Abstract
Background: Somatic embryogenesis (SE) is one of the most promising processes for large-scale dissemination of elite varieties. However, for many plant species, optimizing SE protocols still relies on a trial and error approach. We report the first global scale transcriptome profiling performed at all developmental stages of SE in coffee to unravel the mechanisms that regulate cell fate and totipotency. Results: RNA-seq of 48 samples (12 developmental stages × 4 biological replicates) generated 90 million high quality reads per sample, approximately 74% of which were uniquely mapped to the Arabica genome. First, the statistical analysis of transcript data clearly grouped SE developmental stages into seven important phases (Leaf, Dedifferentiation, Primary callus, Embryogenic callus, Embryogenic cell clusters, Redifferentiation and Embryo) enabling the identification of six key developmental phase switches, which are strategic for the overall biological efficiency of embryo regeneration. Differential gene expression and functional analysis showed that genes encoding transcription factors, stress-related genes, metabolism-related genes and hormone signaling-related genes were significantly enriched. Second, the standard environmental drivers used to control SE, i.e. light, growth regulators and cell density, were clearly perceived at the molecular level at different developmental stages. Third, expression profiles of auxin-related genes, transcription factor-related genes and secondary metabolism-related genes were analyzed during SE. Gene co-expression networks were also inferred. Auxin-related genes were upregulated during dedifferentiation and redifferentiation while transcription factor-related genes were switched on from the embryogenic callus and onward. Secondary metabolism-related genes were switched off during dedifferentiation and switched back on at the onset of redifferentiation. Secondary metabolites and endogenous IAA content were tightly linked with their respective gene expression. Lastly, comparing Arabica embryogenic and non-embryogenic cell transcriptomes enabled the identification of biological processes involved in the acquisition of embryogenic capacity. Conclusions: The present analysis showed that transcript fingerprints are discriminating signatures of cell fate and are under the direct influence of environmental drivers. A total of 23 molecular candidates were successfully identified overall the 12 developmental stages and can be tested in many plant species to optimize SE protocols in a rational way.