MTAP, 5'-deoxy-5'-methylthioadenosine, and the dysregulation of symmetric dimethylarginine in cancer

MTAP、5-脱氧-5-甲硫腺苷和癌症中对称二甲基精氨酸的失调

• 批准号: 10414804
• 负责人: WARREN D KRUGER
• 金额: $ 41.92万
• 依托单位: RESEARCH INST OF FOX CHASE CAN CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10414804
• 关键词: Adenosine; Affect; Antibodies; Arginine; Cell Line; Cell physiology; Cells; Cyclic AMP; Data; Enzymes; Epitopes; Family; Gene Expression Profile; Genes; Genetic; Genetic Screening; Goals; Hematologic Neoplasms; Housekeeping; Human; Lead; Light; Liquid substance; Location; Lye; Malignant Neoplasms; Mediating; Messenger RNA; Metabolic; Methionine; Modification; Multienzyme Complexes; Mutation; Pathway interactions; Pharmacology; Phosphorylases; Play; Process; Protein-Arginine N-Methyltransferase; Proteins; Purinergic P1 Receptors; Role; Signal Transduction; Solid; System; Testing; Tumor Suppressor Genes; Tumor Suppressor Proteins; Western Blotting; Work; cancer cell; dimethylarginine; extracellular; inhibitor; knock-down; migration; neoplastic cell; novel; novel strategies; novel therapeutic intervention; response; small hairpin RNA; tandem mass spectrometry; transcriptome sequencing; tumor; tumorigenesis

Project Summary Methylthioadenosine phosphorylase (MTAP) is a key enzyme in the methionine salvage pathway whose function is to convert 5′-deoxy-5′-methylthioadenosine (MTA) into methionine. Inactivation of MTAP, often by homozygous deletion, is found in both solid and hematologic malignancies and is one of the most frequently observed genetic alterations in human cancer. Previous work from our lab and others has established that MTAP can act as a tumor suppressor gene. However the precise mechanism by which MTAP loss promotes tumorigenesis is still unclear. One possible mechanism involves the accumulation of MTA, which is secreted by MTAP-deleted tumor cells. Structurally, MTA closely resembles adenosine and evidence indicates that it can interact with adenosine receptors. Large-scale genetic screens using shRNA have established that MTAP- deleted cells are especially sensitive to knockdown of a specific protein arginine methyltransferase enzyme (PRMT5), which is responsible for the post-translational symmetric dimethylation modification of arginine residues (sDMA). This modification is frequently observed in proteins involved in mRNA maturation. In preliminary data, we demonstrate that loss of MTAP or addition of extracellular MTA causes a dramatic reduction of the steady-state levels of sDMA-containing proteins. Significantly, when extracellular MTA is added, no increase in intracellular MTA occurs, suggesting that the reduction in sDMA-ylation is due to a signal transduction process. However, our data also suggests that enzymatically inactive MTAP protein itself, independent of MTA, can affect mRNA levels and antagonize the effects of MTA. Thus the overall goal of this proposal is to explore the hypothesis MTAP mediates its tumor suppressor function via two different mechanisms: one involving MTA as an oncometabolite and the other a direct role for the MTAP protein. The specific aims of the study are: (1) Identify specific proteins and arginine residues that are differentially methylated in response to MTAP; (2) Determine the roles of intracellular and extracellular MTA in the mechanism of sDMA-lyation in cancer cells; and (3) Clarification of the enzymatic vs. non-enzymatic functions of MTAP. These studies are significant because they provide a potential mechanism for understanding the role that MTAP deletion plays in tumorigenesis and may lead to novel therapeutic strategies for MTAP-deleted tumors. In addition, these studies will shed light on how a “housekeeping” metabolic enzyme can have a novel role as a tumor suppressor gene.

项目摘要 甲基己醇磷酸化酶（MTAP）是蛋氨酸拯救途径的关键酶，其功能 是将5'-脱氧5'-甲基硫代腺苷（MTA）转化为甲基氨酸。 MTAP失活，通常是 纯合缺失，在固体和血液系统恶性肿瘤中都发现，并且是最常见的一种 观察到人类癌症的遗传改变。我们实验室和其他人的先前工作已经确定了MTAP 可以充当肿瘤抑制基因。但是，MTAP损失促进的精确机制 肿瘤发生尚不清楚。一种可能的机制涉及MTA的积累，该机制由 MTAP缺失的肿瘤细胞。在结构上，MTA与腺苷非常相似，证据表明它可以 与腺苷接收器互动。使用SHRNA的大规模遗传筛选已经确定 删除的细胞对特定蛋白精氨酸甲基转移酶的敲低特别敏感 （prmt5），负责精氨酸的翻译后对称二甲基化修饰 残留物（SDMA）。在参与mRNA成熟的蛋白质中经常观察到这种修饰。在 初步数据，我们证明了MTAP的丢失或添加细胞外MTA会引起巨大的减少 含SDMA的蛋白质的稳态水平。值得注意的是，当添加细胞外MTA时，否 细胞内MTA的增加发生，这表明SDMAyLation的降低是由于信号转导引起的 过程。但是，我们的数据还表明，酶无活性MTAP蛋白本身，独立于MTA， 可以影响mRNA水平并拮抗MTA的作用。该提议的总体目标是探索 该假设MTAP通过两种不同的机制介导了其肿瘤抑制功能：一种涉及MTA 作为oncometabolite，另一个是MTAP蛋白的直接作用。该研究的具体目的是：（1） 鉴定特定的蛋白质和精氨酸保留，这些蛋白会响应于MTAP而甲基化的甲基化。 （2） 确定细胞内和细胞外MTA在癌细胞中SDMA-裂解机理中的作用；和 （3）阐明MTAP的酶促与非酶函数。这些研究很重要，因为 它们提供了了解MTAP缺失在肿瘤发生和 可能导致新的MTAP骨骼肿瘤的热策略。此外，这些研究将阐明 “家政”代谢酶如何作为肿瘤抑制基因具有新颖的作用。