Fluorescent polysome profiling reveals stress-mediated regulation of HSPA14-ribosome interactions

Abstract

Ribosome-associated factors play important roles in regulation of translation in response to various physiological and environmental signals. Here we present fluorescent polysome profiling, a new method that provides simultaneous detection of UV and fluorescence directly from polysome gradients. We demonstrate the capabilities of the method by following the polysome incorporation of different fluorescently tagged ribosomal proteins in human cells. Furthermore, we used fluorescent polysome profiling to examine chaperone-ribosome interactions, and characterized their changes in response to proteotoxic stress conditions. We revealed dynamic regulation of HSPA14-polysome association in response to heat shock, with a marked heat shock-mediated increase in the polysome-association of HSPA14. Our data suggest a model whereby HSPA14 dimerization is increased upon heat shock. We therefore established fluorescent polysome profiling as a streamlined method, that can enhance and facilitate the broad exploration of ribosome-associated factors and their regulation.