Project Summary When sick with an infectious, autoimmune, or neoplastic disease, humans report sleepiness and excess or unrefreshing sleep. The features of human sleep during sickness are shared with those in Drosophila. Sickness behavior in fruit flies as in humans and other species is induced by stressors such as bacterial infection or aseptic injury, which reduce activity and increase sleep. While the causal mechanisms underlying altered sleep during sickness are unknown, immune system and lipid dysregulation are implicated. We will exploit the Drosophila genetic model to directly test specific interactions between the innate immune system and lipid homeostasis to determine how these interactions contribute to sleep need during sickness. We recently identified a novel brain-expressed antimicrobial peptide (AMP) nemuri (nur), which links the immune response with sleep. nur is strongly induced by infection and sleep deprivation and loss of it reduces sleep under these conditions. Our preliminary data indicate that induction of nur is mediated by Nuclear Factor κB transcription factors (NFκB). Overexpression of nur in the brain also promotes sleep and is associated with dysregulation of lipid metabolism in a manner that parallels that of patients with myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). Preliminary data also indicate lipid depletion by sustained sleep deprivation. Based on these findings, we hypothesize that prolonged expression of nemuri, either genetically or as induced by chronic sleep deprivation or infection, causes lipid bilayer stress (LBS). Under normal circumstances, the unfolded protein response, mediated by IRE1/XBP1 signaling, resolves LBS (UPRLBS) and terminates the acute stress-induced sleep response. However, prolonged activation of nur disrupts NFκB activity, which is central to the innate immune response. The suppression of NFκB signaling subsequently prevents an appropriate response to LBS by reducing the adaptive lipid homeostatic UPRLBS, thereby increasing sleep need. We will use behavioral genetic, lipidomic, biochemical, and pharmacological approaches to test key aspects of this hypothesis. Given that immune, lipid metabolic, and UPR pathways are highly conserved between flies and mammals, results of our proposed studies will provide a framework for understanding a mechanism for sleep during sickness, with implications for unrefreshing sleep and fatigue associated with numerous illnesses in humans.