PROJECT SUMMARY Characterized by progressive cognitive decline, Alzheimer's disease (AD) is the most common type of dementia, primarily affecting the aging population. Inflammatory activation of microglia is an early pathological feature of Alzheimer’s disease (AD) and contributes to brain damages. However, the underlying mechanisms that mediate microglial inflammatory activation in Aβ-rich milieus are not completely understood. Stimulator of interferon genes (STING) is an innate immune adaptor protein that is abundantly expressed by cells of myeloid origin including microglia. Despite its roles in interferon-based antiviral immunity, recent studies have revealed an important contribution of STING signaling to inflammatory damages in non-communicable disorders. cyclic GMP- AMP (cGAMP) synthase (cGAS) is a cytosolic DNA sensor and signals through STING to promote interferon signaling and production of inflammatory mediators. Cytosolic accumulation of endogenous DNA especially mitochondrial DNA (mtDNA) fragments constitutes an important source of endogenous “sterile” activator of cGAS-STING signaling. In our preliminary study on patients and an AD animal model with AD-like amyloidopathy, we observed increased microglial STING activity, which contributed to early microglial activation in 5xFAD mice. In addition, microglia from patients and 5xFAD mice demonstrated cytosolic mtDNA accumulation; and deficiency of cGAS attenuated microglial STING activation in 5xFAD mice, implicating an association of cytosolic mtDNA sensing by cGAS with STING activation. Finally, our data suggest that microglial mitochondrial stress in Aβ milieus may contribute to mtDNA instability and leakage into the cytosol. We therefore hypothesize that microglial STING signaling triggered via cytosolic mtDNA sensing by cGAS contributes to Aβ- induced microglial activation, culminating in neuroinflammation and neuronal stress in AD. Here, we aim to establish a link between microglial STING activity and microglial activation in 5xFAD mice. Next, we will determine the contribution of microglial STING activation to microglial deregulation-associated brain damages in 5xFAD mice. In addition, we will address the mechanisms that mediate microglial mtDNA release in Aβ-rich milieus and determine the importance of mtDNA leakage for the activation of microglial cGAS-STING signaling in Aβ-rich milieus. Taken together, the results from this study will answer important questions about STING in shaping microglial phenotype, and will suggest a novel mitochondrial mechanism of microglial activation and new avenues for the treatment of AD.