Cell-Free Formation of RNA Granules




Han, Tina Wei

Journal Title

Journal ISSN

Volume Title


Content Notes


Asymmetric RNA localization is a mechanism by which a cell can spatially and temporally regulate the translation of RNAs. This mechanism is essential for many developmental processes such as germ cell formation in Drosophila embryos, as well as establishment of cell polarity and synaptic plasticity in the brain. In many instances, asymmetric RNA localization is achieved through transport and sequestration by RNA granules. RNA granules are large, non-membrane bound ribonucleoparticles that have been observed in various biological contexts. Unfortunately, because RNA granules are highly heterogeneous and weakly associating aggregates, they can be difficult to study biochemically, which constitutes a major impediment for gaining a more detailed understanding of the mechanisms governing RNA granule assembly. Here we describe two in vitro models for studying RNA granule assembly. The first method is based on the precipitation activity of a 3,5-disubstituted isoxazole compound that can be used as a quick and efficient pharmacological tool to probe the function and regulation of RNA granules. The second method utilizes a three-dimensional protein-retaining hydrogel formed from a recombinant protein. Polypeptides of low amino acid complexity were found to be the sequence determinants of isoxazole precipitation and hydrogel retention. Next generation sequencing was used to identify RNAs that partitioned with granule components in both isoxazole and hydrogel models and were found to be enriched in mRNAs known to be constituents of neuronal transport granules for dendritic localization. The overrepresented gene ontology categories for these RNAs included cell adhesion, extracellular matrix, and synaptic proteins. The average length of the 3’UTR of these RNAs was found to be longer than the 3’UTRs of RNAs excluded from the cell-free RNA granule preparations. These two in vitro models for studying RNA granule assembly offer a novel approach to identify candidate targets recruited to RNA granules by specific RNA-binding proteins.

General Notes

Table of Contents


Related URI