Azacitidine (5-AzaC)-treatment and mutations in DNA methylase genes affect embryogenic response and expression of the genes that are involved in somatic embryogenesis in Arabidopsis

Daria Grzybkowska,Barbara Wójcikowska,Małgorzata Danuta Gaj,Joanna Morończyk

doi:10.1007/s10725-018-0389-1

Daria Grzybkowska, Barbara Wójcikowska + Show 2 more

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https://doi.org/10.1007/s10725-018-0389-1

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Journal: Plant Growth Regulation	Publication Date: Mar 29, 2018
Citations: 53	License type: open-access

Affiliation: University of Silesia

Abstract

Epigenetic processes including DNA methylation play a pivotal role in regulating the genes that control plant development. In contrast to in planta development, the contribution of DNA methylation to the morphogenic processes that are induced in vitro are much less recognised. Hence, in the present study, we analysed the impact of DNA methylation on somatic embryogenesis (SE) that was induced in Arabidopsis. The results demonstrated a decrease in the global DNA methylation level during SE that contrasted with the up-regulation of MET1 and CMT3 DNA methylases and the down-regulation of DNA demethylases (ROS1, DME and DML2). Hence, the global DNA methylation level appears not to correlate with the transcriptional activity of the genes encoding DNA methylases/demethylases, thereby implying the complexity of the regulatory mechanism that controls the DNA methylation status of the SE-epigenome. Moreover, distinct changes in the expression level of the SE-regulatory genes were indicated in the 5-AzaC-treated and DNA methylase mutant cultures. Accordingly, a significant repression of the LEC2, LEC1 and BBM genes was found in the 5-AzaC-treated culture that was incapable of SE induction. In contrast, the distinct up-regulation of these genes was observed in the drm1drm2 and drm1drm2cmt3 mutant cultures with an improved embryogenic response. The modulated expression of DNA methylase genes and the significantly modified embryogenic response of the met1 and drm mutants imply that both the maintenance and the de novo pathway of DNA methylation are engaged in the regulation of SE in Arabidopsis.

Highlights

Somatic embryogenesis (SE) is a plant-specific developmental process that involves the induction of the embryogenic programme in somatic cells, which results in the formation of somatic embryos that are capable of regenerating complete plants
RT-qPCR analysis was used to analyse the expression of the genes encoding the transcription factors (TFs) of the documented regulatory function in SE induction in Arabidopsis including LEC1 (LEAFY COTYLEDON1), LEC2 (LEAFY COTYLEDON2) and BBM (BBM BABY BOOM)
The results demonstrated that the expression profiles of LEC1, LEC2 and BBM genes were similar in the dd and ddc mutant cultures and that the analysed genes showed a significantly increased expression in the mutant explants that were induced on the E5 medium (Fig. 3)

Summary

Introduction

Somatic embryogenesis (SE) is a plant-specific developmental process that involves the induction of the embryogenic programme in somatic cells, which results in the formation of somatic embryos that are capable of regenerating complete plants. The methylation of plant DNA involves the addition of a methyl group to the carbon-5 of cytosine at the CpG, CpNpG and CpNpN (where N could be any nucleotide except G) sequences in DNA, which results in an increase of the content of 5-methyl cytosine (5 mC) in the genomic DNA (Law. Plant Growth Regulation (2018) 85:243–256 and Jacobsen 2010). Three types of DNA methylases have been found in plants including METHYLTRANSFERASE 1 (MET1), CHROMOMETHYLASE 3 (CMT3) and DOMAINS REARRANGED METHYLTRANSFERASE (DRM1 and DRM2) (Lindroth et al 2001; Cao and Jacobsen 2002a; Zhang et al 2006). In Arabidopsis thaliana, DRMs are required for de novo methylation while MET1 and CMT3 maintain the methylation pattern during DNA replication (Zhang et al 2010)

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