Figure - available from: Protoplasma
This content is subject to copyright. Terms and conditions apply.
Transmission electron microscopy of A. angustifolia proembryo.a–c Cap cells showing a cell wall (cw); b starch grains (s), vacuoles (v), and nuclei (n) with nucleoli (nu); c golgi complex (g) and mitochondria (m). d–f Embryonic cells showing d nucleus (n) with nucleolus (nu); e euchromatin (ec), heterochromatin (hc), mitochondria (m), and nuclear envelope (ne); f lipid bodies (lb) and cell wall (cw) with plasmodesmata (arrow). g–i Suspensor cells showing g starch grains (s), vacuoles (v), nuclei (n), nuclear envelope (ne) dismantling and presence of nuclear pore complexes (ncp with arrows) in cytoplasm; h plastid-like leucoplast becoming a plastolysome-like structure (pl) and autophagic vacuoles (v) with cytoplasm portion (arrow); i cellular corpse represented by only the cell wall. Bars: a = 10 μm; d, i = 5 μm; b, c, g, h = 2 μm; and e, f = 1 μm
Source publication
This study addresses gaps in our understanding of pre-fertilization and archegonia development and reinterprets embryonic ontogenesis from Burlingame (Bot Gaz 59:1–39, 1915) to the present based on timescale and structural features allowing us to determine functionally and developmentally accurate terminology for all these stages in A. angustifolia...
Citations
... Therefore, many aspects of the embryonic ontogenesis of A. angustifolia were updated by Goeten et al. (2020), and Herting and Stützel (2020) described the A. araucana seed scale ontogeny discussing the homology of this structure. The overall pollination mechanism of Araucaria was analyzed, which includes wind pollination, the presence of nonsaccate pollen, the absence of pollination drops, and extraovular pollen germination (e.g., Burlingame 1915;Haines et al. 1984;Owens et al. 1998), in which pollen grains laid on the seed scale, germinate and grow toward the seed cone axis straight to the micropyle. ...
Key message
In Araucaria angustifolia, the seed scale is part of the ovule, the female gametophyte presents a monosporic origin and arises from a coenocytic tetrad, and the pollen tube presents a single axis.
Abstract
The seed cone of conifers has many informative features, and its ontogenetic data may help interpret relationships among function, development patterns, and homology among seed plants. We reported the seed cone development, from pollination to pre-fertilization, including seed scale, ovule ontogeny, and pollen tube growth in Araucaria angustifolia. The study was performed using light microscopy, scanning electron microscopy, and X-ray microcomputed tomography (μCT). During the pollination period, the ovule arises right after the seed scale has emerged. From that event to the pre-fertilization period takes about 14 months. Megasporogenesis occurs three weeks after ovule formation, producing a coenocytic tetrad. At the same time as the female gametophyte's first nuclear division begins, the pollen tube grows through the seed scale adaxial face. Until maturity, the megagametophyte goes through the free nuclei stage, cellularization stage, and cellular growth stage. Along its development, many pollen tubes develop in the nucellar tissue extending straight toward the female gametophyte. Our observations show that the seed scale came out of the same primordia of the ovule, agreeing with past studies that this structure is part of the ovule itself. The formation of a female gametophyte with a monosporic origin that arises from a coenocytic tetrad was described for the first time in conifers, and the three-dimensional reconstruction of the ovule revealed the presence of pollen tubes with only one axis and no branches, highlighting a new pattern of pollen tube growth in Araucariaceae.
... Brazilian conservation laws impose limits on the exploitation of A. angustifolia (BRAZIL 2006;CONAMA 2001). Seeds of A. angustifolia are critical source materials to capture genetic diversity and propagate trees (Stefenon et al. 2009;Zechini et al. 2018;Goeten et al. 2020). However, seeds of this species quickly lose viability after drying and cannot be stored for more than a year in conventional seed banks (Farias-Soares et al. 2013;Gasparin et al. 2020). ...
... Sensitivity to desiccation of mature seeds is linked to the punctuation of metabolism occurring between late embryogenesis and early germination programs (Farrant et al. 1989;Pammenter and Berjak 2014;Leprince et al. 2017). Most of these studies involve angiosperm species, but there are few studies of maturation and germination metabolism in desiccation sensitive gymnosperm seeds Podocarpus henkelii (Henkel yellowwood) (Dodd et al. 1989)d angustifolia (Farrant et al. 1989), which might vary considerably from angiosperm seeds vis a vis important differences in early histodifferentiation as well as role of megagametophyte as a food source for the embryo (Goeten et al. 2020). ...
... Toward that end, we are compiling markers of the embryogenic program for A. angustifolia. Broad changes in polyamines (Astarita et al. 2003b;Oliveira et al. 2016), proteins (Santos et al. 2002;Silveira et al. 2008;Balbuena et al. 2009Balbuena et al. , 2011, carbohidratos (Navarro et al. 2017), amino acid ((Astarita et al. 2003a and histological characteristics (Rogge-Renner et al. 2013;Goeten et al. 2020) do not reveal differences in development of A. angustifolia embryos relative to other embryogenic programs that might explain differences in sensitivity to desiccation. The purpose of this paper is to map a broad portion of the embryogenic program using light microscopy and concomitant analyses of carbohydrate and protein markers known to indicate the onset of seed maturity and desiccation tolerance in other species. ...
