen receptor-α (mERα). We examined heterotrimeric G protein involvement leading to extracellular-regulated kinase (ERK) activation in GH3/B6/F10 rat anterior pituitary tumor cells that express abundant mERα, and smaller amounts of mERβ and GPR30. A combination of microarrays, immunoblots, and quantitative immunoassays demonstrated the expression of members of all α, β, and γ G protein classes in these cells. Use of selective inhibitors showed that the G αi subtype was the primary initiator of downstream ERK signaling. Using antibodies against the GTP-bound form of G α protein subtypes I and s, we showed that xenoestrogens (bisphenol A, nonylphenol) activated G αi at 15-30 s; all alkylphenols examined subsequently suppressed activation by 5 min. GTP-activation of G αi for all estrogens was enhanced by irreversible cumulative binding to GTPγS. In contrast, G αs was neither activated nor deactivated by these treatments with estrogens. ERα and G αi co-localized outside nuclei and could be immuno-captured together. Interactions of ERα with G αi and caveolin I were demonstrated by epitope proximity ligation assays. An ERα/β antagonist (ICI182780) and a selective disruptor of caveolar structures (nystatin) blocked estrogen-induced ERK activation. Conclusions: Xenoestrogens, like physiologic estrogens, can evoke downstream kinase signaling involving selective interactions of ERα with G αi and caveolin I, but with some different characteristics, which could explain their disruptive actions.
AB - Multiple physiologic estrogens (estradiol, estriol, and estrone), as well as xenoestrogenic compounds (including alkylphenols and bisphenol A), can act via nongenomic signaling initiated by liganding of the plasma membrane estrogen receptor-α (mERα). We examined heterotrimeric G protein involvement leading to extracellular-regulated kinase (ERK) activation in GH3/B6/F10 rat anterior pituitary tumor cells that express abundant mERα, and smaller amounts of mERβ and GPR30. A combination of microarrays, immunoblots, and quantitative immunoassays demonstrated the expression of members of all α, β, and γ G protein classes in these cells. Use of selective inhibitors showed that the G αi subtype was the primary initiator of downstream ERK signaling. Using antibodies against the GTP-bound form of G α protein subtypes I and s, we showed that xenoestrogens (bisphenol A, nonylphenol) activated G αi at 15-30 s; all alkylphenols examined subsequently suppressed activation by 5 min. GTP-activation of G αi for all estrogens was enhanced by irreversible cumulative binding to GTPγS. In contrast, G αs was neither activated nor deactivated by these treatments with estrogens. ERα and G αi co-localized outside nuclei and could be immuno-captured together. Interactions of ERα with G αi and caveolin I were demonstrated by epitope proximity ligation assays. An ERα/β antagonist (ICI182780) and a selective disruptor of caveolar structures (nystatin) blocked estrogen-induced ERK activation. Conclusions: Xenoestrogens, like physiologic estrogens, can evoke downstream kinase signaling involving selective interactions of ERα with G αi and caveolin I, but with some different characteristics, which could explain their disruptive actions.
KW - Alkylphenols
KW - Bisphenol A
KW - GH3 cells
KW - GPR30
KW - Membrane estrogen receptor
KW - Nongenomic
UR - http://www.scopus.com/inward/record.url?scp=84858341505&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84858341505&partnerID=8YFLogxK
U2 - 10.1016/j.steroids.2011.12.025
DO - 10.1016/j.steroids.2011.12.025
M3 - Article
C2 - 22230296
AN - SCOPUS:84858341505
SN - 0039-128X
VL - 77
SP - 424
EP - 432
JO - Steroids
JF - Steroids
IS - 5
ER -