Identification and characterization of functional genetic variants associated with human longevity

摘要 人类遗传学和基因组学的最新进展为鉴定和鉴定 药物靶标的确认。在模型生物中的研究表明，衰老的速度和 年龄相关疾病的频率和严重程度受到保守的遗传途径和因素的影响 当从遗传学、药理学或饮食调节方面进行靶向时，它可以延长寿命和 健康跨度。这一压倒性的证据为延缓衰老过程和减轻衰老的新药带来了希望。 通过调节这些保守的衰老途径，人类中与年龄相关的疾病。在项目2(旧项目1)中， 我们一直在进行的实验正是为了这个目标：识别和功能特征 与人类健康衰老和极端衰老相关的保守衰老途径中的遗传变异 延年益寿用于治疗调整，以提高人类的健康寿命和寿命。我们通过细胞演示了 SIRT6、USP35和UBE3C中与长寿相关的罕见编码变体改变蛋白质功能和/或 在人体组织和细胞衰老过程中对抗与年龄相关的有害变化的表达。 我们在Smad3和Smad3中发现了与长寿相关的非编码变体相同的抗衰老作用 参与PKC/NF-B信号通路的基因有NFKBIA、CLU和PRKCH。在续签申请中， 项目2建议通过采取系统的多学科方法来证实和扩大我们的观察结果。 项目2的目标是确定并直接测试其遗传扰动的因果变异的影响 这是与长寿有关的基础。我们的方法是使用高通量筛选方法，例如 编码变异和大规模平行的整合蛋白质相互作用组扰动筛查(InPOINT) 非编码变体的报告程序分析(MPRA)，并使用CRISPR-来研究潜在的机制- 工程化人多能干细胞，多细胞分化范式，多组学 方法，以及由分子、生化和细胞分析组成的功能分析。我们的职能部门 对SIRT6、USP35和UBE3C中与长寿相关的变异的分析表明，它们在保护中具有潜在的作用 抗老年性认知功能减退和阿尔茨海默病风险及细胞表型的研究 衰老，它越来越多地与神经退行性疾病有关。因此，我们将重点关注老龄化- SIRT6、UBE3C和USP35编码变体在认知健康和治疗中的拮抗作用 对AD的防护。项目2的最终影响在于它可能揭示保守的路径，如 人类衰老的基本机制，并作为健康衰老的治疗目标，密切合作 项目1、3、4和核心B。

Abstract Recent advances in human genetics and genomics provide novel opportunities for the identification and validation of drug targets. Studies in model organisms have demonstrated that the rate of aging and the frequency and severity of age-related pathologies are influenced by conserved genetic pathways and factors which, when targeted genetically, pharmacologically, or through dietary modulation, can extend lifespan and healthspan. This overwhelming evidence raises hopes for new drugs that slow the aging process and attenuate age-related disease in humans by modulating these conserved pathways of aging. In Project 2 (old Project 1), we have been conducting the experiments aimed at exactly this goal: To identify and functionally characterize genetic variants in the conserved pathways of aging that are associated with human healthy aging and extreme longevity for therapeutic modulation to improve human healthspan and lifespan. We demonstrated through cell models that longevity-associated rare coding variants in SIRT6, USP35, and UBE3C alter protein function and/or expression which antagonize age-related deleterious changes in human tissues and during cellular senescence. We found the same aging-antagonizing effects of longevity-associated non-coding variants in SMAD3 and the 3 genes (NFKBIA, CLU and PRKCH) involved in the PKC/NF-B signaling pathways. In the renewal application, Project 2 proposes to confirm and extend our observations by taking a systematic multidisciplinary approach. The objective of Project 2 is to identify and directly test the impact of causal variants, whose genetic perturbations underlie the association with longevity. Our approach is to use high-throughput screening methods, such as INtegrated PrOtein INteractome perTurbation screening (InPOINT) for coding variants and Massively Parallel Reporter Assay (MPRA) for non-coding variants, and to investigate the underlying mechanisms using CRISPR- engineered human pluripotent stem cells (hPSC), multiple cellular differentiation paradigms, multiomics approaches, and functional analysis composed of molecular, biochemical, and cellular assays. Our functional analyses of longevity-associated variants in SIRT6, USP35, and UBE3C point to their potential role in protection against age-related cognitive decline and risk of Alzheimer’s disease (AD) and on the phenotypes of cellular senescence, which are increasingly implicated in neurodegenerative disease. Thus, we will focus on the aging- antagonizing effects of the SIRT6, UBE3C, and USP35 coding variants in the context of cognitive health and protection from AD. The ultimate impact of Project 2 lies in its potential to reveal conserved pathways as fundamental mechanisms of aging in humans and as therapeutic targets for healthy aging, in close collaboration with Projects 1, 3, 4 and Core B.