Project Summary Synthetic biology offers transformative technologies for constructing or rewiring gene networks to create novel biological functions. However, the rational engineering of complex biological traits remains very challenging. To address this, we propose to integrate synthetic biology and computational modeling to engineer cellular aging, an intricate, multifaceted biological process crucially related to various diseases, including cancers and neurodegenerative disorders. In this proposal, we choose to focus on replicative aging of budding yeast Saccharomyces cerevisiae, a genetically tractable model for aging of mitotic cells in mammals, such as stem cell. We previously found that genetically-identical yeast cells age through two distinct routes with different molecular and cellular changes. About half of the cells age with chromatin silencing loss, leading to nucleolar decline, whereas the other half age with heme depletion, resulting in mitochondrial deterioration. We further revealed that the fate decision and commitment to the two aging paths are governed by a core gene circuit comprised of the lysine deacetylase Sir2 and heme-activated protein (HAP) transcriptional complex. Sir2 and HAP inhibit each other, resembling a toggle switch to drive the divergence in aging. Very recently, we used computational modeling to guide our network engineering and rewired the Sir2-HAP circuit to create a synthetic oscillator that dramatically slows aging by avoiding the cell’s commitment to either of the two natural aging paths. Building on these results, we hypothesize that engineered oscillations in pro-longevity factors (e.g. Sir2 and HAP) can be a general strategy for enabling dynamic homeostasis to slow cell deterioration and promote longevity. To test this, we will systematically tune the amplitude and period of the Sir2-HAP oscillator to optimize its effect on longevity (Aim 1). We will then apply the same design principles to engineer oscillations in the proteasome (Aim 2) and energy sensing systems (Aim 3), both of which are considered as pro-longevity factors, but their constitutive overexpression/over-activation negatively impact cell physiology and lifespan. We will systematically evaluate the effects of oscillations on damage accumulation, aging phenotypes/hallmarks, as well as the lifespan, and will evaluate the mechanisms underlying their effects. We will construct computational modeling to guide the design of synthetic gene networks and to quantitatively analyze the dynamics and effects of engineered oscillations to advance our mechanistic understanding of aging processes. If successful, the proposed research will not only further our basic understanding of aging in natural systems but also establish a general strategy for promoting longevity applicable to diverse aging processes and organisms.