Translational dynamics revealed by genome wide profiling of ribosome footprints in Arabidopsis
Significance Plant survival in a highly varied environment requires flexibility in gene regulation. To capture dynamics of mRNA translation at the genome scale, we precisely mapped individual ribosomes to mRNAs of whole seedlings under control and low oxygen conditions. The results demonstrate nearly 100 fold variation in the efficiency of translation of individual mRNAs under both conditions and provide unique insights into posttranscriptional and translational regulation modulated by low energy stress in Arabidopsis thaliana. Translational regulation contributes to plasticity in metabolism and growth that enables plants to survive in a dynamic environment. Here, we used the precise mapping of ribosome footprints (RFs) on mRNAs to investigate translational regulation under control and sublethal hypoxia stress conditions in seedlings of Arabidopsis thaliana. Ribosomes were obtained by differential centrifugation or immunopurification and were digested with RNase I to generate footprint fragments that were deep sequenced. Comparison of RF number and position on genic regions with fragmented total and polysomal mRNA illuminated numerous aspects of posttranscriptional and translational control under both growth conditions. When seedlings were oxygen deprived, the frequency of ribosomes at the start codon was reduced, consistent with a global decline in initiation of translation. Hypoxia up regulated gene transcripts increased in polysome complexes during the stress, but the number of ribosomes per transcript relative to normoxic conditions was not enhanced. On the other hand, many mRNAs with limited change in steady state abundance had significantly fewer ribosomes but with an overall similar distribution under hypoxia, consistent with restriction of initiation rather than elongation of translation. RF profiling also exposed the inhibitory effect of upstream ORFs on the translation of downstream protein coding regions under normoxia, which was further modulated by hypoxia. The data document translation of alternatively spliced mRNAs and expose ribosome association with some noncoding RNAs. Altogether, we present an experimental approach that illuminates prevalent and nuanced regulation of protein synthesis under optimal and energy limiting conditions.
Proceedings of the National Academy of Sciences