RNA Granules in Cerebellar Neurodegeneration

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Neurodegenerative diseases represent an ever-increasing societal and economic burden with WHO estimates indicating that they will replace cancer as the 2nd leading cause of death by 2040. In neurodegenerative disease research, a wealth of pathways has been uncovered, but their direct and primary relevance to the respective human disease has been difficult to prove and targeting of pathways has remained difficult. The proposed work will characterize stress granules (SGs) in spinocerebellar ataxia type 2 (SCA2), a hereditary neurodegenerative disease affecting cerebellar Purkinje neurons (PNs) and other neurons in the cerebellum, brainstem and cerebrum. The cause of SCA2 is a gain-of-function CAG expansion in the ATXN2 gene resulting in an expanded polyglutamine (polyQ) in ataxin-2. ATXN2 was previously known to have functions in mRNA metabolism based on its interactions with multiple RNA binding proteins (RBPs) including A2BP1/RBFOX1, DDX6, PABP1, TDP-43, FUS. We have now demonstrated that mutant ATXN2 interacts with the SG protein Staufen1, and this interaction has specific effects on SGs. Staufen expression (but not DDX6 or PABPC1) is increased with ATXN2 mutation, and we have also identified specific downstream consequences of this interaction on the abundance of SG mRNAs directly interacting Staufen. The objective of the proposed research is to characterize mechanistically why Staufen expression increases with ATXN2 mutation, to characterize the proteomic and mRNA composition of SCA2 SGs in SCA2 patient fibroblasts and neurons and in SCA2 transgenic mice. SG elements that are identified by this work might be exploited therapeutically for treating SCA2. Three specific aims are proposed: 1) We will evaluate mutant ATXN2 inhibition of protein autophagy as a reason for staufen overexpression, and will characterize SCA2 SG rate of formation, half-life, and superstructure using superresolution microscopy, as well as SCA2 SG transcriptomes using purified SGs. 2) We will conduct FISH to localize SCA2 SG mRNAs in cultured SCA2 neurons and in cerebellar slices of SCA2 mice. 3) We will determine SCA2 mouse motor and neurophysiological phenotypes in SCA2 mice haploinsufficient for Staufen, and will record Purkinje cell firing frequencies simultaneously with imaging for TIA1 positive SGs and calcium abundance. This will determine whether abnormal SG trafficking/localization associates with abnormal PC physiology in SCA2 mice, depending on Staufen expression. The proposed work will clarify the role for SGs in neurodegeneration and will aid in the identification of new avenues towards treatments of SCA2 and other degenerative ataxias