Abstract
Molecular mechanisms controlling plant totipotency are largely unknown and studies on somatic embryogenesis (SE), the process through which already differentiated cells reverse their developmental program and become embryogenic, provide a unique means for deciphering molecular mechanisms controlling developmental plasticity of somatic cells. Among various factors essential for embryogenic transition of somatic cells transcription factors (TFs), crucial regulators of genetic programs, are believed to play a central role. Herein, we used quantitative real-time polymerase chain reaction (qRT-PCR) to identify TF genes affected during SE induced by in vitro culture in Arabidopsis thaliana. Expression profiles of 1,880 TFs were evaluated in the highly embryogenic Col-0 accession and the non-embryogenic tanmei/emb2757 mutant. Our study revealed 729 TFs whose expression changes during the 10-days incubation period of SE; 141 TFs displayed distinct differences in expression patterns in embryogenic versus non-embryogenic cultures. The embryo-induction stage of SE occurring during the first 5 days of culture was associated with a robust and dramatic change of the TF transcriptome characterized by the drastic up-regulation of the expression of a great majority (over 80%) of the TFs active during embryogenic culture. In contrast to SE induction, the advanced stage of embryo formation showed attenuation and stabilization of transcript levels of many TFs. In total, 519 of the SE-modulated TFs were functionally annotated and transcripts related with plant development, phytohormones and stress responses were found to be most abundant. The involvement of selected TFs in SE was verified using T-DNA insertion lines and a significantly reduced embryogenic response was found for the majority of them. This study provides comprehensive data focused on the expression of TF genes during SE and suggests directions for further research on functional genomics of SE.
Highlights
Most plant cells, in contrast to animal cells, express an amazing developmental plasticity allowing their reprogramming and manifestation of totipotency [1]
Immature zygotic embryos (IZEs) at the late cotyledonary stage of development were carefully excised from siliques 10–12 days after pollination and cultured on solid medium containing the synthetic auxin analog 2,4-dichlorophenoxyacetic acid (2,4-D, 5 mM)
The experiment was designed to monitor the expression of 1,880 transcription factors (TFs) genes at three distinctive stages of immature zygotic embryos (IZEs)-derived embryogenic culture: (i) freshly isolated explants (0 d), (ii) explants subjected to somatic embryogenesis (SE) induction for 5 days (5 d), and (iii) explants at an advanced stage of embryogenesis related to somatic embryo formation (10 d)
Summary
In contrast to animal cells, express an amazing developmental plasticity allowing their reprogramming and manifestation of totipotency [1]. Our current understanding of the genetic mechanisms controlling plant totipotency are largely based on studies on somatic embryogenesis (SE), the process through which already differentiated cells reverse their developmental program during in vitro culture and become embryogenic giving rise to the formation of somatic embryos which develop further into entire plants. The control of plant embryogenesis, similar to other developmental processes, occurs through a complex set of intrinsic signals that are involved in providing information to the dividing and differentiating cells. In adult human somatic cells a specific combination of TFs was found to re-programme differentiated cells into pluripotent embryonic stem cells [8,9]. A combination of only four over-expressed TFs was sufficient to induce the formation of pluripotent stem cells from e.g. adult human fibroblasts [10,11]
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