Key message
Embryos in early-to-late embryo transition stage might be potentially able to respond the dehydration and cryopreservation process and in addition to producing somatic embryos.
Araucaria angustifolia (Bert. O. Kuntze) is a threatened and endemic species and takes almost 24 months after pollination (MAP) for seed maturation. Mature seeds lose viability if dried and do not survive prolonged storage which seems to be a consequence of metabolic profile. To understand how maturation occurs, we examined the biochemical changes within developing seeds. Carbohydrate and total protein contents as well as appearance of dehydrin-like proteins, were analyzed during the proembryogenic (15–6 MAP), early embryogenesis (17–18 MAP) and late embryogenesis (19–23 MAP) stages. Three embryos stages were describing in A. angustifolia seed development. Intense mitotic activity occurs during pro-embryogenic and early embryogenesis stages, which are also characterized by high fructose and glucose and low sucrose contents. During early embryogenesis, embryos had low total and heat-soluble protein contents and dehydrin-like proteins were not present. Interestingly, during late embryogenesis the sucrose to hexose ratio increased markedly and four fractions of dehydrin-like proteins with molecular masses near 21.5, 23, 26 and 27 kDa were detected. Our results show a high diversity of dehydrin-like proteins and carbohydrates over embryo development which are not necessarily associate to desiccation tolerance and this can explain Araucaria seed recalcitrant behavior. Also, it is proposed that embryo cryopreservation should be performed at early-to-late embryo transition stage and somatic embryogenesis might be improved by sucrose addition at maturation media. It was highlighted the seed biochemical composition of a primitive and threatened species including a discussion of wider implications for seed conservation.
... Este período entre la emisión de polen y la formación de semillas es muy extenso. Analizando histológicamente el desarrollo de los megaestróbilos, Goeten et al. (2020) determinaron que las semillas de A. angustifolia están maduras cuando alcanzan tonalidades marrones y 7 cm de largo, 20 meses después de ocurrida la polinación. Se utiliza el término polinación para indicar la emisión del polen por las anteras antes de que se deposite en el órgano sexual femenino (Faegri y van der Pijl 1979). ...
The reproductive cycle of Araucaria angustifolia, a critically endangered species due to the reduction of its original habitat and the lack of natural regeneration, has been studied. The scarcity of seeds in its distribution area makes reforestation a challenging restoration tool. This work analyses the influence of climate on pollen and seed production. Aerobiological monitoring and seed collection were performed during the 2015-2017 reproductive period in two regions of Argentina: San Antonio (SA), Misiones, a subtropical site; and 25 de Mayo (25M), Buenos Aires, a temperate area. The seasonality of pollination was similar between the sites (spring). However, the pollen amount was four times higher in 25M, with high hourly values throughout the day, and a shorter duration of about half the number of days compared to SA. Seed production was also higher in 25M (six fold). Reproduction proved to be favoured by the temperate climate due to higher summer temperatures prior to pollination (period of strobili initiation), lower winter temperatures (during pollen grain formation), and lower rainfall during pollen transport. Management guidelines involving ex-situ conservation of this native forest species are proposed.
... The A. angustifolia embryogenesis processes were analyzed during three developmental zygotic embryo stages (globular, cotyledonal, and mature) and two contrasting embryogenic cultures (Responsive and Blocked) in proliferation and maturation phases (Fig. 1). The globular stage refers to the early development of the zygotic embryo, and the cotyledonal and mature stages to the late embryo development (Goeten et al. 2020), with a difference of 60 days between them. The globular stage was constituted of the zygotic embryo surrounded by the megagametophyte. ...
Embryogenesis is a complex developmental process dependent on the intercommunication of coordinated signals. Among these signals, carbohydrate metabolism seems to be central, modulating responses according to the spatio-temporal environmental variations. Nitric oxide (NO) is a molecule that is extensively involved in various growth processes; however, the regulatory mechanism effect of NO signaling on carbohydrate metabolism is still largely unknown. Thus, in this study, we investigated the interaction between carbohydrate metabolism and NO homeostasis. First, the expression values of sucrose, raffinose, and starch metabolisms target genes were evaluated during Araucaria angustifolia zygotic and early somatic embryo development. This result demonstrated that starch metabolism targets highlighted the differences between zygotic embryo development and progression in early somatic embryo establishment. Therefore, non-structural carbohydrate content, NO emission, and starch metabolism-related genes were evaluated under treatments with donor, scavenger, or inhibitor of NO biosynthesis. These analyses were conducted at the somatic embryogenesis proliferation phase. In this work, the results showed that NO acted on the starch content, and in a different way between cell lines with contrasting responsiveness for the development of early somatic embryos. This indicates that the modification of starch accumulation by NO might be an effective strategy to optimize the in vitro conditions for A. angustifolia somatic embryo development.
