Developmental and Cytochemical Features of Female Gametophyte in Endemic Lathyrus undulatus (Fabaceae).
The presented study describes developmental and cytochemical features of female gametophyte in Lathyrus undulatus Boiss. (Fabaceae), which belongs to subfamily Papilionoideae and is endemic to northwestern Turkey. The ovary is monocarpellate and the mature ovule is anatropous, bitegmic and crassinucellate. Differentiation of the outer integument determines a Y- shaped micropyle aperture. The nucellar epidermis is poorly developed. The inner parietal layers of the integument differentiate into a thin layer of endothelium. The hypodermally oriented archesporium is distinguished as one cell. Archesporium produces a parietal cell and a primary sporogenous cell. The primary sporogenous cell enlarges to form megasporocyte. After meiotic division, megaspores organize in an isobilateral manner of tetrad. Only one megaspore functions and it is one of the chalazal members of the tetrad. The functional megaspore develops into an embryo sac, which gives rise to a Polygonum type.
The synergids lying above the egg cell are typical in structure. The antipodals are ephemeral and degenerate earlier without leaving any remnants. Embryo development is of Onagrad type. Suspensor degenerates at the globular embryo stage. The primary endosperm nucleus divides mitotically in a free nuclear manner and it is rich for insoluble polysaccharides and protein. (c) 2013 Friends Science Publishers
Keywords: Lathyrus undulatus; Megasporogenesis; Megagametogenesis; Embryo; Endosperma
The genus Lathyrus L. (Fabaceae) comprises approximately 200 species, most of which are annual and perennial plants, predominantly centered in the Mediterranean region. Lathyrus is represented by 78 Lathyrus taxa, 24 of which are endemic to Turkey (Davis, 1970; 1988; Gunes and Cirpici, 2008; 2011). Lathyrus species have high ecological and economical importance including drought resistance, erosion preventation, food, agricultural processes and ornamental purposes (Gunes, 2011).
Anatomical, palynological, caryological and seed morphological studies on some species of Lathyrus have been performed to elucidate the phylogenetic relationship of Lathyrus genus within the subfamily Papilionoideae (Perveen and Qaiser, 1998; Tosheva et al., 2004; Tosheva and Tonkov, 2005; Abou-El-Enain et al., 2007; Gunes and Cirpici, 2008; 2010; Gunes and Aytug, 2010).
It has been known that there are some taxonomic problems between the members of this genus that can not be solved by anatomical and morphological characters (Mantar et al., 2003). However, despite the usefulness of the embryological characters in this type of analysis, there are only a few studies on male and female gametophyte development in Lathyrus genus (Latter, 1925; Rembert, 1969; Davies and Williams, 1985). Recently, developmental, cytochemistry and programmed cell death studies were reported on the anthers of Lathyrus undulatus in detail (Vardar and Unal, 2011a; 2011b; 2011c; 2012).
Besides, the developmental features of megagametophyte and embryo seem to be very significant not only for systematic comparisons, but also for the knowledge of development and fertilization process. To the best of our knowledge, only the megasporogenesis of L. latifolius (Rembert, 1969a) and L. sativus (Roy, 1933) were carried out and there is no previous data available on the megagametophyte and embryo development of L. undulatus as well as the other members of Lathyrus genus.
The present study reports the first observations on the development and cytochemistry of ovule in L. undulatus Boiss. (Fabaceae), which belongs to Papilionoideae subfamily and is endemic to northwestern Turkey. This study will provide a detailed understanding of the events that leads to embryo formation. Information on the development of the female reproductive structures in L. undulatus will advance our understanding of its reproductive process, and contribute to understand taxonomic relationship with closely related taxa within the Papilionoideae.
Materials and Methods
Flower buds of Lathyrus undulatus Boiss. (Fabaceae)
growing in natural habitats in the vicinity of Beykoz-Istanbul (Turkey) was collected in March-April 2008. Flower buds were fixed in acetic:alcohol (1:3, v/v) for 24 h at room temperature. After dehydration in a graded series of ethanol, the material was embedded in paraffin. Sections (8-10 um) were cut using a Leica RM2125RT microtome and stained with Delafield's hematoxylin.
Insoluble polysaccharides were localized according to periodic acid-Schiff's (PAS) method of Feder and O'Brien (1968). The sections were immersed in a solution of 1% periodic acid dissolved in 96% ethanol for 20 min. Staining in Schiff's reagent was carried out for 30 min in the dark. The presence of carbohydrate in the tissue is indicated by the appearance of a purplish-red color. Proteins were localized according to a method of Fisher (1968). The sections were stained with 0.2% Coomassie Brilliant Blue dissolved in a methanolic solution, methanol:acetic acid:
water (5:1:4), for 30 min at 60degC. The presence of protein is indicated by the appearance of a blue color.
