Non-canonical ERAD as a Regulator of Cardiac Hypertrophy

非典型 ERAD 作为心脏肥大的调节剂

• 批准号: 10817347
• 负责人: Chris Glembotski
• 金额: $ 4.01万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10817347
• 关键词: Binding; Binding Proteins; Cardiac; Cardiac Myocytes; Cardiovascular Diseases; Cessation of life; Ectopic Expression; Endoplasmic Reticulum; Endoplasmic Reticulum Degradation Pathway; Glucocorticoids; Growth; Heart; Heart Diseases; Heart Hypertrophy; Heart failure; Interruption; Knowledge; Leucine Zippers; Mediating; Methods; Molecular; Mus; Pathologic; Pathology; Peptides; Phosphotransferases; Protein Biosynthesis; Protein Overexpression; Proteins; Role; Serum; Serum Proteins; Sgk protein; Signal Transduction; System; Therapeutic; cancer cell; cell type; innovation; knock-down; misfolded protein; mortality; multicatalytic endopeptidase complex; novel; pressure; protein folding

PROJECT SUMMARY Increases in protein synthesis during pathological cardiac hypertrophy places demands on protein-folding machinery to avert the accumulation of toxic misfolded proteins. Associated with the endoplasmic reticulum (ER), where many important proteins are synthesized in cardiac myocytes, is a system that recognizes and degrades misfolded proteins, i.e. ER associated (protein) degradation, or ERAD. We discovered a different, non-canonical role for ERAD as a regulator of the levels of the growth-promoting kinase, serum glucocorticoid kinase 1 (SGK1). SGK1 is a cytosolic kinase involved in growth of other cell types, such as cancer cells. Interestingly, SGK1 can traffic to the ER where it is ubiquitylated by ERAD machinery and subsequently degraded by cytosolic proteasomes; however, this unique regulatory mechanism has not been studied in the heart. Our preliminary evidence shows that non-canonical ERAD could regulate SGK1 levels in the heart and, thereby regulating cardiac growth under pathological conditions. We also found that an SGK1-binding protein, called glucocorticoid-inducible leucine zipper protein (GILZ), can bind to and protect SGK1 from non-canonical ERAD-mediated degradation, which we believe increases SGK1-mediated growth of the heart during pressure overload. Accordingly, our hypothesis is that SGK1 is a major inducer of pressure overload-induced cardiac pathology. During pressure overload, SGK1 levels, and thus, SGK1-mediated cardiac hypertrophy and subsequent pathology, are increased by GILZ-dependent diversion of SGK1 away from the ER, which decreases SGK1 degradation by non-canonical ERAD. Ectopic expression of an SGK1 peptide disrupts the GILZ-SGK1 interaction, increases SGK1 degradation, thus decreasing SGK1-mediated cardiac hypertrophy and subsequent pathology. This hypothesis will be examined in mice subjected to pressure overload-induced cardiac pathology in our specific aims, which are to 1-couple cardiac-specific SGK1 deletion with AAV9 encoding SGK1-WT (active; ER-targeted), SGK1-KD (kinase-dead ER-targeted) or SGK1-∆60 (active; not ER-targeted) to examine the effect of SGK1 and ERAD on overload-induced cardiac pathology, 2-combine AAV9-SGK1-WT or AAV9- SGK1-∆60 with AAV9-mediated GILZ overexpression or knockdown to determine whether GILZ diverts SGK1- WT from the ER and protects it from ERAD in the heart, 3-evaluate the potential therapeutic, antihypertrophic effects of a novel SGK1 peptide that interrupts GILZ-SGK1 binding, and increases non-canonical ERAD- mediated SGK1 degradation. These studies are significant because they will reveal previously unappreciated roles for SGK1, GILZ and ERAD in pathologic cardiac hypertrophy. We will use an innovative molecular strategy to mechanistically dissect roles for GILZ and non-canonical ERAD as regulators of SGK1 signaling and cardiac pathology. Peptide-based disruption of the SGK1-GILZ interaction could be a highly specific method for inhibiting the maladaptive pathological effects of SGK1 in the heart by selective degradation of SGK1.

项目摘要 病理性心肥大的蛋白质合成的增加对蛋白质折叠的需求 避免有毒错误折叠蛋白的积累的机械。与内质网 （ER），许多重要的蛋白质在心肌细胞中合成，是一个识别和 降解错误折叠的蛋白质，即ER相关（蛋白质）降解或Erad。我们发现了一个不同的 ERAD作为促进生长激酶水平血清糖皮质激素水平的调节剂的非传统作用 激酶1（SGK1）。 SGK1是一种参与其他细胞类型（例如癌细胞）生长的胞质激酶。 有趣的是，SGK1可以通过ERAD机械和随后的急诊室流动。 被胞质蛋白酶体降解；但是，这种独特的调节机制尚未研究 心。我们的初步证据表明，非典型的ERAD可以调节心脏中的SGK1水平，并且，，，，，， 从而调节病理条件下的心脏生长。我们还发现SGK1结合蛋白， 称为糖皮质激素诱导的亮氨酸拉链蛋白（GILZ）可以结合并保护SGK1免受非典型的影响 ERAD介导的降解，我们认为这会增加SGK1介导的心脏在压力下的生长 超载。据此，我们的假设是SGK1是压力超负荷引起的心脏的主要诱导剂 病理。在压力超负荷，SGK1水平，因此SGK1介导的心脏肥大和 随后的病理学通过gilz依赖的sgk1转移而增加，从ER降低了 SGK1通过非典型的ERAD降解。 SGK1辣椒的异位表达破坏了Gilz-SGK1 相互作用，增加SGK1降解，从而降低SGK1介导的心脏肥大，随后 病理。该假设将在受压超负荷引起的心脏病理的小鼠中进行检查 在我们的特定目的中，用AAV9编码sgk1-wt，是1套偶心特异性SGK1删除 （活跃；靶标），sgk1-kd（激酶死于靶标）或sgk1-∆60（活动;未靶向ER）以检查 SGK1和ERAD对超载诱导的心脏病理，2-Combine AAV9-SGK1-WT或AAV9-的影响 SGK1-∆60具有AAV9介导的GILZ过表达或敲低，以确定GILZ是转移SGK1- wt从急诊室中保护并保护其免受心脏的侵蚀，3-评估潜在的疗法，抗营养 新型的SGK1胡椒的影响，它会中断gilz-sgk1结合，并增加非典型的eRAD- 介导的SGK1降解。这些研究很重要，因为它们会揭示先前未欣赏的 SGK1，GILZ和ERAD在病理心脏肥大中的作用。我们将使用创新的分子策略 为了机械剖析吉尔兹和非规范剥离的作用，作为SGK1信号和心脏的调节剂 病理。 SGK1-GILZ相互作用的基于肽的破坏可能是抑制高度特异性的方法 SGK1的选择性降解，SGK1在心脏中的不良病理性作用。