Calcium-dependent phosphorylation processes control brain aromatase in quail
Calcium-dependent phosphorylation processes control brain aromatase in quail
复制标题
DOI：
10.1046/j.1460-9568.2003.02598.x
复制
发表时间：
2003-04-01
影响因子：
3.4
通讯作者：
Ball, GF
中科院分区：
文献类型：
--
作者：
Balthazart, J;Baillien, M;Ball, GF
脑内雌激素合成酶（芳香化酶）活性调控的重要途径之一是性类固醇与其同源受体结合而激活的基因转录增加。这种控制机制相对较慢（数小时至数天），但最近的数据表明，暴露于增强Ca2+依赖性蛋白磷酸化的条件下，鹌鹑视前下丘脑匀浆中的芳香酶活性也会迅速（几分钟内）受到影响。我们在这里证明，Ca2+依赖性磷酸化由多种蛋白激酶的活性控制，包括PKC，也可能是PKA和CAMK，可以快速下调脑匀浆中的芳香酶活性。这些磷酸化直接影响芳香化酶分子本身。免疫沉淀纯化的芳香酶的Western blotting实验显示，磷酸化的丝氨酸、苏氨酸和酪氨酸残基在酶上的浓度随着磷酸化条件的增加而增加。鹌鹑芳香化酶的克隆和测序鉴定出一个1541 bp的开放阅读框，该阅读框编码一个预测的490个氨基酸的蛋白，包含先前在哺乳动物和鸟类芳香化酶中描述的所有功能域。在该序列中确定了15个预测的共识磷酸化位点，但其中只有两个（苏氨酸455和486）与药理学实验中显示影响芳香酶活性的蛋白激酶（即PKC和PKA）对应的共识序列相匹配。这表明，其中一个或两个残基的磷酸化至少部分地代表了芳香酶活性快速变化的机制。
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.