The sections photographed with the ProgRes Capture Pro 2.6 software, assisted by a Jenoptik 122CU color camera and an Olympus BX-51 microscope.
In L. undulatus the ovary is monocarpellate. The mature ovule is anatropous, bitegmic and crassinucellate. The inner integument initiates first, and then the outer integument develops into a small protuberance at the megaspore mother cell stage. Throughout the development the outer integument becomes multilayered and swells at the micropylar region where it consists of 9-10 layers of cells.
The differentiation of the outer integument, in the proximal flank of the primordium determines a Y-shaped micropyle aperture (Fig. 1a). Besides, the inner integument remains uniformly two layered which include numerous starch grains at the stage of mature embryo sac. The asymmetrical integument growth creates the anatropous curvature. The nucellar epidermis is poorly developed and composed of 4-5 layers of cuboidal cells. The inner parietal layers of the integument differentiate into a thin layer of endothelium with cuboidal cells comprises strong PAS positive reaction (Fig. 1b, c).
Megasporogenesis is initiated by the development of an archesporium hypodermally oriented in the nucellus. The archesporium is distinguished as one cell, which represents a large volume, dense cytoplasm and distinct nucleolus. Development continues with the mitotic division of the archesporium cell, producing a parietal cell and a primary sporogenous cell (Fig. 2a). Parietal cell remains undivided in the nucellus. The primary sporogenous cell enlarges to form the megasporocyte (megaspore mother cell-MMC) (Fig. 2b). The MMC undergoes meiosis I to form equally-sized dyad. In L. undulatus as distinct from the common at the end of meiosis I the wall formation is longitidunal (Fig. 2c-f). The meiotic division in the MMC is synchronic with the microspore mother cells.
At the conclusion of the meiotic divisions in the ovule of L. undulatus, an unusual arrangement of tetrad is formed (Fig. 2g, h). The megaspores organize in an isobilateral manner of tetrad. Only one megaspore functions and it is always one of the two larger chalazal members of the tetrad (Fig. 2i). At about the functional megaspore stage the ovule is completely anatropous.
The functional megaspore develops into an embryo sac (ES), which increases in length, extending into the micropylar region. Mitotic divisions in the functional megaspore result in 2, 4 and 8-nucleate ES (Fig. 3a-c). These nuclei reorganize to give rise to a Polygonum type of ES. Three nuclei from the micropylar quartet develop into an egg apparatus consisting of an egg and two synergids, which are PAS positive (Fig. 3d, e). The synergids lying above the egg cell are typical in structure. The antipodals, formed from the three nucleus of chalazal quartet, are situated on the periphery at the chalazal end of the ES (Fig. 3f). The antipodals are ephemeral and degenerate earlier without leaving any remnants. The fourth nuclei from the micropylar and chalazal quartet form the polar nuclei (Fig. 3g). The two polar nuclei fuse to form a secondary nucleus before fertilization, which is situated by the side of the egg. The central cell contains PAS positive granules (Fig. 3h).
After fertilization, the synergids degenerate simultaneously (Fig. 3i), and secondary nucleus gives rise to the primary endosperm nucleus.
Embryo development is of Onagrad type. Zygote presents at the micropylar pole of the ES and smaller than the egg cell (Fig. 4a). The volume decrement is due to the reduction of vacuole. The first division of zygote is transverse giving rise to an apical cell ca and a basal cell cb almost equal in size (Fig. 4b). Apical cell ca divides transversely. The two daughter cells of ca commonly divide by a vertical wall and constitues to form a proembryo of four cells. Subsequent divisions give rise to globular embryo. Basal cell cb divides also transversely forming two superimposed cells (Fig. 4c, d). The basal cell constitutes the suspensor, which becomes larger. The suspensor persists until globular embryo and then degenerates (Fig. 4e, f).
The primary endosperm nucleus prior to division in the zygote divides mitotically in a free nuclear manner. Free nuclei remain embedded in a cytoplasmic sheet around the central vacuole (Fig. 5a). The endosperm is rich for insoluble polysaccharides and protein carried out by PAS and Coomassie Brilliant Blue, respectively (Fig. 5b, c).
