Mechanism of Disease-causing mutations in PCNA

PCNA 致病突变机制

• 批准号: 10699962
• 负责人: Brian Anthony Kelch
• 金额: $ 38.37万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-07 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10699962
• 关键词: Address; Affect; Affinity; Alleles; Antigen Targeting; Apoptosis; Automobile Driving; Binding; Binding Sites; Biochemical; Biological Assay; Cell Cycle Regulation; Cell Proliferation; Cell physiology; Cells; Chromatin; Chromatin Modeling; Chromatin Structure; Clinical; Closure by clamp; Complex; DNA; DNA Integration; DNA Repair; DNA Repair Disorder; DNA Repair Enzymes; DNA Repair Gene; DNA Repair Inhibition; DNA Repair Pathway; DNA biosynthesis; Defect; Development; Diagnosis; Disease; Dissociation; Enzymes; Epigenetic Process; Fibroblasts; Genetic Recombination; Health; Human; Ligands; Longevity; Malignant Neoplasms; Manuscripts; Measures; Mediating; Molecular; Molecular Conformation; Molecular Machines; Mutation; NMR Spectroscopy; Neoplasm Metastasis; Pathway interactions; Patients; Phenotype; Point Mutation; Positioning Attribute; Process; Proliferating Cell Nuclear Antigen; Proteins; Rare Diseases; Role; Scaffolding Protein; Shapes; Skin Cancer; Slide; Structure; Symptoms; Therapeutic; Thermodynamics; Time; Variant; Work; X-Ray Crystallography; antigen binding; autosome; biophysical techniques; cell behavior; design; disabling symptom; disease-causing mutation; early onset; experience; genome integrity; human disease; insight; multidisciplinary; novel therapeutic intervention; premature; protein folding; protein function; rare genetic disorder; repaired; replication factor A; residence; small molecule; therapeutic target; therapy outcome; treatment strategy; tumor progression

Abstract PCNA is a critical regulator and facilitator of many cellular processes such as DNA replication, DNA repair, recombination, chromatin structure and apoptosis. PCNA is a ring-shaped complex that acts as a sliding platform on DNA for the arrangement of scores different proteins to assemble and act on chromatin. Abnormal PCNA activity is associated with the development and metastasis of cancer; consequently, PCNA is a target for development of chemotherapeutics. Two separate point mutations (Ser228Ile and Cys148Ser) in PCNA cause an autosomal recessive disorder (PCNA Associated DNA Repair Disorder or PARD) that results from defects in DNA repair. We hypothesize that these mutations disrupt PCNA’s stability and/or ability to bind to partner proteins. We further hypothesize that the PCNA defect in folding stability results in a shorter lifetime on DNA, thereby preferentially inhibiting DNA repair proteins that function near the end of PCNA’s lifetime on DNA. Our preliminary studies show that the S228I mutation disrupts the binding site for PCNA partners, yet some partners overcome this disruption to bind PCNA. We hypothesize that the S228I mutation disrupts the structure and dynamics of the binding site such that the DNA repair pathway is disproportionately perturbed. We further find that both PARD mutations disrupt PCNA stability, which could decrease the lifetime of PCNA on DNA. In support of this hypothesis, we find that PCNA levels on chromatin are abnormally low in patient-derived fibroblast cells. We will address these hypotheses with three specific aims: (1) To determine how the PARD mutations alter PCNA structure, stability, and dynamics, (2) to determine the biochemical effects of PARD variants on PCNA longevity and on partner binding and activity, and (3) to identify which cellular factors and pathways are disrupted. Our studies will determine the robustness or fragility of PCNA-mediated pathways such as DNA repair and DNA replication, which will define how PCNA affects cancer. Our work will also uncover how PCNA structure and dynamics controls partner binding, which will guide efforts to develop small molecules that disrupt specific PCNA-mediated.

抽象的 PCNA是许多细胞过程的关键调节剂，也是 DNA复制，DNA修复，重组，染色质结构和凋亡。 PCNA是一种环形复合物，充当DNA的滑动平台 分数的排列不同的蛋白质以组装和作用于染色质。 PCNA活性异常与发展和转移有关 癌症;因此，PCNA是化学治疗剂开发的靶标。 PCNA中的两个单独的点突变（Ser228ile和Cys148ser）导致 常染色体隐性障碍（PCNA相关的DNA修复障碍或PARD） 这是由于DNA修复缺陷所致。我们假设这些突变 破坏PCNA的稳定性和/或与伴侣蛋白结合的能力。我们进一步 假设折叠稳定性中的PCNA缺陷导致寿命较短 在DNA上，优先抑制在附近起作用的DNA修复蛋白 PCNA在DNA上的生命结束。我们的初步研究表明S228i 突变破坏了PCNA合作伙伴的绑定站点，但一些合作伙伴 克服这种破坏以结合PCNA。我们假设S228i突变 破坏结合位点的结构和动力学，使DNA修复 途径不成比例地扰动。我们进一步发现两个 突变破坏PCNA稳定性，这可能会降低PCNA的寿命 脱氧核糖核酸。为了支持这一假设，我们发现染色质上的PCNA水平是 交替使用患者衍生的成纤维细胞。 我们将以三个特定的目的解决这些假设：（1）确定如何 PARD突变改变了PCNA结构，稳定性和动力学，（2）至 确定帕德变体对PCNA寿命和对PCNA寿命的生化效应 伴侣结合和活动，（3）确定哪些细胞因素和 途径被干扰。我们的研究将确定 PCNA介导的途径，例如DNA修复和DNA复制，将 定义PCNA如何影响癌症。我们的工作还将发现PCNA的结构如何 动态控制合作伙伴的绑定，这将指导努力发展小型 破坏特定PCNA介导的分子。