Project Summary Alzheimer's disease (AD) is the most common cause of elderly dementia and currently there are no effective treatments. Selective neuronal vulnerability, appearance and accumulation of amyloid-β (Aβ) plaques, altered circuitry function, synaptic loss and degeneration are hallmarks of AD. Microglia and astrocytes, two major subtypes of glial cells, contribute to these neuropathological changes. However, the precise mechanisms controlling these processes remain elusive. One of the reasons may be that we still lack the information about the cell type-specific alterations and how they contribute to the onset and progression of AD. Here, we postulate that by revealing detailed changes in functional networks of genes in microglia and astrocytes, and how those processes intersect, we may be able to determine their contribution to neuropathological changes in AD. We will utilize Translational Ribosomal Affinity Purification combined with novel bioinformatics tools to construct functional network models of glial-specific functions and integrate these with AD quantitative genetic data. Using this framework, we will detect glial-specific genes most likely associated with AD pathology in an unbiased data-driven way and further investigate the functional network modules between astrocytes and microglia. Relevant candidate genes from this analysis will be examined in vitro for their role in AD progression, and emphasis will be given to cytokine-cytokine receptor pathways, especially interleukin 1 and colony stimulating factor 1 hubs. We will utilize primary cell cultures and co-cultures with astrocytes and microglia isolated from healthy and 5xFAD mice. Furthermore, we will investigate only genes that are relevant to the human pathology. To achieve this, we will test the appearance of AD hallmarks and/or the rescue phenotype in the microglia and astrocytes derived from the induced pluripotent stem cells (iPSC) isolated from AD patients and healthy controls. Finally, the most important gene candidates will be tested in vivo by gene expression manipulation in the appropriate mice lines. We will measure: i) Aβ accumulation, uptake, and degradation by microglia and astrocytes, ii) neuronal degeneration, iii) synaptic function, including synaptic markers and functional circuitry, iv) plaque formation, and v) cognitive function, assessed with a battery of behavioral tests. The proposed study will reveal the molecular profile of each cell type and examine their interaction. Generated datasets and bioinformatics tools will be shared with the public via web-based interface. These data will offer new insights into the appearance of the AD hallmarks and elucidate the basic mechanisms of the disease. Studying these changes, especially on the whole genome level, will engender tremendous insights into alterations in the individual genes, as well as pathways, and may offer new approaches to studying the cause of AD, as well as reveal novel therapy targets.