Tyrosine phosphates act on steroidogenesis through the activation of arachidonic acid release
Tyrosine phosphates act on steroidogenesis through the activation of arachidonic acid release
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DOI：
10.1081/erc-200043795
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发表时间：
2004-01-01
影响因子：
2.1
通讯作者：
Maciel, FC
中科院分区：
文献类型：
--
作者：
Castillo, F;Cano, F;Maciel, FC
ACTH信号通路包括PKA活化以及PKA依赖性酪氨酸磷酸酶活化。此外，这种激素的作用还包括通过两种酶的协同作用调节花生四烯酸（AA）的细胞内水平：酰基CoA-硫酯酶和酰基-CoA-合成酶（ACS 4）。本工作描述了一个特定的ACS 4抗体的生产和表征，这是用来分析ACTH对Y1肾上腺皮质细胞中ACS 4蛋白水平的影响和酪氨酸磷酸酶和ACS 4之间的关系。免疫家兔获得抗血清。该免疫原在细菌中产生，并在部分纯化的细菌裂解物的SDS-PAGE分离后从丙烯酰胺凝胶洗脱。当用于蛋白质印迹分析时，获得的抗体仅特异性识别Y1细胞和几种大鼠组织中对应于ACS 4的分子量的一种蛋白质。使用这里描述的抗体，我们分析了ACTH刺激对ACS 4蛋白水平的影响。激素产生的增加，这种酰基辅酶A合成酶在Y1肾上腺皮质细胞。此外，这种效果是模仿cAMP和部分减少酪氨酸磷酸酶抑制剂。我们建议，ACTH调节ACS 4蛋白水平通过PKA依赖的机制，也可能涉及PTP活性。
The ACTH signaling pathway includes both PKA activation as well as PKA-dependent tyrosine phosphatase activation. In addition, the action of this hormone also includes the regulation of the intracellular levels of arachidonic acid (AA) by the concerted action of two enzymes: an acylCoA-thioesterase and an acyl-CoA-synthetase (ACS4). This work describes the production and characterization of a specific ACS4 antibody, which was used to analyze the effect of ACTH on ACS4 protein level in Y1 adrenocortical cells and the putative relationship between tyrosine phosphatases and ACS4. The antiserum was obtained from rabbits immunized with the recombinant ACS4. This immunogen was produced in bacteria and eluted from an acrylamide gel after SDS-PAGE separation of a partially purified bacteria lysate. When used in Western blot analysis, the antibody obtained specifically recognized only one protein of the molecular mass corresponding to ACS4, in Y1 cells and in several rat tissues. Using the antibody described here, we analyzed the effect of ACTH stimulation on ACS4 protein level. The hormone produced an increase of this acyl-CoA synthetase in Y1 adrenocortical cells. Moreover, this effect was mimicked by cAMP and partially reduced by a tyrosine phosphatase inhibitor. We propose that ACTH regulates ACS4 protein levels through a PKA-dependent mechanism that could involve also PTP activity.