Priming Maritime Pine Megagametophytes during Somatic Embryogenesis Improved Plant Adaptation to Heat Stress

Abstract

In the context of global climate change, forest tree research should be addressed to provide genotypes with increased resilience to high temperature events. These improved plants can be obtained by heat priming during somatic embryogenesis (SE), which would produce an epigenetic-mediated transgenerational memory. Thereby, we applied 37 °C or 50 °C to maritime pine (Pinus pinaster) megagametophytes and the obtained embryogenic masses went through the subsequent SE phases to produce plants that were further subjected to heat stress conditions. A putative transcription factor WRKY11 was upregulated in priming-derived embryonal masses, and also in the regenerated P37 and P50 plants, suggesting its role in establishing an epigenetic memory in this plant species. In vitro-grown P50 plants also showed higher cytokinin content and SOD upregulation, which points to a better responsiveness to heat stress. Heat exposure of two-year-old maritime pine plants induced upregulation of HSP70 in those derived from primed embryogenic masses, that also showed better osmotic adjustment and higher increases in chlorophyll, soluble sugars and starch contents. Moreover, ɸPSII of P50 plants was less affected by heat exposure. Thus, our results suggest that priming at 50 °C at the SE induction phase is a promising strategy to improve heat resilience in maritime pine.

Changing Temperature Conditions during Somatic Embryo Maturation Result in Pinus pinaster Plants with Altered Response to Heat Stress

“…Thus, needles of M28 plants showed significantly higher contents of ABA and total CK, mainly of CK bases and glucosides, and of cZand tZ-types, while M18 plants accumulated mainly iP-types, and both M18 and M28 plants showed higher contents of DZ-types. Higher basal levels of ABA, IAA, and CKs were also observed in primed maritime pine plants studied in our previous work, in which short temperature treatments at 37 or 50 • C were applied during the induction phase of the SE propagation protocol [18]. Variation in the hormone profiles of plants derived from somatic embryos matured at different temperatures resulted in significant coefficients of correlation between basal levels of ABA and total CKs (ρ = 0.804, p = 0.002); in addition, the total CK contents correlated with the content of active CK bases (ρ = 0.719, p = 0.001).…”

“…De Diego et al [51] also reported high levels of proline in radiata pine after recovery of a drought stress treatment, suggesting the possible implication of this metabolite in improving tolerance against future stress situations. In a previous work, we demonstrated that heat priming (50 • C) during SE induction improved performance under heat stress in Pinus pinaster, since primed plants showed better osmotic adjustment and higher increases in CKs, chlorophyll, soluble sugars, and starch contents, and their photosynthesis rates were less affected [18].…”

“…Efficient SE protocols have been developed for commercial pine species [11], and in particular to maritime pine materials from different Spanish provenances [6,16,17]. In addition, we have recently reported that a short-time high-temperature priming applied to megagametophytes induced heat tolerance in SE-derived maritime pine plants [18]. The high temperatures used for priming in these experiments (up to 50 • C) affected the efficiency of the SE process, as occurred when milder temperatures were employed along the different steps of the protocol.…”

“…Priming for improving thermotolerance in Pinus spp. by short heat treatments during SE induction affected DNA methylation and changed the expression pattern of stress-related genes in Pinus halepensis [28] and in P. radiata [29], and also altered gene expression patterns and the hormonal profile in P. pinaster plants [18]. Temperature during induction also had significant effects on the cytokinin (CK) profile of embryogenic lines of Pinus halepensis [30] and P. radiata [31].…”

Heat Stress in Pinus halepensis Somatic Embryogenesis Induction: Effect in DNA Methylation and Differential Expression of Stress-Related Genes

“…Furthermore, HSPs are closely involved in cellular protection, its structures and responses to heat stress being highly conserved amongst several organisms [29,56]. According to this, in maritime pine, when heat priming was performed at immature megagametophytes through SE, similar levels of expression were observed in primed and control EMs for HSP70 and SOD, but the expression of HSP70 at the derived in vitro somatic plants was higher in control plants [57]. In P. radiata, the gene coding for a heat shock protein (HSP20) was down-regulated in proliferating EMs and somatic plants coming from EMs initiated at high temperatures [42].…”