Abstract

mRNA secondary structure influences translation. Proteins that modulate the mRNA secondary structure around the translation initiation region may regulate translation in plastids. To test this hypothesis, we exposed Arabidopsis thaliana to high light, which induces translation of psbA mRNA encoding the D1 subunit of photosystem II. We assayed translation by ribosome profiling and applied two complementary methods to analyze in vivo RNA secondary structure: DMS-MaPseq and SHAPE-seq. We detected increased accessibility of the translation initiation region of psbA after high light treatment, likely contributing to the observed increase in translation by facilitating translation initiation. Furthermore, we identified the footprint of a putative regulatory protein in the 5′ UTR of psbA at a position where occlusion of the nucleotide sequence would cause the structure of the translation initiation region to open up, thereby facilitating ribosome access. Moreover, we show that other plastid genes with weak Shine-Dalgarno sequences (SD) are likely to exhibit psbA-like regulation, while those with strong SDs do not. This supports the idea that changes in mRNA secondary structure might represent a general mechanism for translational regulation of psbA and other plastid genes.

Highlights

  • The secondary structure of mRNA is important for many translation related processes in bacteria and bacteria-derived eukaryotic organelles

  • The intrinsic mRNA features that determine the efficiency of start codon recognition in plastids of higher plants, and the efficiency of translation initiation, are well characterized: (a) Shine-Dalgarno sequences hybridize to the anti-Shine-Dalgarno sequence at the tail of the 16S rRNA and thereby position the start codon so that it can bind to the initiator tRNA; (b) local minima of mRNA

  • We tested whether changes in mRNA secondary structure would influence translation in chloroplasts using a well-known example for translation regulation as a starting point: the high light induced upregulation of psbA translation, the mRNA coding for the D1 subunit of photosystem II [30,31]

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Summary

Introduction

The secondary structure of mRNA is important for many translation related processes in bacteria and bacteria-derived eukaryotic organelles. This includes the efficiency of translation initiation [1,2,3,4,5], the recognition of start codons [6,7,8], and ribosome pausing [9,10,11]. It has been proposed that changes in mRNA secondary structure regulate translation in plastids, i.e., plant organelles derived from cyanobacteria [17,18,19,20]. The intrinsic mRNA features that determine the efficiency of start codon recognition in plastids of higher plants, and the efficiency of translation initiation, are well characterized: (a) Shine-Dalgarno sequences hybridize to the anti-Shine-Dalgarno sequence at the tail of the 16S rRNA and thereby position the start codon so that it can bind to the initiator tRNA; (b) local minima of mRNA secondary structure around the start codon make it accessible for the ribosome, whereas other AUGs are masked by folded RNA [6,8,25,26,27,28]

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