Defective PKA Signaling in Cushing's Syndrome

摘要 当肾上腺释放过多的应激激素皮质醇时，库欣综合症就会发生。这 内分泌紊乱自然是由肾上腺或脑下垂体瘤引起的，女性的发病率要高出五倍。 比男人更频繁。长期接触皮质醇还会增加血糖和血压。 本研究旨在探讨肾上腺库欣综合征的分子发病机制。这种疾病通常是一种 肾上腺皮质cAMP信号缺陷的结果。精准医学的进展已经确定了要点 蛋白激酶A催化亚基(PKAc)的突变和插入是这种疾病的驱动因素。最多的 流行的突变是PKAc-L205R，在大约45%的库欣患者中发现。其他遗传损害包括 PKAc-W196R。这两种致病突变都驻留在与 它的监管亚单位。这种蛋白质-蛋白质界面对于自身抑制激酶活性是必要的，但也 通过结合A激酶锚定蛋白来指导PKA全酶的亚细胞靶向 (AKAP)。我们的生化、基于细胞和临床的分析推断，携带PKAc-L205R的突变等位基因 或者W196R足以取代库欣患者正常的亚细胞作用部位的激酶。这 导致了一个工作假说，即失去对蛋白激酶A的时空控制是肾上腺库欣病的基础 综合症。AIM 1将测试i)PKA锚定的恢复是否能纠正皮质醇的释放，ii)确定哪些AKAP 协调皮质醇生物合成和iii)考虑了一个相反的前提，即PKAc变异体的底物重新连接 或者酶稳定性的改变会导致库欣综合症。乍一看，PKAc-L205R和PKAc-W196R 变种人看起来几乎一模一样。然而，我们已经发现，这些是生理和化学上截然不同的激酶 参与不同的细胞信号通路。对患者组织的分析显示PKAc-L205R对河马具有动员作用 控制器官大小的信号通路。相反，肾上腺特异性敲入小鼠的表型特征表明 PKAc-W196R激活ERK丝裂原激活级联反应。AIM 2将评估i)女性PKAcWT/W196R 小鼠更容易患库欣合并症，II)测试PKAc-L205R是否参与河马信号级联反应 推动皮质醇过度生产和iii)确定PKAc-W196R是否协调ERK信号事件以促进 肾上腺增生症。这项提议探索了两个将改变我们思考方式的变革想法 库欣综合征。1)失去对蛋白激酶A的时空控制是肾上腺库欣综合征的基础 病理学。2)PKAc的相邻和相同突变导致肾上腺增生和皮质醇高分泌 在库欣的病人身上。

ABSTRACT Cushing’s syndrome occurs when the adrenal glands release too much of the stress hormone cortisol. This endocrine disorder is naturally caused by adrenal or pituitary tumors and afflicts women five times more frequently than men. Chronic exposure to cortisol also increases blood glucose and elevates blood pressure. This proposal investigates the molecular pathogenesis of adrenal Cushing’s syndrome. This disease is often an outcome of defective cAMP signaling in the adrenal cortex. Advances in precision medicine have identified point mutations and insertions in protein kinase A catalytic subunits (PKAc) as drivers of this disease. The most prevalent mutant is PKAc-L205R, which is found in ~45% of Cushing’s patients. Other genetic lesions include PKAc-W196R. Both disease-causing mutations reside within a region of the kinase domain that interfaces with its regulatory subunits. This protein-protein interface is necessary for autoinhibition of kinase activity, but also directs the subcellular targeting of PKA holoenzymes through association with A kinase anchoring proteins (AKAPs). Our biochemical, cell-based, and clinical analyses infer that carrying a mutant allele of PKAc-L205R or W196R is sufficient to displace the kinase from normal subcellular sites of action in Cushing’s patients. This has led to a working hypothesis that losing spatiotemporal control of protein kinase A underlies adrenal Cushing’s syndrome. Aim 1 will test if i) restoration of PKA anchoring amends cortisol release, ii) identify which AKAPs coordinate cortisol biosynthesis and iii) considers an opposing premise that “substrate rewiring” of PKAc variants or altered enzyme stability drives Cushing’s syndrome. At first glance, the PKAc-L205R and PKAc-W196R mutants look virtually identical. Yet we have discovered that these are physiochemically distinct kinases that engage different cell signaling pathways. Analyses of patient tissue reveal that PKAc-L205R mobilizes the Hippo signaling pathway to control organ size. Conversely phenotypic profiling of adrenal-specific knock-in mice indicate that PKAc-W196R activates the ERK mitogenic kinase cascade. Aim 2 will evaluate i) if female PKAcWT/W196R mice are more prone to Cushing’s comorbidities, ii) test if PKAc-L205R engages the Hippo signaling cascade to drive cortisol hyperproduction and iii) ascertain if PKAc-W196R coordinates ERK signaling events to promote adrenal hyperplasia. This proposal explores two transformative ideas that will change how we think about Cushing’s syndrome. 1) Losing spatiotemporal control of protein kinase A underlies adrenal Cushing’s pathologies. 2) Adjacent and identical mutations in PKAc drive adrenal hyperplasia and cortisol hypersecretion in Cushing’s patients.