Defective PKA Signaling in Cushing's Syndrome

库欣综合征中 PKA 信号传导缺陷

• 批准号: 10582988
• 负责人: John D Scott
• 金额: $ 49.96万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-24 至 2028-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10582988
• 关键词: A kinase anchoring protein; Adrenal Cortex; Adrenal Gland Neoplasms; Adrenal Glands; Alleles; Amendment; Anabolism; Arginine; Back; Biochemical; Blood Circulation; Blood Glucose; Catalytic Domain; Cell Nucleus; Cells; Chronic; Clinical; Cushing Syndrome; Cushingoid facies; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Defect; Development; Diabetes Mellitus; Disease; Endocrine; Endocrine System Diseases; Enzyme Stability; Enzymes; Event; Exclusion; Exposure to; Female; Glucose; Holoenzymes; Hormones; Hydrocortisone; Hypertension; Island; Knock-in Mouse; Left; Lesion; Molecular; Mus; Mutation; Operative Surgical Procedures; Organ Size; Other Genetics; Outcome; Pathogenesis; Pathogenicity; Pathologic; Pathology; Patients; Pharmaceutical Preparations; Phenotype; Phosphotransferases; Pituitary Neoplasms; Point Mutation; Proteins; Public Health; Radiation; Rosaniline Dyes; Sampling; Signal Pathway; Signal Transduction; Signaling Protein; Site; Stains; Stress; Symptoms; Testing; Tissues; Variant; Weight Gain; Woman; adenoma; adrenal hyperplasia; blood pressure elevation; chemotherapy; comorbidity; disease-causing mutation; intracellular protein transport; men; mouse model; mutant; precision medicine; protein distribution; restoration; spatiotemporal; transmission process; virtual

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.

摘要 当肾上腺释放过多的应激激素皮质醇时，就会发生库欣综合征。这 内分泌失调是由肾上腺或垂体肿瘤引起的，女性的发病率是男性的五倍多。 往往比男人。长期接触皮质醇也会增加血糖和血压。 本研究旨在探讨肾上腺皮质库欣综合征的分子发病机制。这种疾病往往是一种 肾上腺皮质cAMP信号传导缺陷的结果。精准医疗的进步已经确定了 蛋白激酶A催化亚基（PKAc）中的突变和插入是这种疾病的驱动因素。最 流行的突变体是PKAc-L205 R，其在约45%的库欣患者中发现。其他遗传性病变包括 PKAc-W196R。两种致病突变都位于激酶结构域的一个区域内， 其调节亚基。这种蛋白质-蛋白质界面对于激酶活性的自抑制是必需的，而且 通过与A激酶锚定蛋白的结合指导PKA全酶的亚细胞靶向 （AKAP）。我们的生化、细胞和临床分析推断，携带PKAc-L205 R突变等位基因的人， 或W196 R足以将该激酶从库欣病患者的正常亚细胞作用位点置换。这 导致了一个工作假设，即蛋白激酶A的时空控制丧失是肾上腺库欣综合征的基础 综合征目标1将测试i）PKA锚定的恢复是否修正皮质醇释放，ii）识别哪些AKAP 协调皮质醇生物合成和iii）考虑相反的前提，即PKAc变体的“底物重新布线” 或酶稳定性的改变导致了库欣综合征。乍一看，PKAc-L205 R和PKAc-W196 R 突变体看起来几乎一模一样。然而，我们已经发现，这些是物理化学上不同的激酶， 参与不同的细胞信号通路。对患者组织的分析表明PKAc-L205 R动员了Hippo 控制器官大小的信号通路。相反，肾上腺特异性基因敲入小鼠的表型分析表明， PKAc-W196 R激活ERK促有丝分裂激酶级联。目标2将评价i）雌性PKAcWT/W196 R 小鼠更容易患库欣氏合并症，ii）测试PKAc-L205 R是否参与Hippo信号级联反应， iii）确定PKAc-W196 R是否协调ERK信号传导事件以促进皮质醇的过度产生 肾上腺增生这份提案探讨了两个变革性的想法，它们将改变我们对 库欣综合征。1)蛋白激酶A时空调控的丧失是肾上腺库欣综合征的基础 病理学2)PKAc的相邻和相同突变驱动肾上腺增生和皮质醇分泌过多 在库欣的病人中。

项目成果

AKAP Signaling Islands: Venues for Precision Pharmacology.

• DOI: 10.1016/j.tips.2020.09.007
• 发表时间: 2020-12
• 期刊: Trends in pharmacological sciences
• 影响因子: 13.8
• 作者: Omar MH;Scott JD
• 通讯作者: Scott JD

Classification of Cushing's syndrome PKAc mutants based upon their ability to bind PKI

• DOI: 10.1042/bcj20230183
• 发表时间: 2023-06-01
• 期刊: BIOCHEMICAL JOURNAL
• 影响因子: 4.1
• 作者: Omar，Mitchell H.;Kihiu，Maryanne;Scott，John D.
• 通讯作者: Scott，John D.

Proximity biotinylation to define the local environment of the protein kinase A catalytic subunit in adrenal cells.

• DOI: 10.1016/j.xpro.2022.101992
• 发表时间: 2023-03-17
• 期刊: STAR protocols
• 作者: Omar MH;Lauer SM;Lau HT;Golkowski M;Ong SE;Scott JD
• 通讯作者: Scott JD

Mislocalization of protein kinase A drives pathology in Cushing's syndrome.

• DOI: 10.1016/j.celrep.2022.111073
• 发表时间: 2022-07-12
• 期刊: CELL REPORTS
• 影响因子: 8.8
• 作者: Omar, Mitchell H.;Byrne, Dominic P.;Jones, Kiana N.;Lakey, Tyler M.;Collins, Kerrie B.;Lee, Kyung-Soon;Daly, Leonard A.;Forbush, Katherine A.;Lau, Ho-Tak;Golkowski, Martin;McKnight, G. Stanley;Breault, David T.;Lefrancois-Martinez, Anne-Marie;Martinez, Antoine;Eyers, Claire E.;Baird, Geoffrey S.;Ong, Shao-En;Smith, F. Donelson;Eyers, Patrick A.;Scott, John D.
• 通讯作者: Scott, John D.

Neurotensin as a source of cyclic AMP and co-mitogen in fibrolamellar hepatocellular carcinoma.

神经降压素作为纤维板层肝细胞癌中环磷酸腺苷和共丝裂原的来源。

• DOI: 10.18632/oncotarget.27149
• 发表时间: 2019
• 期刊: Oncotarget
• 作者: Riehle,KimberlyJ;Kenerson,HeidiL;Riggle,KevinM;Turnham,Rigney;Sullivan,Kevin;Bauer,Renay;Scott,JohnD;Yeung,RaymondS
• 通讯作者: Yeung,RaymondS