Moderate stress responses and specific changes in polyamine metabolism characterize Scots pine somatic embryogenesis

Abstract

Somatic embryogenesis (SE) is one of the methods with the highest potential for the vegetative propagation of commercially important coniferous species. However, many conifers, including Scots pine (Pinus sylvestris L.), are recalcitrant to SE and a better understanding of the mechanisms behind the SE process is needed. In Scots pine SE cultures, embryo production is commonly induced by the removal of auxin, addition of abscisic acid (ABA) and the desiccation of cell masses by polyethylene glycol (PEG). In the present study, we focus on the possible link between the induction of somatic embryo formation and cellular stress responses such as hydrogen peroxide protection, DNA repair, changes in polyamine (PA) metabolism and autophagy. Cellular PA contents and the expression of the PA metabolism genes arginine decarboxylase (ADC), spermidine synthase (SPDS), thermospermine synthase (ACL5) and diamine oxidase (DAO) were analyzed, as well as the expression of catalase (CAT), DNA repair genes (RAD51, KU80) and autophagy-related genes (ATG5, ATG8) throughout the induction of somatic embryo formation in Scots pine SE cultures. Among the embryo-producing SE lines, the expression of ADC, SPDS, ACL5, DAO, CAT, RAD51, KU80 and ATG8 showed consistent profiles. Furthermore, the overall low expression of the stress-related genes suggests that cells in those SE lines were not stressed but recognized the ABA+PEG treatment as a signal to trigger the embryogenic pathway. In those SE lines that were unable to produce embryos, cells seemed to experience the ABA+PEG treatment mostly as osmotic stress and activated a wide range of stress defense mechanisms. Altogether, our results suggest that the direction to the embryogenic pathway is connected with cellular stress responses in Scots pine SE cultures. Thus, the manipulation of stress response pathways may provide a way to enhance somatic embryo production in recalcitrant Scots pine SE lines.

Thermospermine Synthase (ACL5) and Diamine Oxidase (DAO) Expression Is Needed for Zygotic Embryogenesis and Vascular Development in Scots Pine

“…Therefore, we suggest that PAs may have more potential for biotechnological applications in development related processes such as somatic embryogenesis and the control of wood formation in Scots pine. During the induction of Scots pine somatic embryogenesis, the triggering of the embryo-producing pathway was connected with consistent changes in PA gene expression (Salo et al, 2016). Thus, the specific manipulation of PA gene expression might provide a way to enhance somatic embryo production in recalcitrant Scots pine lines.…”

“…For both clones one representative graft was selected and used repeatedly for cone collection in different years of the PA research (e.g. Vuosku et al, 2006;Vuosku et al, 2012;Muilu-Mäkelä et al, 2015a;Salo et al, 2016). For the present study, cones were collected four times during the growing season of 2004, on July 5 (sampling date I), July 12 (sampling date II), July 19 (sampling date III), and July 26 (sampling date IV).…”

“…In conifers PA contents and ratios followed developmental stage dependent profiles during zygotic embryogenesis (Minocha et al, 1999;Astarita et al, 2003;Vuosku et al, 2006;de Oliveira et al, 2017). Likewise, the triggering of somatic embryogenesis pathway modulated the PA metabolism at gene expression, enzyme activity, and metabolite levels (Minocha et al, 1999;Gemperlová et al, 2009;Jo et al, 2014;Salo et al, 2016). However, the reactions of embryogenic cell masses to exogenous PAs and PA biosynthesis inhibitors have proved to be complex and dependent on both the conifer species and the developmental stage of somatic embryos (Santanen and Simola, 1992;Minocha et al, 1993;Sarjala et al, 1997;Kong et al, 1998;Niemi et al, 2002;Steiner et al, 2007).…”

The Transcriptomic Responses of Pinus massoniana to Drought Stress

“…Drought stress results in the overproduction of ROS in plants [4]. The activation of many antioxidants that occurs due to drought is considered to be a protective mechanism against drought damage [47].…”

“…The involved genes are typically divided into genes encoding functional proteins and those encoding regulatory proteins. Functional proteins directly protect plants and include scavengers of ROS (reactive oxygen species) [4], aquaporins [5], dehydrins [6] and others. In contrast, regulatory proteins control gene expression networks and signal transduction pathways involved in stress responses [2].…”