Regulation of epigenome stability by SIRT6 during Aging

该项目的长期目标是确定调控表观基因组的分子机制 衰老过程中的稳定性以及哺乳动物sirtuin，SIRT 6在这一过程中的作用。越来越多的证据表明 表观基因组结构随着年龄的增长而受损，这可能是与年龄相关的衰退的根本原因。 细胞和器官功能。SIRT 6作为与衰老相关的多种途径的关键调节剂出现，例如 基因组和表观基因组稳定性、肿瘤发生、炎症和糖酵解。此外，SIRT 6 过表达延长了小鼠的寿命。 我们的实验室已经证明SIRT 6是DNA双链断裂（DSB）的上游调节因子 修复.我们证明，SIRT 6是由JNK 1/2磷酸化，以响应氧化应激对氨基酸 S10的磷酸化是刺激DSB修复所必需的。我们已经证明，除了 SIRT 6控制DSB修复，通过抑制转座元件维持基因组稳定性，并且 氧化应激导致SIRT 6从转座因子的启动子重新定位到DNA断裂。 SIRT 6具有两种生物化学活性脱乙酰酶（deacetylase）和单ADP核糖基酶。功能 SIRT 6脱乙酰酶活性的最佳表征。相比之下，SIRT 6的单ADP核糖基化活性更高， 研究较少。我们的工作表明这种活性与DNA修复和表观基因组稳定性有关。我们最近未发表的 分析百岁老人SIRT 6变异体生化功能的数据显示，百岁老人SIRT 6 具有降低的脱乙酰化活性和增强的单ADP核糖基化活性。这表明SIRT 6 mono-ADP核糖基化活性对于长寿是重要的。因此，我们着手确定功能和新的 SIRT 6单ADP核糖基化的靶点。使用质谱法，我们确定了SIRT 6单克隆抗体的新靶点， ADP核糖基化包括组蛋白H1、H2 A、H2A.J和染色质重塑物BRG 1和SMARCC 2。 此外，我们发现SIRT 6介导的SMARCC 2的核糖基化是激活Nrf 2靶标所必需的。 基因对氧化应激的反应。 因此，我们理想地定位于对SIRT 6单ADP的作用进行进一步的机制研究。 表观基因组稳定性中的核糖基化活性。我们将追求以下具体目标：（1）研究 SIRT 6在细胞衰老和老化背景下维持表观基因组稳定性中的作用;（2）检查SIRT 6在细胞衰老和老化背景下维持表观基因组稳定性的作用。 SIRT 6对表观基因组的作用机制;具体而言，H1、H2 A、H2A.J、BRG 1和 （3）确定SIRT 6单ADP核糖基化活性在SMARCC 2单ADP核糖基化中的作用。 通过分析表达核糖基化缺陷的基因敲入小鼠模型， SIRT 6突变。这项拟议的研究将描绘SIRT 6调控的新途径，这些途径与以下方面有关： 表观基因组稳定性和老化。因此，我们希望这些实验将揭示关键的新信息， 关于衰老过程，并将有助于开发治疗与年龄有关的疾病的新策略。

The long-term goal of this project is to define the molecular mechanisms which regulate epigenome stability during aging and the role of the mammalian sirtuin, SIRT6, in this process. Growing evidence indicates that epigenome structure becomes compromised with age which may be the root cause of age-related decline in cell and organ function. SIRT6 emerged as a critical regulator of multiple pathways related to aging such as genome and epigenome stability, tumorigenesis, inflammation and glycolysis. Additionally, SIRT6 overexpression extends the lifespan of mice. Our laboratory has demonstrated that SIRT6 is an upstream regulator of DNA double strand break (DSB) repair. We demonstrated that SIRT6 is phosphorylated by JNK1/2 in response to oxidative stress on amino acid S10 and this phosphorylation is required for the stimulation of DSB repair. We have shown that, in addition to controlling DSB repair, SIRT6 maintains genome stability by repressing transposable elements, and that oxidative stress causes re-localization of SIRT6 from the promoters of transposable elements to the DNA breaks. SIRT6 has two biochemical activities deacetylase (deacetylase) and mono-ADP ribosylase. The function of SIRT6 deacetylase activity is best characterized. In contrast, mono-ADP ribosylation activity of SIRT6 is much less studied. Our work has implicated this activity in DNA repair and epigenome stability. Our recent unpublished data analyzing biochemical functions of a SIRT6 variant found in centenarians, showed that centenarian SIRT6 has reduced deacetylation activity and enhanced mono-ADP ribosylation activity. This suggests that SIRT6 mono-ADP ribosylation activity is important for longevity. Therefore, we set out to identify the function and new targets of SIRT6 mono-ADP ribosylation. Using mass spectrometry we identified novel targets for SIRT6 mono- ADP ribosylation including histone H1, H2A, H2A.J, and chromatin remodelers BRG1 and SMARCC2. Furthermore, we showed that SIRT6-mediated ribosylation of SMARCC2 is required for activation of Nrf2 target genes in response to oxidative stress. Thus, we are ideally positioned to conduct further mechanistic studies of the role of SIRT6 mono-ADP ribosylation activity in epigenome stability. We will pursue the following specific aims: (1) examine the role of SIRT6 in maintaining epigenome stability in the context of cellular senescence and aging; (2) examine the mechanisms of SIRT6 effect on epigenome; specifically, the biological function of H1, H2A, H2A.J, BRG1 and SMARCC2 mono-ADP ribosylation; and (3) determine the role of SIRT6 mono-ADP ribosylation activity in epigenome stability and longevity by analyzing the knock-in mouse model which expresses ribosylation deficient SIRT6 mutation. The proposed research will delineate new pathways regulated by SIRT6, which are relevant to epigenome stability and aging. As such, we expect that these experiments will reveal critical, new information about the aging process, and will help to develop new strategies for treating age-related diseases.