Comparison of mitochondrial gene expression and polysome loading in different tobacco tissues

Muhammad Waqar Hameed,Ralph Bock,Ilona Juszczak,Joost Thomas van Dongen

doi:10.1186/s13007-017-0257-4

Abstract

BackgroundTo investigate translational regulation of gene expression in plant mitochondria, a mitochondrial polysome isolation protocol was established for tobacco to investigate polysomal mRNA loading as a proxy for translational activity. Furthermore, we developed an oligonucleotide based microarray platform to determine the level of Nicotiana tabacum and Arabidopsis thaliana mitochondrial mRNA.ResultsMicroarray analysis of free and polysomal mRNAs was used to characterize differences in the levels of free transcripts and ribosome-bound mRNAs in various organs of tobacco plants. We have observed higher mitochondrial transcript levels in young leaves, flowers and floral buds as compared to fully expanded leaves and roots. A similar pattern of abundance was observed for ribosome-bound mitochondrial mRNAs in these tissues. However, the accumulation of the mitochondrial protein COX2 was found to be inversely related to that of its ribosome-bound mRNA.ConclusionsOur results indicate that the association of mitochondrial mRNAs to ribosomes is largely determined by the total transcript level of a gene. However, at least for Cox2, we demonstrated that the level of ribosome-bound mRNA is not reflected by the amount of COX2 protein.

Highlights

To investigate translational regulation of gene expression in plant mitochondria, a mitochondrial polysome isolation protocol was established for tobacco to investigate polysomal mRNA loading as a proxy for translational activity
We showed for the COX2 protein that, higher ribosome-bound mRNA levels were associated with lower protein levels and lower ribosome bound mRNA levels were associated with higher protein levels
It is tempting to use ribosome binding of mRNAs as a proxy for translational activity, polysome association is not necessarily positively correlated with the actual protein abundance. This is due to the impacts of post-translational modifications, regulated proteolysis, protein folding and assembly into complexes on the lifetime of a protein. This consideration is valid for the results presented here, where the ribosome-bound mRNA levels did not correspond to the actual protein abundance in that organs with high mitochondrial ribosome bound cox2 mRNA levels showed low COX2 protein abundance

Summary

Introduction

To investigate translational regulation of gene expression in plant mitochondria, a mitochondrial polysome isolation protocol was established for tobacco to investigate polysomal mRNA loading as a proxy for translational activity. Mitochondria evolved when a free-living α-proteobacterium was engulfed by a single-celled protist [1, 2]. The engulfed cell was not digested but rather domesticated by the host cell to establish an endosymbiotic association. During the course of evolution, the mitochondrial genome was significantly reduced, due to the loss of genes or their transfer to the nuclear genome of the host cell. Regardless of the many gene translocations, mitochondria in all multicellular organisms and most lineages of protists retained their own genome [6,7,8].

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