Functional Characterization and Physiological Consequences of Human Longevity-Associated IGF1R Variants

 描述(申请人提供)：尽管有压倒性的证据表明胰岛素/胰岛素样生长因子-1(IGF1)信号转导减少(IIS)在无脊椎动物和小鼠模型中延长了寿命，但这一进化保守的途径对人类寿命的影响尚不清楚。Suh实验室的研究导致在IGF-1受体基因(IGF1R)中发现了两个百岁富集型错义突变A37T(M1)和R407H(M2)，与非携带者建立的淋巴细胞中IIS降低有关(Suh等人，PNAS，2008年)。与野生型人IGF1R相比，M1和M2变异体在Igf1r缺失的小鼠胚胎成纤维细胞中表达时，可导致IGF1信号减弱、IGF1激活基因表达减少以及细胞周期进展延迟(Tazearslan等人，Aging Cell，2011)。虽然IIS与长寿之间的联系是已知的，但IIS在调节人类寿命方面的具体作用，包括IIS组件的错义突变如何改变生存和疾病风险，尚不清楚。这项建议的目的是利用表达百岁富集型变异受体的细胞和携带百岁相关变异的小鼠来阐明IGF-1受体错义突变促进长寿的分子和生理机制。IGF-1受体已经在小鼠身上进行了关于衰老的研究，但只在基因敲除或杂合子缺乏的情况下进行。IGF-1受体在细胞和生物体的生理调节中具有多种作用，包括非受体功能和通过二聚化修饰其他膜结合受体。此外，IGF-1受体的刺激通过特定的配体-受体相互作用激活了多个不同的细胞内信号级联反应，我们的初步数据表明，IGF-1受体的百岁相关突变以组织特有的方式不同地影响这些下游信号通路。为了解决这一目标，我们将(1)在基于细胞的分析中检测IGF1R变体的处理、组装、蛋白质间的相互作用以及对信号转导的影响；(2)利用我们制作的敲入模型，确定与百岁老人相关的IGF1R变体对小鼠的生长、代谢和体内胰岛素/IGF-1信号的影响；(3)确定与百岁老人相关的IGF1R变体对高脂饮食反应的影响，该模型是年龄相关代谢性疾病的模型。我们假设，百岁老人相关的IGF1R变异对细胞和生物体生理学有功能上的相关影响，这是由于蛋白质功能的变化造成的。我们进一步假设，这些作用在功能上部分不同于简单地降低IGF-1受体水平，包括对IGF1R下游信号转导和非受体功能的不同影响。最后，我们预测，这些分子变化和细胞效应将导致敲入小鼠模型中有益的系统功能变化，这是与人类长寿相关的基础，包括在高脂饮食范例中对代谢应激的抵抗。了解自然发生的百岁老人相关IGF1R基因变异的生理作用，将为未来基于我们描述的分子机制预防和治疗年龄相关疾病的治疗策略的发展铺平道路。

 DESCRIPTION (provided by applicant): While there is overwhelming evidence that reduced insulin/insulin like growth factor-1 (IGF1) signaling (IIS) extends lifespan in invertebrate and murine models, the impact of this evolutionarily conserved pathway on human longevity remains unclear. Research in the Suh lab has led to the discovery of two centenarian- enriched missense mutations in the IGF-1 receptor gene (IGF1R), A37T (M1) and R407H (M2), associated with decreased IIS in lymphocytes established from carriers as compared to non-carriers (Suh et al., PNAS, 2008). The M1 and M2 variants were shown to cause attenuation of IGF1 signaling, reduced expression of IGF1-activated genes, and delayed cell cycle progression relative to wild-type human IGF1R when expressed in Igf1r null mouse embryonic fibroblasts (Tazearslan et al., Aging Cell, 2011). While the association of IIS with longevity is known, the specific role of IIS in regulating human lifespan, including how missense mutations in IIS components modify survival and disease risk, is not known. The goal of this proposal is to elucidate the molecular and physiological mechanisms of longevity promotion by missense mutations in the IGF-1 receptor using cells expressing centenarian enriched variant receptor and mice carrying a knock-in of a centenarian associated variant. The IGF-1 receptor has been studied in mice in respect to aging but only in the context of knockout, or heterozygous deficiency. The IGF-1 receptor has multiple roles in regulating cellular and organism physiology including non-receptor functions and modification of other membrane bound receptors through dimerization. In addition, stimulation of the IGF-1 receptor activates multiple distinct intracelluar signaling cascades through specific ligand-receptor interactions and our preliminary data shows that centenarian associated mutations in IGF-1 receptor differentially affect these downstream signaling pathways in a tissue specific manner. In order to address this goal we will (1) examine IGF1R variant processing, assembly, protein-protein interaction, and impact on signaling in cell based assays; (2) define the effects of a centenarian associated IGF1R variant on growth, metabolism, and in vivo insulin/IGF-1 signaling in mice using the knock-in model we have produced; (3) determine the impact of the centenarian associated IGF1R variant on the response to high- fat diet, a model for age-related metabolic disease. We hypothesize that centenarian associated IGF1R variants have functionally relevant effects on cellular and organism physiology resulting from changes to protein function. We further hypothesize that these effects will be partly functionally distinct from simply reducing levels of IGF-1 receptor, including differential effects on downstream signaling and non-receptor functions of IGF1R. Lastly, we predict that these molecular changes and cellular effects will lead to beneficial systemic functional changes in the mouse knock-in model that underlie the association to human longevity, including resistance to metabolic stress in the high-fat diet paradigm. Understanding the physiological role of naturally occurring centenarian associated IGF1R genetic variation will pave the way for future development of therapeutic strategies for preventing and treating age-related disease based on the molecular mechanisms we describe.