Regulation of Mineralocorticoid Receptor Action

盐皮质激素受体作用的调节

• 批准号: 10553631
• 负责人: Celso Enrique Gomez-Sanchez
• 金额: --
• 依托单位: G V SONNY MONTGOMERY VA MEDCIAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-01 至 2024-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10553631
• 关键词: Affect; Affinity; Alcohol Oxidoreductases; Aldosterone; Amino Acids; Angiotensin II; Antibodies; Area; Binding; Blood Pressure; Brain; CXCR3 gene; Cardiovascular system; Cell Line; Cells; Complications of Diabetes Mellitus; Congestive Heart Failure; Corticosterone; Cortisone; Cultured Cells; Dahl Hypertensive Rats; Data; Development; Diet; Distal; Dominant-Negative Mutation; Enzymes; Epithelium; Essential Hypertension; Gene Expression; Genomics; Glucocorticoid Receptor; Heart; Histocytochemistry; Hormones; Human; Hydrocortisone; Hydroxysteroid Dehydrogenases; Hyperaldosteronism; Hypertension; Immunofluorescence Immunologic; Intercalated Cell; Kidney; Ligand Binding Domain; Ligands; Luciferases; Measures; Mediating; Mineralocorticoid Receptor; Mouse Mammary Tumor Virus; Movement; Nephrons; Normal Range; Organ; Pathogenicity; Pathologic; Patients; Phosphorylation; Phosphotransferases; Plasma; Positioning Attribute; Post-Translational Protein Processing; Potassium; Protein Tyrosine Kinase; Rattus; Receptor Activation; Regulation; Reporter Genes; Reporting; Role; Serine; Serine/Threonine Phosphorylation; Site; Sleep Apnea Syndromes; Sodium; Sodium Chloride; Specificity; Steroids; System; TBK1 gene; Threonine; Tissues; Transactivation; Water; cell type; inorganic phosphate; kidney cell; non-genomic; normotensive; overexpression; receptor; receptor binding; response; serine receptor; side effect; steroid hormone; transcription factor

The mineralocorticoid receptor (MR) is a transcription factor expressed in many cells that regulates the expression of genes in involved myriad cell type-specific functions. Inappropriate activation of the MR causes hypertension and pathological tissue remodeling even in conditions of normal aldosterone levels. Activity of the MR is regulated by post-translational modifications including phosphorylation of serines and threonines. Phosphorylation activates or suppresses MR activity depending upon the amino acid involved. The kinase ULK1 was recently reported to phosphorylate serine 843 of the MR, thereby suppressing its activity, and to be expressed with the MR only in intercalated cells of the distal nephron. Our own specific antibodies reveal more extensive co-expression of ULK1 and MR in the kidney. ULK1 expression is widespread in the body and in multiple cell lines. Other kinases also have the potential to phosphorylate the MR at S843, thus inhibit MR activity. The MR has similar affinity for aldo, cortisol or corticosterone. Specificity for aldo is conferred by conversion of cortisol and corticosterone to the inactive cortisone and 11-dehydrocorticosterone by the 11β-hydroxysteroid dehydrogenase 2 (11-HSD2), however this enzyme is not present in most non-epithelial aldo target tissues. Cortisol and corticosterone are also converted to 20β-dihydro-cortisol and -corticosterone by carbonyl reductase 1 (Cbr1). We that found the 20β-dihydro metabolites do not activate the MR, thus providing an alternative mechanism for MR specificity for aldo where 11-HSD2 is not expressed. Hypotheses: 1) Phosphorylation of the MR at serine 843 is an important negative regulator of MR action that occurs in multiple tissues. 2) Reduction of the 20-keto of corticosterone and cortisol by carbonyl reductase 1 regulates ligand selectivity for the MR by converting corticosterone and cortisol into the inactive metabolites 20β-dihydrocorticosterone (20β-DHC) and 20β-dihydrocortisol. Both potentially modulate MR activity. Specific Aim 1: Study the role of phosphorylation of the MR at serine 843 (human) on genomic and non- genomic activity of the MR. Study the effects of over-expression and suppression of several kinases, ULK1, ULK2, TBK1, NEK2 and PAK1, with putative ability to phosphorylate the MR at S843 in cells expressing an MR reporter gene system. Study the distribution and co-localization of the MR with ULK1, ULK2 and other kinases that catalyze S843 phosphorylation in the kidney, heart, vessels and brain. Specific Aim 2: Study the role of the 20-keto reduction of cortisol and corticosterone on MR ligand selectivity. Determine the dynamics of the 20-keto reduction of corticosterone and cortisol by the human and rat Cbr1 in cells. Measure the conversion of corticosterone to 20β-dihydrocorticosterone in various tissues including in micropunches from specific areas of the rat brain where we demonstrate that Cbr1 and MR are co-expressed by immunofluorescent histochemistry. Measure plasma levels of corticosterone and 20β-DHC in plasma and tissues of rats fed different amounts of salt. The mechanisms for the cell-specific modulation of MR activity will never be known if not explored and are crucial for the development of rational therapy with minimal side effects for inappropriate MR activation.

