Project Summary/Abstract Alzheimer’s disease (AD) is the most common neurodegenerative disease affecting over 3 million Americans yearly, with extracellular accumulation of β-amyloid and intracellular tau aggregates, and aberrant glial activation pathologies. Glial dysfunction can trigger excitotoxicity and neuroinflammation, which is invariably involved in AD pathogenesis. However, how astrocyte and microglia become reactive in the pathogenesis of AD is not clear. Previously, I have identified a novel ER component, membralin (TMEM259), as an important disease modifier in the pathogenesis of AD and Amytrophic Lateral Sclerosis (ALS). First, I find that membralin can modulate the integrity and activity of the γ-secretase complex. I found that knocking down of membralin expression in a mouse model of AD (TgCRND8) can exacerbated Aβ pathology and memory impairment. More recently, I have identified a non-cell-autonomous glutamate clearance mechanism in astrocytes mediated by membralin through regulation of the glutamate transporter, EAAT2. Elevation of membralin through AAV virus injection can significantly increase EAAT2 levels and extend the lifespan of the SOD1G93A ALS mice. Interestingly, astrocyte-deletion of membralin can lead a severe neuroinflammatory pathologies, as shown by remarkable elevation of gliosis markers: GFAP (astrocytes), IBA1 and CD68 (microglia). Transcriptomic analysis of astrocyte conditional knockout animals confirms the upregulation of genes associated to gliosis, neuroinflammation and abnormal immune response. I found reduced membralin levels in brain samples from both AD and ALS patients. Excitotoxicity, EAAT2 dysfunction and gliosis are common pathological features in AD and ALS. Moreover, a recent genome-wide association (GWAS) study has shown that the membralin gene locus (also known as C19ORF6 in human) is located within 500 bp of a single nucleotide polymorphism (SNP, rs117481827) tightly associated with late-onset AD, and splicing of membralin transcripts has been reported to be significantly altered in AD. Thus, I hypothesize that upregulation of astrocytic membralin pathways can attenuate glutamate excitotoxicity and modulate microglial-dependent pathogenic effects in AD. In the K99 phase of this study, I will characterize molecular mechanisms underlying membralin-associated astrocyte function and dissect the induction mechanisms of reactive astrocytes (Aim 1). I will determine whether modulation of astrocytic membralin neuroinflammatory pathways can alter pathogenic effects in an AD mouse model (Aim 2). In the R00 phase of this study, I will investigate modulation of a membralin-dependent astrocytic TREM2-dependent DAM switch in microglia (Aim 3). The proposed study characterizing the gliosis induction mechanisms in AD, will provide insight into neuroprotective membralin-associated astrocyte pathways that can limit glutamatergic excitotoxicity and neuroinflammation through cell-autonomous and non-cell autonomous ...