One tissue, two fates: different roles of megagametophyte cells during Scots pine embryogenesis

Abstract

In the Scots pine (Pinus sylvestris L.) seed, embryos grow and develop within the corrosion cavity of the megagametophyte, a maternally derived haploid tissue, which houses the majority of the storage reserves of the seed. In the present study, histochemical methods and quantification of the expression levels of the programmed cell death (PCD) and DNA repair processes related genes (MCA, TAT-D, RAD51, KU80, and LIG) were used to investigate the physiological events occurring in the megagametophyte tissue during embryo development. It was found that the megagametophyte was viable from the early phases of embryo development until the early germination of mature seeds. However, the megagametophyte cells in the narrow embryo surrounding region (ESR) were destroyed by cell death with morphologically necrotic features. Their cell wall, plasma membrane, and nuclear envelope broke down with the release of cell debris and nucleic acids into the corrosion cavity. The occurrence of necrotic-like cell death in gymnosperm embryogenesis provides a favourable model for the study of developmental cell death with necrotic-like morphology and suggests that the mechanism underlying necrotic cell death is evolutionary conserved.

Pine embryogenesis

“…The vacuole collapse triggers a brief culmination phase which results in the complete removal of the protoplast. 2 Finally, the dying subordinate embryos break down and may be used for the nutrition of the leading embryo via the CC fluid 7 ( Fig. 1D).…”

From Seed to Tree: The Functioning and Evolution of DNA Repair in Plants

“…The megagametophyte houses the majority of the storage reserves of a seed (King & Gifford, 1997) and provides nutrition for the developing embryo during seed development as well as for the young seedling during early germination (Fig.1C). We have shown that, in Scots pine seed, the megagametophyte tissue stays alive from the early phases of embryo development until the imbibition phase of early germination of mature seed, except for the cells in the ESR (Vuosku et al, 2009). Positive signals in TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling) assay indicate DNA fragmentation in the nuclei of the megagametophyte cells at the late embryogeny (Fig.1D).…”

“…1B), while subordinate embryos, as well as suspensor tissue, are deleted by programmed cell death (PCD) during the progress of seed development (Filonova et al 2002). Megagametophyte cells in the embryo surrounding region (ESR) die through necrotic-like cell death (Vuosku et al, 2009), and in addition, the maternal cells of the nucellar layers face destruction during early embryogenesis (Hiratsuka et al, 2002;Vuosku et al, 2009). In a gymnosperm seed, the megagametophyte tissue develops from a haploid megaspore before the actual fertilization of the eggs (Singh 1978).…”

“…Instead, the DNA fragmentation may be a consequence of DNA strand breaks caused by maturation drying or by the DNA breaks with free 3'-OH ends that appear during DNA repair. During the seed development, the expression of RAD51 gene decrease, whereas the expression of the KU80 and DNA ligase (LIG) genes remain constant, which suggests that the proportion of mitotic cells decrease and the DNA breaks are mainly repaired by NHEJ pathway (Vuosku et al, 2009). Nuclear DNA fragmentation is currently one of the most frequently used sign of PCD.…”

“…Nuclear DNA fragmentation is currently one of the most frequently used sign of PCD. However, in the Scots pine seed, the megagametophyte cells remain metabolically active until the imbibition phase of germination despite DNA fragmentation in the nuclei already during late seed development (Vuosku et al, 2009). In plants, both the tolerance of DNA fragmentation and effective DNA repair mechanisms may be adaptations to the special energy metabolism as well as to a sessile life style which exposes cells to various endogenous and exogenous stresses.…”