Project Summary/Abstract This R35 proposal builds on experiments supported by a Javits R37 and a U01 CREATE Bio award and the longstanding success of the principal investigator’s group leading from mechanistic discoveries in spinocerebellar ataxias and translation to novel treatments, one of which is now in a phase 1 human trial (BIIB105). The proposal’s unifying theme is a focus on RNA-binding proteins (RBPs) in the ATXN2-complex of proteins that direct several aspects of RNA metabolism and that are prone to phase-separation and aggregation. Building on our recent discoveries in polyglutamine-mediated neurodegeneration and neuronal staufen-1 (STAU1) overabundance, we will apply multi-dimensional approaches to define novel pathogenic mechanisms in their intersections with autophagy, the unfolded protein response (UPR), and with RNA metabolism and transport. We postulate that these responses, while compensatory in the short term, become maladaptive with sustained stress and that an RBP network response leads to progressive deterioration of autophagic flux and amplification of apoptotic signaling. We have established a broad suite of innovative tools and unique models in an exceptional research environment with established local and national collaborators with whom we have shared resources over many years. Our approach has been to characterize cerebellar degeneration at multiple time points using genome-wide transcriptomic, proteomic, morphologic, physiologic and behavioral techniques. These foundations will allow us to rapidly progress from cellular to animal models modifying dosage of specific genes in vitro and in vivo. We will now extend this approach from Purkinje cells to spinal motor neurons enabling us to address fundamental issues of broad relevance to inherited and sporadic neurodegenerative diseases. These topical issues include: the role of cytoplasmic RBPs, especially ATXN2 and STAU1, in response to nutrient, chemical and mutant protein stress; their regulation and interplay with each other, and key proteins determining autophagic flux and the UPR; their overall effect on neuronal death; and finally, their promise as therapeutic targets using small molecules and RNA-directed therapies. R35 funding will allow us to identify novel functions of proteins in the ATXN2-complex in neurodegeneration and define shared features across different neurodegenerative diseases with particular relevance not only to inherited, but also to sporadic forms of ataxia and motor neuron disease. The broad scope of our proposed studies will enable participation of scientists with diverse backgrounds in a laboratory and departmental culture of inclusivity and diversity. With our established commitment to reproducibility of animal models and gene targeting approaches, the novel mechanistic insights hold promise for translation into developing novel treatments for neurodegenerative disorders.