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Detection of calcium after pollination in Agave. a) Different intensities of calcium in the synergid cells. b) Optical section that allows to see the entrance of the pollen tube (pt) to the embryo sac. c) Helobial endosperm. d) Calcium in the zygote. sc=synergid cell, dsc=degenerated synergid cell, ec =egg cell, m=micropyle, ch =chalaza, pt =pollen tube, z=zygote, zn=zygote nucleus. Inset in b indicates synergid cell degenerated (asterisk) by the arrival of the pollen tube, arrow head in inset in d indicates nucleolus product of the male and female nucleoli fusion prior to the zygote elongation, and arrow heads in c indicate nucleoli. Bar = 20 μm (a–c) and bar = 10 μm (d)
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Calcium is a secondary messenger that regulates and coordinates the cellular responses to environmental cues. Despite calcium being a key player during fertilization in plants, little is known about its role during the development of the endosperm. For this reason, the distribution, abundance, and dynamics of cytosolic calcium during the first stag...
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Asparagaceae's large embryo sacs display a central cell nucleus polarized toward the chalaza, which means the sperm nucleus that fuses it during double fertilization migrates a long distance before karyogamy. Because of the size and inverted polarity of the central cell in Asparagaceae, we hypothesize that the second fertilization process is suppor...
Citations
... During reproduction of higher plants, actin filaments also play an important role; they are involved in pollen tube elongation (Vidali et al., 2001), vesicle and organelle transport (Drøbak et al., 2004;Cai and Cresti, 2009) and self-incompatibility responses (Roldán et al., 2012). Moreover, actin is involved in the female gametophyte development (Huang et al., 1999;Kawashima and Berger, 2015), in double fertilization (Huang and Sheridan, 1998;Kawashima et al., 2014) and the subsequent processes of endosperm (Świerczyńska and Bohdanowicz, 2003;Barranco-Guzmán et al., 2019) and embryo development in the seed (Kimata et al., 2016). ...
During plant sexual reproduction, F-actin takes part in the elongation of the pollen tube and the movement of sperm cells along with it. Moreover, F-actin is involved in the transport of sperm cells throughout the embryo sac when double fertilization occurs. Different techniques for analysis of F-actin in plant cells have been developed: from classical actin-immunolocalization in fixed tissues to genetically tagged actin with fluorescent proteins for live imaging of cells. Despite the implementation of live cell imaging tools, fixed plant tissue methods for cytoskeletal studies remain an essential tool for genetically intractable systems. Also, most of the work on live imaging of the cytoskeleton has been conducted on cells located on the plant's surface, such as epidermal cells, trichomes, and root hairs. In cells situated in the plant's interior, especially those from plant species with thicker organ systems, it is necessary to utilize conventional sectioning and permeabilization methods to allow the label access to the cytoskeleton. Studies about the role of F-actin cytoskeleton during double fertilization in plants with crassinucellate ovules (e.g., Agave, Yucca, Polianthes, Prochnyantes, and Manfreda) remain scarce due to the difficulties to access the female gametophyte. Here, we have developed a straightforward method for analysis of F-actin in the female gametophyte of different Agavoideae sub-family species. The procedure includes the fixation of whole ovules with formaldehyde, followed by membrane permeabilization with cold acetone, a prolonged staining step with rhodamine-phalloidin, and Hoechst 33342 as a counterstain and two final steps of dehydration of samples in increasing-concentration series of cold isopropanol and clarification of tissues with methyl salicylate. This technique allows the analysis of a large number of samples in a short period, cell positioning relative to neighbor cells is maintained, and, with the help of a confocal microscope, reconstruction of a single 3D image of F-actin structures into the embryo sac can be obtained.
... There are reports about the close phylogenetic relationship of Agave, Polianthes and Manfreda (Asparagaceae) (Bogler and Simpson, 1996;Eguiarte et al., 2000;Barba-Gonzalez et al., 2012;Thiede, 2012). Among these, there are reports on the female gametophyte development of Agave lechuguilla (Grove, 1941), A. virginica (Regen, 1941), A. fourcroydes and A. angustifolia (Piven et al., 2001) A. tequilana (Escobar-Guzman et al., 2008;González-Gutiérrez et al., 2014), Polianthes tuberosa (González-Gutiérrez and Rodríguez-Garay, 2016), A. colimana (Barranco-Guzmán, 2018) and A. salmiana (Barranco-Guzmán et al., 2019). ...
Manfreda Salisb. belongs to the Asparagaceae family and is a genus of a high ornamental potential due to the beauty and varied morphology of its leaves, as well as it provides flowers of different species throughout the year. In spite of its economic relevance, little is known about its sexual reproduction. This work aimed to characterize the female gametophyte development of Manfreda elongata through a stain-clearing technique of whole ovules and then analyzed by confocal microscopy. The female gametophyte development of M. elongata is of the monosporic Polygonum-type. The megaspore mother cell of M. elongata contains a dense and prominent nucleus, and at megasporogenesis, when meiosis II is completed, linear and T-shaped tetrads of megaspores were observed. Of this, only the megaspore closest to the chalaza remains functional. In the process of megagametogenesis, three rounds of free nuclear mitosis produced an eight-nuclei embryo sac. At maturity, a seven-celled embryo sac is formed by an egg apparatus (egg cell and two synergids) at the micropylar end, a central cell (with a diploid nucleus resulted from the fusion of the two polar nuclei). Thus, at the end of its formation, the female gametophyte development of M. elongata is of the monosporic, Polygonum-type.
Agave colimana, subgenus Littaea (Asparagaceae) is a wild plant that belongs to the Asparagaceae family, and in this species, the formation of its female reproductive system is unknown, probably because it has no
commercial use. However, A. colimana recently has been considered as a candidate for marketing as a succulent plant because of its floral and growth architecture. Knowledge about aspects of the formation of the embryo sac is useful for understanding its sexual reproduction, which is important for propagation by seed and breeding purposes.
Megasporogenesis and megagametogenesis were studied using Feulgen staining under confocal microscopy. Megasporogenesis resulted in the formation of a linear tetrad with the functional megaspore attached to the chalazal end. Subsequently, megagametogenesis from the functional megaspore was followed by three mitotic divisions generating an embryo sac with seven cells: two synergid cells, one egg cell, three antipodal cells, and one central cell with a 2n nucleus. The embryo sac of Agave colimana was found to be monosporic of the Polygonum-type considered as the typical type in the majority of angiosperm plants. Also, abnormal embryo sacs were observed, following possible errors during meiosis.
