Characterization of TMEM164 as novel multi-pass transmembrane enzyme and its role in ferroptosis

TMEM164作为新型多次跨膜酶的表征及其在铁死亡中的作用

• 批准号: 10607957
• 负责人: Timothy Brandon Ware
• 金额: $ 6.95万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10607957
• 关键词: Acyltransferase; Apoptosis; Attenuated; Biochemistry; Biological Assay; Cancer Model; Cell Death; Cell Death Induction; Cell membrane; Cells; Cellular biology; Cessation of life; Chemicals; Chemoresistance; Coenzyme A Ligases; Collaborations; Coupled; Cysteine; Data; Degenerative Disorder; Diabetes Mellitus; Diabetic Nephropathy; Disease; Drug Targeting; Endothelium; Enzymes; Esters; Ethers; Family; Family member; Fatty Acids; Funding; Genes; Genetic; Goals; Homeostasis; Human; Hyperglycemia; Injury; Integral Membrane Protein; Iron; Iron Overload; Kidney; Libraries; Link; Lipid Peroxidation; Lipid Peroxides; Lipids; Lysophosphatidylcholines; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Membrane; Metabolic; Metabolic Pathway; Methods; Non-Insulin-Dependent Diabetes Mellitus; Orphan; Orthologous Gene; Pharmaceutical Chemistry; Phospholipid Metabolism; Phospholipids; Physiological; Play; Polyunsaturated Fatty Acids; Predisposition; Principal Investigator; Production; Proteins; Proteomics; Recombinant Proteins; Recombinants; Renal Cell Carcinoma; Research Personnel; Risk; Role; Severities; Signal Transduction; Structure; System; Testing; Threonine; Training; Tubular formation; Variant; attenuation; desensitization; diabetic; drug candidate; experience; fatty acid metabolism; genome editing; glutathione peroxidase; human disease; hydroxy fatty acid; inhibitor; kidney cell; kidney vascular structure; lipid metabolism; lipidomics; mutant; new therapeutic target; novel; oxidation; pharmacologic; premature; small molecule; therapeutic target

Project Summary/Abstract The goal of this proposal is to understand how transmembrane protein 164 (TMEM164) attenuates polyunsaturated fatty acid (PUFA) metabolism and ferroptotic signaling. Ferroptosis is a novel iron-dependent form of cell death implicated in a broad range of human diseases, including cancer and type 2 diabetes mellitus (T2DM). In T2DM, the severity of diabetic state increases risk for renal tubular injury related to iron overload and lipid peroxidation, two key features of ferroptosis. Characterizing enzymes that regulate the PUFA content of human cells would not only deepen our understanding of ferroptosis, but also identify therapeutic targets for treating diseases like T2DM where this form of cell death is dysregulated. Recent genome editing screens have identified TMEM164, a multi-pass transmembrane protein of uncharacterized function, as a key regulator of ferroptosis. We hypothesize, based on its Alphafold-predicted structure, that TMEM164 is a novel type of cysteine-dependent transmembrane enzyme that regulates cellular PUFA content. Here we seek to test this hypothesis and identify the specific enzymatic functions performed by TMEM164 in PUFA lipid metabolism, as well as their contribution to conferring ferroptosis sensitivity to human cells. In Aim 1, I will determine the lipid profiles of cells genetically deleted for TMEM164 and test whether wild type, but not a putative catalytic cysteine mutant form of TMEM164 can rescue these lipid perturbations. From these data, I will define candidate physiological substrates for TMEM164, which will be tested in recombinant protein systems. In Aim 2, I will leverage our lab’s longstanding expertise in covalent inhibitor discovery to identify electrophilic compounds that block TMEM164 activity through modifying the putative catalytic cysteine of the protein. Successful completion of this project will broaden our fundamental understanding of the lipid metabolic pathways involved in ferroptosis and identify new candidate drug targets for suppressing the contribution of ferroptosis to human degenerative diseases like T2DM.

项目总结/摘要 该提案的目标是了解跨膜蛋白164（TMEM 164）如何减弱 多不饱和脂肪酸（PUFA）代谢和铁蛋白信号传导。铁蛋白沉积症是一种新的铁依赖性 细胞死亡的形式与多种人类疾病有关，包括癌症和2型糖尿病 （T2DM）。在T2 DM中，糖尿病状态的严重程度增加了与铁过载相关的肾小管损伤的风险， 脂质过氧化是铁缺乏症的两个关键特征。表征调节PUFA含量的酶 人类细胞不仅加深了我们对铁凋亡的理解，而且还确定了治疗靶点， 治疗像T2 DM这样的疾病，其中这种形式的细胞死亡失调。最近的基因组编辑屏幕 鉴定了TMEM 164，一种功能不明的多通道跨膜蛋白，作为一种关键的调节因子， 铁性下垂基于其Alphafold预测的结构，我们假设TMEM 164是一种新型的 调节细胞PUFA含量的半胱氨酸依赖性跨膜酶。在这里我们试图验证这一点 假设并确定TMEM 164在PUFA脂质代谢中执行的特定酶功能， 以及它们对赋予人细胞铁凋亡敏感性的贡献。在目标1中，我将确定脂质 TMEM 164基因缺失的细胞谱，并测试是否为野生型，但不是推定的催化半胱氨酸 TMEM 164的突变形式可以挽救这些脂质扰动。从这些数据中，我将定义候选人 TMEM 164的生理底物，将在重组蛋白系统中进行测试。在目标2中，我将 利用我们实验室在共价抑制剂发现方面的长期专业知识， 通过修饰蛋白质的推定催化半胱氨酸来阻断TMEM 164活性。成功完成 本项目的研究将拓宽我们对铁凋亡中脂质代谢途径的基本理解 并确定新的候选药物靶点，用于抑制铁凋亡对人类退行性疾病的贡献， T2 DM等疾病。