Calcium‐dependent phosphorylation processes control brain aromatase in quail

Abstract

Increased gene transcription activated by the binding of sex steroids to their cognate receptors is one important way in which oestrogen synthase (aromatase) activity is regulated in the brain. This control mechanism is relatively slow (hours to days) but recent data indicate that aromatase activity in quail preoptic-hypothalamic homogenates is also rapidly (within minutes) affected by exposure to conditions that enhance Ca2+-dependent protein phosphorylation. We demonstrate here that Ca2+-dependent phosphorylations controlled by the activity of multiple protein kinases including PKC, and possibly also PKA and CAMK, can rapidly down-regulate aromatase activity in brain homogenates. These phosphorylations directly affect the aromatase molecule itself. Western blotting experiments on aromatase purified by immunoprecipitation reveal the presence on the enzyme of phosphorylated serine, threonine and tyrosine residues in concentrations that are increased by phosphorylating conditions. Cloning and sequencing of the quail aromatase identified a 1541-bp open reading frame that encodes a predicted 490-amino-acid protein containing all the functional domains that have been previously described in the mammalian and avian aromatase. Fifteen predicted consensus phosphorylation sites were identified in this sequence, but only two of these (threonine 455 and 486) match the consensus sequences corresponding to the protein kinases that were shown to affect aromatase activity during the pharmacological experiments (i.e. PKC and PKA). This suggests that the phosphorylation of one or both of these residues represents the mechanism underlying, at least in part, the rapid changes in aromatase activity.

Gossypol Increases Expression of the Pro-apoptotic BH3-only Protein NOXA through a Novel Mechanism Involving Phospholipase A2, Cytoplasmic Calcium, and Endoplasmic Reticulum Stress

“…Inhibition of these enzymes can be explained by either PLA2 activity or increased cytoplasmic calcium. For example, AA has been shown to inhibit 3␣-hydroxysteroid dehydrogenase (51), whereas an increase in calcium concentration has been shown to inhibit telomerase and aromatase (52,53).…”

Mechanistic Basis and Functional Roles of Long-Term Plasticity in Auditory Neurons Induced by a Brain-Generated Estrogen

“…The rapid (MAPKindependent) effects of E2 on neural response properties are owed to its impact on the synaptic machinery (Tremere et al, 2009) and likely the fast regulation of aromatase activity. In fact, in NCM and other avian brain areas, aromatase is localized to presynaptic terminals and its activity is rapidly regulated by neuronal activity, in a calcium-dependent fashion (Balthazart et al, 2003;Peterson et al, 2005;Rohmann et al, 2007;Saldanha et al, 2011). These findings suggest that E2 operates more akin to a neuromodulator than a steroid hormone proper (Balthazart and Ball, 2006).…”

Functional significance of the rapid regulation of brain estrogen action: Where do the estrogens come from?

“…This inhibition is blocked by compounds chelating divalent ions such as EGTA and EDTA or by kinases inhibitors such as inhibitors of protein kinase A (PKA) or C (PKC) indicating that it is indeed caused by phosphorylation processes [16,18]. The identification of putative phosphorylation sites for PKC on the predicted quail aromatase sequence and westernblotting experiments on quail brain aromatase partially purified by immunoprecipitation confirmed that phosphorylation sites located on the aromatase protein itself are affected by the phosphorylating conditions that influence the enzymatic activity [18,19].…”