盐皮质激素受体 (MR) 是一种在许多细胞中表达的转录因子，可调节 涉及多种细胞类型特异性功能的基因表达。 MR 激活不当的原因 即使在醛固酮水平正常的情况下，也会出现高血压和病理性组织重塑。的活动 MR 受到翻译后修饰的调节，包括丝氨酸和苏氨酸的磷酸化。 磷酸化激活或抑制 MR 活性取决于所涉及的氨基酸。激酶 最近报道 ULK1 磷酸化 MR 的丝氨酸 843，从而抑制其活性，并且 MR 仅在远端肾单位的闰细胞中表达。我们自己的特异性抗体揭示了更多 ULK1 和 MR 在肾脏中广泛共表达。 ULK1 表达广泛存在于体内 多个细胞系。其他激酶也有可能磷酸化 MR S843，从而抑制 MR 活动。 MR 对醛、皮质醇或皮质酮具有相似的亲和力。 ALDO 的特异性是通过转化而赋予的 皮质醇和皮质酮通过 11β-羟基类固醇转化为无活性的可的松和 11-脱氢皮质酮 脱氢酶 2 (11-HSD2)，但大多数非上皮醛靶组织中不存在该酶。 皮质醇和皮质酮也通过羰基转化为20β-二氢皮质醇和20β-二氢皮质酮 还原酶 1 (Cbr1)。我们发现 20β-二氢代谢物不会激活 MR，从而提供了 不表达 11-HSD2 的 ALDO MR 特异性的替代机制。 假设：1) MR 丝氨酸 843 处的磷酸化是 MR 作用的重要负调节因子 发生在多个组织中。 2) 羰基还原酶 1 还原皮质酮和皮质醇的 20-酮基 通过将皮质酮和皮质醇转化为无活性代谢物来调节 MR 的配体选择性 20β-二氢皮质酮（20β-DHC）和20β-二氢皮质醇。两者都可能调节 MR 活动。 具体目标 1：研究 MR 丝氨酸 843（人）磷酸化对基因组和非基因组的作用 MR 的基因组活性。研究几种激酶 ULK1 的过度表达和抑制的影响， ULK2、TBK1、NEK2 和 PAK1，假定能够在表达 MR 的细胞中磷酸化 S843 处的 MR 报告基因系统。研究 MR 与 ULK1、ULK2 和其他激酶的分布和共定位 催化肾脏、心脏、血管和大脑中的 S843 磷酸化。 具体目标 2：研究皮质醇和皮质酮的 20-酮还原对 MR 配体选择性的作用。 确定人和大鼠 Cbr1 对皮质酮和皮质醇 20-酮还原的动态 细胞。测量各种组织中皮质酮向 20β-二氢皮质酮的转化，包括 来自大鼠大脑特定区域的微穿孔，我们证明 Cbr1 和 MR 是共表达的 通过免疫荧光组织化学。测量血浆和血浆中皮质酮和 20β-DHC 的血浆水平 给老鼠的组织喂食不同量的盐。 如果不进行探索和研究，细胞特异性调节 MR 活性的机制将永远无法得知。 对于开发合理的治疗方法至关重要，并且对于不适当的 MR 激活具有最小的副作用。