Abstract/Summary The hypotheses to test are that Alu-derived RNAs via activation of double-stranded RNA (dsRNA) sensors stimulate production of both anti-inflammatory type 1 IFNs and IL-33 in the absence of stimulation of the pro- inflammatory inflammasome as well as pathways producing TNF-a. Further, our results argue that tandem repeats of Alu elements will have greater activity than single Alu RNAs to activate these responses and will serve as therapeutic candidates. Our long-term goal will be to identify potent Alu dsRNAs that activate endogenous dsRNA sensors and determine if these Alu dsRNAs block and prevent disease progression in rodent models of multiple sclerosis, such as experimental autoimmune encephalomyelitis, and promote neuronal re-myelination in rodent models of neuronal damage. Alu elements are unique to primates and about 1,000,000 Alu elements exist in the human genome. An Alu element is about 300 bp in length, thus Alu elements account for about 10% of the human genome, are known to be transcribed into RNAs, and since they are composed of highly conserved inverted repeats have the potential to form double-stranded structures if transcribed into RNA and stimulate dsRNA sensors. We find that during the relapsing-remitting phase of relapsing-remitting multiple sclerosis (RRMS) there is an elevated type 1 IFN response in peripheral leukocytes that is mediated by dsRNA. This dsRNA fraction is largely composed of Alu elements and the proportion of highly expressed dsRNA Alu elements is markedly increased in RRMS patients compared to healthy control (HC). Finally, in vitro transcribed single element Alu RNA will trigger a type 1 IFN response at concentrations ~100-fold less than the synthetic dsRNA mimic, poly I/C. We also show that genomic positions of highly transcribed Alu elements are close in proximity (< 5kb) to genomic positions of leukocyte transcriptional enhancers. Thus, are working hypothesis is that these Alu containing RNAs that activate type 1 interferon responses are derived from leukocyte enhancers with embedded Alu elements. Besides elevated type 1 IFN responses during the relapsing-remitting phase of RRMS, IL-33 levels are also elevated and IL33 is known to require interferon regulatory factors (IRF) via an interferon-stimulated response element (ISRE) for its transcription. Further, like type 1 interferons and interferon-response genes, poly I/C also stimulates expression of IL33. These data support the notion that these dsRNA Alu elements will also induce IL33 expression, which is relevant to our proposal since IFN-b is a proven therapy for RRMS. Both poly I/C and IL-33 induce expression of myelin basic protein (MBP) and myelin oligodendrocyte glycoprotein (MOG) and stimulate re-myelination in a demyelinating rodent model of gliotoxic in...