Initiation of leaf somatic embryogenesis involves high pectin esterification, auxin accumulation and DNA demethylation in Quercus alba

Abstract

Somatic embryogenesis is considered a convenient tool for investigating the regulating mechanisms of embryo formation; it is also a feasible system for in vitro regeneration procedures, with many advantages in woody species. Nevertheless, trees have shown recalcitrance to somatic embryogenesis, and its efficiency remains very low in many cases. Consequently, despite the clear potential of somatic embryogenesis in tree breeding programs, its application is limited since factors responsible for embryogenesis initiation have not yet been completely elucidated. In the present work, we investigated key cellular factors involved in the change of developmental program during leaf somatic embryogenesis initiation of white oak (Quercus alba), aiming to identify early markers of the process. The results revealed that pectin esterification, auxin accumulation and DNA demethylation were induced during embryogenesis initiation and differentially found in embryogenic cells, while they were not present in leaf cells before induction or in non-embryogenic cells after embryogenesis initiation. These three factors constitute early markers of leaf embryogenesis and represent processes that could be interconnected and involved in the regulation of cell reprogramming and embryogenesis initiation. These findings provide new insights into the mechanisms underlying plant cell reprogramming, totipotency and embryogenic competence acquisition, especially in tree species for which information is scarce, thus opening up the possibility of efficient manipulation of somatic embryogenesis induction.

Microspore embryogenesis: targeting the determinant factors of stress-induced cell reprogramming for crop improvement

“…characterized by large nuclei and homogenous euchromatin (Grafi et al, 2011). During microspore embryogenesis, changes in global DNA methylation levels have been observed in various herbaceous and woody species (Corredoira et al, 2017;El-Tantawy et al, 2014;Rodriguez-Sanz et al, 2014b;Solis et al, 2012;De-la-Peña et al, 2015;. Using quantitative biochemical assays and immunolocalization of 5deoxy-methyl-cytosine (5mdC), these studies demonstrated that microspore reprogramming and embryogenesis initiation involve a global DNA hypomethylation.…”

Inhibition of Histone H3K9 Methylation by BIX-01294 Promotes Stress-Induced Microspore Totipotency and Enhances Embryogenesis Initiation

“…Previous studies have shown modifications in global DNA methylation that accompanied the change in developmental program of the microspore toward embryogenesis, in very different plant species, like Brassica napus , Hordeum vulgare , Quercus suber and Q. alba (Solis et al, 2012; Testillano et al, 2013; El-Tantawy et al, 2014; Rodriguez-Sanz et al, 2014a; Corredoira et al, 2017), indicating an epigenetic reprogramming after microspore induction to a totipotent state and embryogenesis initiation. More recently, it has been demonstrated that treatment with the DNA demethylating agent azacytidine reduces global DNA methylation of isolated microspore cultures and concomitantly increases embryogenesis initiation rates (Solis et al, 2015).…”

Novel dye for detection of callus embryo by confocal laser scanning fluorescence microscopy

“…Moreover, selection of suitable explants is a significant step for successful obtaining of somatic embryos and high frequency of plant regeneration achievement . Mainly, for this purpose scientists utilize histology methods that are time consuming and relatively expensive . Therefore, it is important to evolve the fluorescent dyes for fast diagnostics of somatic embryos obtained in in vitro conditions.…”

“…Consequently, there is a necessity for simple, fast and relatively inexpensive procedure to test the suitability of high regeneration method in early stage of callus cultivation. Mainly, histology methods that are time consuming and relatively expensive are utilized for this purpose . In previous studies, potential‐sensitive fluorescent dyes were used to visualize plant mitochondria and nuclei, cell‐to‐cell transport trough plasmodesmata and pigment metabolic pathways .…”