RNA-binding proteins (RBPs) function in higher-order assemblages such as RNA granules to regulate RNA localization and translation. The Fragile X homolog FXR2P is an RBP essential for formation of neuronal Fragile X granules that associate with axonal mRNA and ribosomes in the intact brain. However, the FXR2P domains important for assemblage formation in a cellular system are unknown. Here we used an EGFP insertional mutagenesis approach to probe for FXR2P intrinsic features that influence its structural states. We tested 18 different in-frame FXR2PEGFP fusions in neurons and found that the majority did not impact assemblage formation. However, EGFP insertion within a 23 amino acid region of the low complexity (LC) domain induced FXR2PEGFP assembly into two distinct fibril states that were observed in isolation or in highly-ordered bundles. FXR2PEGFP fibrils exhibited different developmental timelines, ultrastructures and ribosome associations. Formation of both fibril types was dependent on an intact RNA-binding domain. These results suggest that restricted regions of the LC domain, together with the RNA-binding domain, may be important for FXR2P structural state organization in neurons.
CITATION STYLE
Stackpole, E. E., Akins, M. R., Ivshina, M., Murthy, A. C., Fawzi, N. L., & Fallon, J. R. (2019). EGFP insertional mutagenesis reveals multiple FXR2P fibrillar states with differing ribosome association in neurons. Biology Open, 8(8). https://doi.org/10.1242/bio.046383
Mendeley helps you to discover research relevant for your work.