醛固酮产生腺瘤是一种良性肾上腺肿瘤，具有自主产生醛固酮的能力，这会导致高血压和过度心血管风险。蛋白磷酸化调节肾上腺皮质细胞中的醛固酮分泌，但在醛固酮产生腺瘤中信号网络的重构仍不清楚。我们用四维无标签技术对15例醛固酮产生腺瘤和10例非功能性肾上腺皮质肿瘤（NFAT）进行了综合蛋白质组和磷酸化蛋白质组分析。我们进一步在扩大的醛固酮产生腺瘤样本、小鼠和肾上腺皮质细胞系中验证了我们的主要发现。 醛固酮产生腺瘤和NFAT蛋白质组和磷酸化蛋白质组分析量化了6500种蛋白质和9797种磷酸肽。我们突出了差异表达和磷酸化的蛋白质，它们调节了醛固酮的合成和分泌。由于胞内钙是醛固酮合成的中心信号，我们的综合钙信号网络表明，Wolframin在控制胞内钙流和醛固酮合酶（CYP11B2）激活中起到重要作用（Wolframin在APA和NFAT中比例为6.411，P <0.001）。在97例醛固酮产生腺瘤的验证中，Wolframin表达水平较高的患者在卡普托普利挑战试验后血浆醛固酮浓度和收缩压均较高。体外实验表明，Wolframin过表达可增强醛固酮的分泌，而敲低Wolframin会抑制钾或血管紧张素II刺激下的醛固酮分泌。进一步的体内和体外数据证明了Wolframin-钙轴对CYP11B2表达和醛固酮的产生具有重要调节作用。Wolframin是醛固酮增多症的调节蛋白。REM构象钙离子的转运和线粒体功能参与了Wolframin相关的醛固酮分泌。

Aldosterone-producing adenoma (APA) is a benign adrenal tumor with autonomous aldosterone production which causes hypertension and excess cardiovascular risk. Protein phosphorylation regulates aldosterone secretion from adrenal cortical cells, but how signaling networks are remodeled in APA remains unknown.We performed an integrated proteomic and phosphoproteomic profiling of 15 APA and 10 matched nonfunctioning adrenocortical tumors (NFAT) based on the 4-dimensional label-free technique. We further validated our main findings in enlarged APA samples, mice, and adrenocortical cell line.The proteomic and phosphoproteomic profiling of APA and NFAT quantified 6500 proteins and 9797 phosphopeptides. We highlighted differentially expressed and phosphorylated proteins which modulated aldosterone synthesis and secretion from APA. As intracellular calcium is the central signal for aldosterone synthesis, our integrated calcium signaling network implicated wolframin in the control of calcium influx and aldosterone synthase (CYP11B2) activation in APA (ratio of wolframin expression in APA to NFAT: 6.411, P<0.001). Among 97 APA cases for validation, a higher expression level of wolframin was associated with a higher plasma aldosterone concentration postcaptopril challenge test and a higher systolic blood pressure. In vitro, the secretion of aldosterone was enhanced by wolframin overexpression, while aldosterone secretion in response to potassium or angiotensin II was inhibited by the knockdown of wolframin. Further in vivo and in vitro data demonstrated the wolframin-calcium axis as an important regulator of CYP11B2 expression and aldosterone production.Wolframin is a regulatory protein in aldosterone hypersecretion. Remodeled calcium transportation and mitochondrial function are involved in wolframin-related aldosterone secretion.