The Tomato Translational Landscape Revealed by Transcriptome Assembly and Ribosome Profiling.

Recent applications of translational control in Arabidopsis () highlight the potential power of manipulating mRNA translation for crop improvement. However, to what extent translational regulation is conserved between Arabidopsis and other species is largely unknown, and the translatome of most crops remains poorly studied. Here, we combined de novo transcriptome assembly and ribosome profiling to study global mRNA translation in tomato () roots. Exploiting features corresponding to active translation, we discovered widespread unannotated translation events, including 1,329 upstream open reading frames (uORFs) within the 5' untranslated regions of annotated coding genes and 354 small ORFs (sORFs) among unannotated transcripts. uORFs may repress translation of their downstream main ORFs, whereas sORFs may encode signaling peptides. Besides evolutionarily conserved sORFs, we uncovered 96 Solanaceae-specific sORFs, revealing the importance of studying translatomes directly in crops. Proteomic analysis confirmed that some of the unannotated ORFs generate stable proteins in planta. In addition to defining the translatome, our results reveal the global regulation by uORFs and microRNAs. Despite diverging over 100 million years ago, many translational features are well conserved between Arabidopsis and tomato. Thus, our approach provides a high-throughput method to discover unannotated ORFs, elucidates evolutionarily conserved and unique translational features, and identifies regulatory mechanisms hidden in a crop genome.