Ribosome Profiling

Abstract

Robust mechanisms exist that serve to dynamically regulate the translation of mRNA into proteins across heterogeneous tissues. These processes ensure timely generation of proteins in quantities that scale with the demands of specific cell types. Importantly, this translational regulation occurs with spatiotemporal precision and is capable of recalibration as conditions change. Aberrant regulation of translation contributes to and exacerbates a wide range of diseases. Although dynamic control of translation is an essential and fundamental process shared by organisms, specific tissues and cell types can be differentially impacted by circumstances that challenge and impair basal translation, highlighting the heterogeneous nature of translational regulation. To understand how translation is differentially regulated during changing environments and across specific cells and tissues, methods capable of profiling translation in specific tissues and cells are crucial. Here, we describe a method for profiling genome-wide translation in specific tissues or cell types in Drosophila melanogaster, in which we combine ribosome affinity purification with ribosome profiling to enable a simplified protocol for robust analysis of translation in specific tissues.