ASH1 mRNP-core factors form stable complexes in absence of cargo RNA at physiological conditions

Abstract

Asymmetric ASH1 mRNA transport during mitosis of budding yeast constitutes one of the best-studied examples of mRNA localization. Recently, 2 studies used in vitro motility assays to prove that motile ASH1 mRNA-transport complexes can be reconstituted entirely from recombinant factors. Both studies, however, differed in their conclusions on whether cargo RNA itself is required for particle assembly and thus activation of directional transport. Here we provide direct evidence that stable complexes do assemble in absence of RNA at physiologic conditions and even at ionic strengths above cellular levels. These results directly confirm the previous notion that the ASH1 transport machinery is not activated by the cargo RNA itself, but rather through protein-protein interactions.

Keywords: ASH1 mRNA; Myo4p; RNA localization; She2p; She3p; budding yeast; in vitro reconstitution; mRNP; macromolecular assembly; myosin.