Rapid estrogenic regulation of extracellular signal-regulated kinase 1/2 signaling in cerebellar granule cells involves a G protein- and protein kinase A-dependent mechanism and intracellular activation of protein phosphatase 2A

In neonatal rat cerebellar neurons, 17 beta-estradiol (E-2) rapidly stimulates ERK1/2 phosphorylation through a membrane-associated receptor. Here the mechanism of rapid E-2-induced ERK1/2 signaling in primary cultured granule cells was investigated in more detail. The results of these studies show that E-2 and ICI182,780, a steroidal antagonist of estrogen receptor transactivation, rapidly increased ERK signaling with a time course similar to the transient activation induced by epidermal growth factor (EGF). However, EGF receptor (EGFR) autophosphorylation was not increased by E-2, and blockade of EGFR tyrosine kinase activity did not abrogate the rapid actions of E-2. The involvement of Src-tyrosine kinase activity was demonstrated by detection of increased c-Src phosphorylation in response to E-2 and by blockade of E-2-induced ERK1/2 activation by inhibition of Src-family tyrosine kinase activity. Inhibition of G alpha i signaling or protein kinase A (PKA) activity blocked the ability of ICI182,780 to rapidly stimulate ERK signaling. Under those conditions, E-2 treatment induced a rapid and transient suppression of basal ERK1/2 phosphorylation. Protein phosphatase 2A (PP2A) activity was rapidly increased by E-2 but not by E-2 covalently linked to BSA. Rapid E-2-induced increases in PP2A activity were insensitive to pertussis toxin. The presented evidence indicates that the rapid effects of estrogens on ERK signaling in cerebellar granule cells are induced through a novel G protein-coupled receptor mechanism that requires PKA and Src-kinase activity to link E-2 to the ERK/MAPK signaling module. Along with stimulating ERK signaling, E-2 rapidly activates PP2A via an independent signaling mechanism that may serve as a cell-specific regulator of signal duration.