Measuring Organ-Specific Translation Elongation Rate in Mice

Abstract

Modern methods of genome editing enable the rapid generation of mouse models to study the regulation of protein synthesis. At the same time, few options are available to study translation in rodents as the animal's complexity severely limits the repertoire of experimental tools. Here we describe a method to monitor translation in mice and other small animals. The technique is based on a ribosome profiling and specifically tailored toward measuring translation elongation. However, it can be easily applied for short upstream reading frames discovery. The advantage of this method is the ability to study translation in fully developed animals without extracting and subculturing cells, therefore, maintaining unperturbed physiological conditions.

Keywords: Elongation; Harringtonine; Injection; Mouse; Organ; Ribosome profiling; Translation.

Fig. 1

Schematic of mouse injection set up.

Fig. 2

Polysome profiles obtained from mouse livers with no injections (left), injected with harringtonine (center), co-injected with harringtonine and cycloheximide (right).

Fig. 3

Representative time-dependent ribosome occupancy plots for the kidney and the liver. 3 time point were tested: 15, 30, and 45 seconds between harringtonine and cycloheximide injections.

Fig. 4

Ribosome occupancy over a representative gene (Acadm). The more time points collected - the more accurate the translation elongation rate estimation.