... Due to the influence of climate and other factors, we attempted to determine which developmental stage of zygotic embryos is the best for SEMs initiation. Based on previous finding [29][30][31] and the morphological of megagametophyte, we defined four stages of whole zygotic embryos developmental from early July to mid-September. At www.nature.com/scientificreports/ the early July, most of embryos were at the pro-embryo stage; they developed into early embryogeny in mid-July. ...
Picea mongolica, a native species with excellent industrial wood quality and strong sand-fixing capacity, may be utilized in construction of urban green spaces in arid areas in China. However, now the sustainability of the ecosystems where this species grows is at serious risk due to a lack of natural regeneration. In this study, we developed an efficient regeneration system and comprehensively analyzed various factors affecting somatic embryogenesis (SE) using zygotic embryos as explants. We identified the optimal plant growth regulators (PGRs) performance and the best donor trees (k81) for the generation of somatic embryos (SEMs). Additionally, we confirmed that the positive developmental window of SEMs initiation was at the end of July to early August, which is when zygotic embryos was at the late embryogeny. In this time period, specific transcripts associated with the regulation of epigenetic modifications, plant hormone-related genes, and embryonic development-related transcription factors play important roles for early SEMs initiation. These results may provide a valuable resource for vegetative propagation of Picea mongolica. Our results may help to establish a reliable protocol for plantlet regeneration, which may facilitate urban greening applications and industrialization in arid areas.
... In conifers, the formation of multiple embryos is typical during the pro-embryonary stage, either by the occurrence of simple and cleavage polyembryony (Buchholz 1920(Buchholz , 1926Steeves and Sussex 1989;Williams 2009). Only polyzygotic polyembryony at early stages occurs in Araucaria (Figs. 1g,h) (Gifford and Foster 1989;Goeten et al. 2020). However, only one embryo remains in the mature seed, and the other polyembryos degenerate (Filonova et al. 2002;Bozhkov et al. 2005). ...
... In A. angustifolia, mature archegonia are observed at the 13th month after pollination, pollen tube germination occurs during the 14th month, and multiple polyembryos are observed at the 15th month (Goeten et al. 2020). During the pro-embryogenic phase, the embryo consists of a small apical region connected to a long tail formed by elongated cells. ...
... In this stage, pro-embryos possess three distinct cell types: cap cells, embryonic cells, and suspensor cells. Seventeen months after pollination occurs, cap cells have completely degenerated, giving rise to the early embryogenic stage of the dominant embryo (Steiner et al. 2015;Goeten et al. 2020). The early embryo presents a mass of embryogenic cells structured into an elongate cylindrical body with a rounded and smooth apical region, and a long suspensor system. ...
Somatic embryogenesis (SE) is a useful biotechnological tool to promote the conservation of plant genetic resources. Araucaria angustifolia, Acca sellowiana, and Bactris gasipaes are forest species with recognized ecological, cultural, and economic importance in the subtropical Atlantic Forest and the tropical Amazon Forest. SE protocols have been studied for these three species for more than 30 years. Despite the advances obtained during this time, some drawbacks remain. Here we reviewed and discussed the advancements and continuing constraints in the SE of these species, pointing out the more successful procedures. Further, we highlight several of the most pressing issues remaining in the SE protocols for each species. Further development of these protocols will aid in the conservation of these three species. Advances and drawbacks of the SE in three forest species of the Atlantic Forest and the Amazon Forest are discussed. Perspectives for overpassing the main shortcomings are presented.
Boswellia serrata is an important source of medicinal oleo-gum resins and boswellic acid, a unique non-steroidal anti-inflammatory, anti-arthritis drug. This experimental study describes the first time and in vitro direct somatic embryogenesis protocol and plantlet formation from immature zygotic embryos of B. serrata . The morphogenic potential of zygotic embryo was tested on MS medium fortified with cytokinins: 6-furfurylaminopurine [kinetin], N6-benzyladenine [BA], 1-Phenyl-3-(1,2,3-thiadiazol-5-yl)-urea [TDZ], auxins: IAA [Indole-3-acetic acid], NAA [α-naphthaleneacetic acid] and 2,4-D [2,4-dichlorophenoxyacetic acid] alone and in combination. MS medium containing 1mg/l TDZ + 0.50 mg/l 2,4-D + 200 mg/l PVP was effective for inducing direct somatic embryo in 89.63% of cultures. On the surface of the zygotic embryo observed, the globular, heart and cotyledonary stages of somatic embryo development. Better germination and conversion to plantlets (73.2 ± 1.6%) of somatic embryos were observed on subculture to GA 3 alone containing medium. The survival of plantlets under ex-vitro conditions was about 72%. Flow cytometry (FCM) confirmed the stable ploidy level in regenerated plantlets. In leaf samples of somatic embryo-derived plantlets, boswellic acid isomers are detected by using UPLC-UV-MS analysis and the content of isomer 3-O-acetyl-11-keto-β-Boswellic acid (AKBA) was 0.13 µg/gm dry weights. The propagation protocol described here provides an important micropropagation technique for this valued plant. Nevertheless, this described propagation method can be used for genetic transformation and medicinally important bioactive boswellic acid production.
