The role of salt inducible kinases in renal PTH action

盐诱导激酶在肾 PTH 作用中的作用

基本信息

批准号：
10207599
负责人：
Michael Mannstadt
金额：
$ 41.09万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10207599
关键词：
3-Dimensional Acute Adaptor Signaling Protein Adenine Autoimmune Binding Biology Blood Bone Diseases Bone Resorption Bypass CYP27B1 gene Calcium Cells Chronic Chronic Kidney Failure Cyclic AMP Cyclic AMP-Dependent Protein Kinases Data Diet Disease Endocrine Enzymes Epithelial Cells Event Excretory function Family Family member Future Gene Expression Genes Genetic Transcription Goals Homeostasis Hormonal Hormone Responsive Hormone use Hormones Human In Vitro Inflammatory Injury to Kidney Inorganic Phosphate Transporter Ions Kidney Kidney Diseases Knock-out Knowledge Light Link Mediating Mediator of activation protein Metabolism Minerals Mission Modeling Modification Molecular Mus National Institute of Diabetes and Digestive and Kidney Diseases Nephrons Nucleic Acid Regulatory Sequences Oral Organ Organoids Osteocytes Osteogenesis Osteoporosis PTH gene Parathyroid Hormone Receptor Pathway interactions Pharmaceutical Preparations Phenotype Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Inhibition Play Post-Translational Regulation Production Protein Isoforms Protein Phosphatase 2A Regulatory Subunit PR53 Proteins Proximal Kidney Tubules Receptor Activation Regulation Regulatory Pathway Research Role Serum Signal Pathway Signal Transduction Testing Therapeutic Therapeutic Uses Tissues Transcriptional Activation United States National Institutes of Health Upstream Enhancer Vitamin D bone bone health bone mass brush border membrane calcium metabolism chromatin immunoprecipitation experimental study hormonal signals hormone resistance human disease in vivo inhibitor/antagonist inorganic phosphate kidney cell kinase inhibitor mouse model novel novel strategies novel therapeutic intervention receptor renal epithelium salt-inducible kinase skeletal skeletal disorder small molecule small molecule inhibitor therapeutic target transcription factor transcriptome transcriptome sequencing treatment effect uptake urinary

项目摘要

Project Summary Parathyroid hormone (PTH) maintains mineral ion homeostasis by interacting with the PTH/PTHrP receptor to induce transcriptional and post-transcriptional modifications in target tissues. PTH stimulates resorption of the mineral matrix in bone, and increases production of active vitamin D, calcium reabsorption, and phosphate excretion in the kidneys. Despite the central role of PTH as a regulatory hormone, many of the downstream intracellular target molecules that orchestrate PTH signaling events have yet to be identified. Characterization of these pathways will expand current knowledge of the mechanisms governing mineral homeostasis, and suggest new therapeutic strategies to counteract or restore PTH action in skeletal and renal disease. Recently, salt- inducible kinases (SIKs) were identified as intracellular mediators of PTH signaling in osteocytes, and small- molecule inhibitors of SIKs were found to mimic PTH action on bone in vivo. The objective of this proposal is to explore whether analogous SIK-dependent PTH signaling pathways are active in the kidney. Aim 1 of this proposal will explore the molecular mechanisms through which PTH/SIK signaling regulates CYP27B1 expression. In renal cells, human kidney organoids, and mice, both PTH and the SIK inhibitor YKL-05-099 increase expression of CYP27B1, the enzyme that converts inactive vitamin D to its active form. In renal cells, PTH treatment results in decreased phosphorylation of SIK3 substrate CRTC2. The remaining intermediaries of the PTH/SIK/CYP27B1 pathway will be characterized using in vitro experiments targeting molecules upstream and downstream of SIK in renal cells and organoids. Chromatin Immunoprecipitation of CRTC family members will identify the regulatory regions necessary for CYP27B1 transcriptional activation by PTH/SIK. Aim 2 will define the role of PTH/SIK signaling in renal phosphate reabsorption. Mice treated with PTH or YKL-05-099 have decreased serum phosphate levels, and YKL-05-099 treatment decreases phosphate transporter Npt2a localization to the renal brush border membrane. These mice will be tested to confirm that SIK inhibition induces phosphaturia in vivo. Three SIK family members will be knocked out in renal epithelial cells and their effect on phosphate uptake assessed. Aim 3 will characterize the phenotype of mice lacking renal SIK1 and SIK3, and explore the therapeutic potential of small molecule SIK inhibitors in CKD-MBD. The experiments described here explore a novel role for SIKs in regulating PTH signaling in the kidney. In addition, mouse models developed for this project will help to shed light on the crucial bone/kidney regulatory axis that controls mineral homeostasis and bone health. This project also offers a novel application for SIK inhibitors, a class of drug currently studied for their application in autoimmune and other inflammatory disorders, as treatment in later stages of chronic kidney disease (CKD) and other disorders of mineral ion homeostasis that are unresponsive to PTH.

项目摘要甲状旁腺激素（PTH）通过与PTH/PTHRP受体相互作用来维持矿物质离子稳态诱导目标组织中的转录和转录后修饰。 PTH刺激吸收骨中的矿物基质，并增加活性维生素D，钙吸收和磷酸盐的产生肾脏排泄。尽管PTH是调节激素的中心作用，但许多下游编排PTH信号事件的细胞内靶标分子尚未确定。表征这些途径将扩大有关矿物质稳态机制的当前知识，并建议应对骨骼和肾脏疾病中的PTH作用的新治疗策略。最近，盐 - 诱导性激酶（SIKS）被鉴定为骨细胞中PTH信号的细胞内介体，而小 - 发现SIK的分子抑制剂模仿体内骨骼的PTH作用。该提议的目的是探索肾脏中类似的SIK依赖性PTH信号通路是否活跃。目标1 提案将探索PTH/SIK信号调节CYP27B1的分子机制表达。在肾细胞中，人类肾脏器官和小鼠，PTH和SIK抑制剂YKL-05-099 增加CYP27B1的表达，即将非活性维生素D转化为活性形式的酶。在肾细胞中， PTH处理导致SIK3底物CRTC2的磷酸化降低。其余的中介 PTH/SIK/CYP27B1途径将使用针对上游分子的体外实验来表征和肾细胞和器官中SIK的下游。 CRTC家庭成员的染色质免疫沉淀将确定PTH/SIK通过CYP27B1转录激活所必需的调节区域。 AIM 2意志定义PTH/SIK信号传导在肾磷酸盐重吸收中的作用。用PTH或YKL-05-099处理的小鼠具有血清磷酸盐水平降低，YKL-05-099治疗降低了磷酸转运蛋白NPT2A 定位到肾刷边框膜。这些小鼠将经过测试以确认SIK抑制作用会诱导体内磷酸。三名SIK家庭成员将在肾上皮细胞中被淘汰，它们对评估了磷酸盐的摄取。 AIM 3将表征缺乏肾脏Sik1和Sik3的小鼠的表型，以及探索CKD-MBD中小分子SIK抑制剂的治疗潜力。这里描述的实验探索SIKS在调节肾脏中PTH信号传导中的新作用。此外，为鼠标开发了该项目将有助于阐明控制矿物稳态的关键骨/肾脏调节轴和骨骼健康。该项目还为SIK抑制剂提供了新颖的应用程序，该抑制剂目前正在研究为了在自身免疫性和其他炎症性疾病中应用，作为慢性阶段的治疗肾脏疾病（CKD）和其他对PTH无反应的矿物离子稳态的疾病。