Gene regulation in seeds: insights into translational dynamics

Seeds are unique structures in the plant life cycle. The variation in the timing of seed maturation, dispersion, and the establishment of seed dormancy and longevity, increases the chances of plant survival and enlarge the distance that plants could disperse in the natural habitat. Seeds contain large amounts of mRNAs and the role of these mRNAs in seed dormancy and germination has been investigated in this thesis regarding their translational dynamics using Arabidopsis thaliana seeds as a model system. This study is facilitated by the translatome profiling technique, in which ribosomes are fractionated based on their engagement in translation followed by the high throughput profiling of the mRNAs associated to these ribosomes.Chapter 1 gives an overview of physiological seed traits (dormancy, germination, longevity) related to regulation of gene expression underlining the importance of translational control as well as a comprehensive description of the mechanisms involved in the regulation of mRNA translation. In Chapter 2, the translational dynamics from a quiescent dry seed to a seedling are described. The levels of polysome-associated mRNAs (translatome) are compared with the levels of the mRNAs in the total mRNA pool (transcriptome) in consecutive stages during seed germination. Two stages with extensive changes in translational efficiency were identified and named the Hydration Shift and Germination Shift stages, based on the time windows at which they were identified and that correspond to seed hydration and germination sensu stricto, respectively. Chapter 3 investigates seed stored mRNAs and mRNA binding proteins for their association with ribosomes (monosome/polysome). We show that mRNAs co-purify with monosomes and approximately 30% of these monosome associated mRNAs are translationally up regulated upon seed imbibition while seed mRNAs that are associated to polysomes are mostly leftovers from translation during late seed maturation. Fully matured seeds can only germinate after a period of dry storage, called after-ripening. Whether gene transcription or translation is the driving force for the changes in germinability during dry storage is explored in Chapter 4. Genes that are differentially transcribed and translated during the imbibition of NILDOG1 dormant and non-dormant seeds are investigated. This revealed that dormancy during imbibition is mainly transcriptionally regulated and related to tryptophan-dependent auxin and indole glucosinolate biosynthesis pathways. In Chapter 5 a novel ATP Binding Cassette gene (ABCI20) is characterized, the knockout of this gene dramatically affects seed dormancy, longevity and ABA sensitivity. With the knowledge gained in Chapter 2-4, transcriptome, translatome and ribosome proteome analysis on abci20 seeds are performed which identify molecular processes that contribute to the distinct dormancy and longevity phenotype of abci20. abci20 is the first identified mutant that shows a negative correlation between seed dormancy and longevity and therefore, provides a most useful tool for investigating the trade-off between these two important seed traits. Finally, in Chapter 6, the findings on translational dynamics in the various aspects of seed physiology are integrated. Furthermore the potential for translational regulation for future research in seed biology and plant breeding are discussed.