Structure and function of the hypertrophic synergid in some species of genus Allium L.
ABSTRACT The egg cell in the embryo sac of flowering plants is generally accompanied by two symmetrical cells, called synergid cells, which usually contains haploid nucleus. However, in some species of genus Allium L. mainly one and sometimes both of the synergid cells enlarge in size, undergo endopolyploidization and become hypertrophic. We have studied structure of the synergid cells of Allium atroviolaceum Boiss., A. rotundum L., A. fistulosum L. and A. cepa L. and determined DNA amount (C value) in the synergid cells of A. atroviolaceum. Cytophotometric study of various Feulgen-stained synergid and integument nuclei revealed clear difference in DNA content both in different types of the cells and in different stages of synergid development. The amount of DNA measured in newly formed synergid was already equal to 2C value found in the integument cell nucleus. At the time of fertilization the DNA amount in synergid is 4C. At the stage of proembryo it is already 6C and before degeneration at the late globular stage of embryo development 8C. No sign of mitotic cell division or formation of metaphase plate have been observed in any of investigated materials. It is assumed that endopolyploidization might determine longer persistent of intact synergid and increase of its trophic function.
- Plant Systematics and Evolution - PLANT SYST EVOL. 01/1957; 104:1-24.
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ABSTRACT: The synergid cells of the female gametophyte play a role in many steps of the angiosperm fertilization process, including guidance of pollen tube growth to the female gametophyte. However, the mechanisms by which the synergid cells become specified and develop their unique features during female gametophyte development are not understood. We identified MYB98 in a screen for Arabidopsis thaliana genes expressed in the female gametophyte. MYB98 is a member of the R2R3-MYB gene family, the members of which likely encode transcription factors. In the context of the ovule, MYB98 is expressed exclusively in the synergid cells, and mutations in this gene affect the female gametophyte specifically. myb98 female gametophytes are affected in two unique features of the synergid cell, pollen tube guidance and the filiform apparatus, but are otherwise normal. MYB98 also is expressed in trichomes and endosperm. Homozygous myb98 mutants exhibit no sporophytic defects, including trichome and endosperm defects. Together, these data suggest that MYB98 controls the development of specific features within the synergid cell during female gametophyte development.The Plant Cell 12/2005; 17(11):2981-92. · 9.25 Impact Factor
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ABSTRACT: In flowering plants, the egg cell is generally accompanied by two symmetrical cells, called synergid cells. As early as the 1870s, synergid cells were distinguished from egg cells and cooperation between synergid and egg cells was proposed; the term "synergid" is derived from the Greek "synergos," which means "working together." The accumulation of morphological and genetic data, and, more recently, the in vitro physiological analysis of the fertilization system of Torenia fournieri, have revealed that synergid cells work together with egg and central cells to accomplish double fertilization. This cooperation is of crucial importance in the attraction and acceptance of the pollen tube. In this review article, I focus on the physiological function and behavior of the synergid cell during the fertilization process.Journal of Plant Research 05/2002; 115(1118):149-60. · 2.06 Impact Factor
Structure and function of the hypertrophic synergid in some species of genus Allium L.
N. Nadirashvili, G. Gvaladze, M. Akhalkatsi
Institute of Botany, Georgian Academy of Sciences
The egg cell in the embryo sac of flowering plants is generally accompanied by two symmetrical
cells, called synergid cells, which usually contains haploid nucleus. However, in some species of
genus Allium L. mainly one and sometimes both of the synergid cells enlarge in size, undergo
endopolyploidization and become hypertrophic. We have studied structure of the synergid cells
of Allium atroviolaceum Boiss., A. rotundum L., A. fistulosum L. and A. cepa L. and determined
DNA amount (C value) in the synergid cells of A. atroviolaceum. Cytophotometric study of
various Feulgen-stained synergid and integument nuclei revealed clear difference in DNA
content both in different types of the cells and in different stages of synergid development. The
amount of DNA measured in newly formed synergid was already equal to 2C value found in the
integument cell nucleus. At the time of fertilization the DNA amount in synergid is 4C. At the
stage of proembryo it is already 6C and before degeneration at the late globular stage of embryo
development 8C. No sign of mitotic cell division or formation of metaphase plate have been
observed in any of investigated materials. It is assumed that endopolyploidization might
determine longer persistent of intact synergid and increase of its trophic function.