The reproductive and embryological knowledge of economically important plants, belonging to Fabaceae, provides useful data in relation to the fields of cell biology, reproductive ecology and taxonomy and for purposes related to seed production and cross-breeding. According to embryological data (Johri et al., 1992), considerable information is available on this family, with greater emphasis on subfamily Papilionoideae. Although anatomy, pollen morphology and the chromosome number were most widely analyzed in Lathyrus genus, which belongs to Papilionoideae, male and female gametophyte development was not noticeable. Recently development, cytochemistry and programmed cell death studies have been performed on the anthers of L. undulatus in detail (Vardar and Unal, 2011a; 2011b; 2011c; 2012). Besides, there are only a few studies on female gametophyte development, which consist only of megasporogenesis in Lathyrus genus (Latter, 1925; Rembert, 1969a; Davies and Williams, 1985).
The ovule characters are important particularly for the Fabaceae family such as ovule shape and type, the integuments and the participation of integuments in micropyle formation. The ovule of L. undulatus is anatropous, bitegmic and crassinucellate consistent with Fabaceae family. Davis (1966) indicated that the micropyle is formed by both integuments and often markedly zig-zag in Papilionaceae (synonym of Fabaceae). Although micropyle is generated by both of the integuments similar with the other family members, it is distinctively Y-shaped in L. undulatus, which was also reported in Adesmia latifolia (Moco and Mariath, 2003). Besides the inner integument consists of two cell layers in L. undulatus, while the outer integument comprises of several layers as common in Fabaceae (Hindmarsh, 1964; Deshpande and Bhasin, 1974; Rembert, 1977; Ashrafunnisa and Pullaiah, 1994; 1999; Moco and Mariath, 2003).
As it was observed in the presented study, the nucellus of the Fabaceae ovules is crassinucellate (Prakash, 1987), and which has been considered to be the primitive condition in angiosperms (Sporne, 1969).
Rembert (1969a) compared megasporogenesis of 16 species of Papilionaceae family and indicated that in this family an archesporium, whether multicellular or a single cell, is characteristically hypodermal. The researcher explained that in hypodermal development a parietal cell and a primary sporogenous cell are produced in 14 of the 16 species and the primary sporogenous cell enlarges to form a single megasporocyte in these 14 species, as it was presented in L. undulatus.
Rembert (1966; 1969a; 1969b; 1971) used the marked variability in the megaspore tetrads to make phylogenetic hypotheses in Fabaceae. The author described the occurence of ten types of tetrads (linear with chalazal or epichalazal functional megaspore, T-shaped, oblique linear, inverted T- shape, anisobilateral, triad and three different types of bisporic patterns) and supposed evolutionary lines with in the family. Roy (1933) reported T-shaped tetrad for L. sativus. Besides, a related species L. latifolius displays a unique pattern of megaspore tetrads as anisobilateral (Rembert, 1969a). According to our results, as distinct from the other Lathyrus species L. undulatus exhibited an isobilateral tetrad.
In all other species of Papilionaceae it is the chalazal megaspore that functions in megagametogenesis and in no case is more than one megaspore known to function in this family. In general chalazal megaspore of tetrad develops into Polygonum type embryo sac (Davis, 1966; Rembert, 1969a). These are also the cases in L. undulatus.
The monosporic, Polygonum type of female gametophyte is typically a seven-celled structure at maturity. However, this structure may be reduced by cell death.
In Arabidopsis the antipodal cells underwent cell death before fertilization (Yadegari and Drews, 2004). Similarly ephemeral antipodals were observed in L. undulatus which is consistent with the Papilionaceae (Davis, 1966). Davis (1966) represented that embryogeny conforms either to the Onagrad, Asterad or Caryophyllad type and the suspensor shows great variation between the species in Papilionaceae. L. undulatus shows Onagrad type embryogeny and large suspansor, which persists until globular embryo.
It has been reported that endosperm formation is nuclear in Papilionaceae (Davis, 1966). In L. annuus and Stylosanthes mucronata wall formation is suppressed and the endosperm remains free-nuclear (Davis, 1966). Similar was the case for L. undulatus.
In conclusion; our data provide a new look at the aspect of sexual reproductive potential of L. undulatus in the genus Lathyrus. On the other hand, developmental and cytochemical features will be the object of fertilization and embryo growth investigations in Lathyrus thereby in subfamily Papilionoideae. Moreover, embryo sac features will provide useful characters in assessing relationships within this genus and family.
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1Marmara University, Science and Art Faculty, Department of Biology, Goztepe, 34722, Istanbul, Turkey
For correspondence: firstname.lastname@example.org
To cite this paper: Vardar, F., 2013. Developmental and cytochemical features of female gametophyte in endemic Lathyrus undulatus (Fabaceae). Int. J. Agric. Biol., 15: 135-139
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|Publication:||International Journal of Agriculture and Biology|
|Date:||Feb 28, 2013|
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