Experimental approaches to studying translation in plant semi-autonomous organelles

Abstract

Plant mitochondria and chloroplasts are semi-autonomous organelles originated from free-living bacteria and retaining respective reduced genomes during evolution. As a consequence, relatively few of the mitochondrial and chloroplast proteins are encoded in the organellar genomes and synthesized by the organellar ribosomes. Since the both organellar genomes encode mainly components of the energy transduction systems, oxidative phosphorylation in mitochondria and photosynthetic apparatus in chloroplasts, understanding the organellar translation is critical to a thorough comprehension of the key aspects of mitochondrial and chloroplast activity affecting plant growth and development. Recent studies have clearly shown that translation is a key regulatory node in the expression of plant organellar genes, underscoring the need for an adequate methodology to study this unique stage of gene expression. The organellar translatome can be analysed by studying newly synthesized proteins or the mRNA pool recruited to the organellar ribosomes. In this review, we present in some detail the experimental approaches used to date for studying translation in the plant bioenergetic organelles. Their benefits and limitations, as well as the critical steps are discussed. Additionally, we briefly mention several recently developed strategies to study organellar translation that have not yet been applied to plants.

Keywords: Arabidopsis; chloroplast; mitochondria; plant; polysome profiling; protein synthesis; radioisotope labeling; ribosome; ribosome profiling; translation assay.