Structured and disordered regions of Ataxin-2 contribute differently to the specificity and efficiency of mRNP granule formation

Ataxin-2 (ATXN2) is a gene implicated in spinocerebellar ataxia type II (SCA2), amyotrophic lateral sclerosis (ALS) and Parkinsonism. The encoded protein is a therapeutic target for ALS and related conditions. ATXN2 (or Atx2 in insects) functions in translational regulation, mRNA stability, and in the assembly of mRNP-granules, a process mediated by intrinsically disordered regions (IDRs). Previous work has shown that the LSm (Like-Sm) domain of Atx2, which mediates translational activation of some target mRNAs, antagonizes mRNP-granule assembly. Here we advance these findings through a series of experiments on Drosophila and human Ataxin-2 proteins. Results of Targets of RNA-Binding Proteins Identified by Editing (TRIBE) experiments indicate that a polyA-binding protein (PABP) interacting, PAM2 motif of Ataxin-2 may be a major determinant of the mRNA content of Ataxin-2 mRNP granules. Co-localization and co-immunoprecipitation analyses show that structured interactions between Ataxin-2 and PABP additionally help determine protein components of Ataxin-2-associated mRNP granules and contribute to Ataxin-2’s association with stress granules. Finally, in vivo experiments in Drosophila indicate that while the Atx2-LSm domain protects against neurodegeneration, structured PAM2- and unstructured IDR- interactions both promote degeneration. Taken together the data: (a) lead to a proposal for how Ataxin-2 interactions are remodelled during different stages of translational control; (b) show how structured and non-structured interactions of Ataxin-2 contribute differently to the specificity and efficiency of RNP granule condensation; and (c) demonstrate that the Ataxin-2 protein contains multiple activities that may respectively prevent or promote neurodegeneration. Elav-Gal4 was used to drive Atx2-ADARcd fusion protein in nervous system and the expression was restricted to adult flies with the use of the temperature sensitive Gal4 inhibitor, GAL80ts. Fly brains were dissected, and mRNA was sequenced using Illumina HiSeq 2500. RNA edits sites were identified using TRIBE pipeline (https://github.com/rosbashlab/TRIBE).