Project Summary/Abstract Title: Insulin regulates aging-associated genes through chromatin topology Insulin signaling plays an important role in controlling metabolic homeostasis, with its dysregulation linked to various metabolic diseases associated with aging, including type 2 diabetes (T2D) and obesity. Inhibiting insulin signaling has been shown to prolong lifespan and postpone aging-associated diseases across various animal species. However, the mechanism by which insulin activity controls aging and related metabolic syndromes remain unclear. In mouse liver, a significant portion of insulin-regulated genes are associated with aging. Among them, the expression level of Tmtc2 (transmembrane and tetratricopeptide repeat containing protein 2), a novel Ca2+ regulator, significantly declines with aging, insulin treatment and insulin-associated metabolic conditions. This project has three objectives. 1) Aim 1.1 (F99): This project determines how Tmtc2 controls cell response to stress and senescence through Ca2+ homeostasis. Tmtc2 will be knocked down followed by the identification of cell response to metabolic stress. Mechanistically, Ca2+ channels and pumps interacting with Tmtc2 will be defined. 2) Aim 1.2 (F99): This project defines how insulin regulates chromatin topology through transcription factor (TF)-mediated alterations in CTCF binding, subsequently regulating the expression of novel aging- associated genes. Chromatin topology will be defined upon insulin treatment followed by the identification of how TFs mediates this process. 3) Aim 2 (K00): This project further identifies how altered DNA methylation remodels the genomic template in aging cells to alter CTCF binding and chromatin loops, shifting the transcriptional impact of insulin and permitting aging-associated metabolic dysfunction. Differential DNA methylation landscape in hepatocytes from young and old mice will be defined, and its functional implications in the alterations of CTCF occupancy and chromatin topology in aging will be identified. This proposal has the potential to provide new genes or DNA elements as targets to achieve liver rejuvenation and the reversal of insulin resistance, which is associated with aging. This project also has the potential to shed light on the fundamental mechanism by which aging shifts cell response to insulin.