Analysis of Translational Regulation Using Polysome Profiling and Puromycin Incorporation

Abstract

Translation is the process of decoding an mRNA transcript to permit the synthesis of a protein. This process occurs in three steps: initiation, elongation, and termination. Each step of translation is regulated by translation factors. By regulating translation, the quantity and quality of proteins can be controlled. When translation becomes dysfunctional, disease can ensue, making translational regulation an important avenue of research. Polysome profiling and puromycin incorporation are experimental techniques used in concert to analyze the translational state of cells or tissues. Polysome profiling evaluates the state of translation by quantifying mRNAs based on the abundance of associated ribosomes. Puromycin incorporation measures the amount of newly synthesized protein. Together these methodologies can decipher stark and subtle changes in the rate and efficiency of translation, and provide the opportunity to dissect alterations to the translation of specific transcripts.