Allium-is gvaris zogierTi saxeobis hipertrofuli sinergidas struqtura da is gvaris zogierTi saxeobis hipertrofuli sinergidas struqtura da
n. nadiraSvili, g. RvalaZe, m. axalkaci
saq. mecn. akademiis botanikis instituti
kvercxujredi Cveulebriv garSemortymulia ori
simetriulad ganlagebuli sinergidaTi, romlebic, rogorc wesi, haploidur birTvs
Seicaven. Tumca, Allium-is gvaris zogierT saxeobaSi erTi an zogjer orive
sinergida imatebs zomaSi, ganicdis endopoliploidizacias da xdeba hipertrofuli.
Cven SeviswavleT sinergidebis struqtura Allium-is gvaris Semdeg saxeobebSi Allium
atroviolaceum Boiss., A. rotundum L., A. fistulosum L. da A. cepa L. da ganvsazRvreT dnm-is
raodenoba A. atroviolaceum-is sinergidas birTvSi. Catarda fiolgeniT SeRebili
sinergidas da integumentis birTvebis citofotometriuli Seswavla, romelmac
gamoavlina mkveTri sxvaoba dnm-is raodenobas Soris, rogorc gansxvavebuli tipis
birTvebSi, ise sinergidas ganviTarebis sxvadasxva stadiaze. dnm-is raodenoba
sinergidas formirebisTanave Seadgenda 2C–s. ganayofierebis win igi udrida 4C-s.
proembrionis stadiaze 6C–s, xolo degeneraciis win, Canasaxis ganviTarebis gvian
globularul stadiaze, 8C-s. navaraudebia, rom endopoliploidizacia ganapirobebs
sinergidas arsebobis gaxangrZlivebas da misi trofikuli funqciis gazrdas.
Key Words: Embryology, ovule, synergid, endopolyploidy
The egg cell in the embryo sac of flowering plants is generally accompanied by two
symmetrical cells, called synergid cells, which usually contains haploid nucleus like as the other
cells of the female gametophyte. However, in some species of genus Allium L. (Allium cepa, A.
nutans, A. rotundum, A. schoenoprasum, A. uniflorum, A. ursinum etc.) mainly one and
sometimes both of the synergid cells enlarge in size, undergo endopolyploidization and become
hypertrophic (Weber, 1929; Tschermak-Woess, 1950; Hasitschka-Jenschke, 1957; Gvaladze,
1962,1976; Sokolov, 1968; Batygina, 1990). The mechanism and the role of this phenomenon,
however, are not known until now.
The general role of the synergids in the embryo sac is assumed to be cooperation with
egg and central cells to accomplish double fertilization. This cooperation is of crucial importance
in the attraction and acceptance of the pollen tube (Higashiyama, 2002). The last develops from
the pollen grain after germination on the stigma and carries two male gametes through the
maternal reproductive tissues to the embryo sac, which contains two female gametes, egg and
central cells. The sperm cell of a flowering plant cannot migrate unaided and it must be
transported by the pollen tube before successful fertilization can occur. The mechanism of
guidance of the pollen tube from stigma to the embryo sac has been studied for more than a
century. Nowadays, it is determined that synergids play most significant role in this process
attracting pollen tube due to chemotropic and diffusible signals (Higashiyama, 2002).
The pollen tube penetrates into one of the synergid cells and releases its two male
gametes leading to the degeneration of the synergid cell. The second persistent synergid remains
intact during some period after fertilization and degenerates gradually. Two male pronuclei enter
the egg and the central cells and accomplish syngamy (fusion of sperm nucleus with egg
nucleus) and triple fusion (unification of the sperm and two polar nuclei). These two processes
represent consistent steps of double fertilization determining formation of embryo and
The function of synergids determines their structure. They develop distinct filiform
apparatus, some kind of cell wall protuberances at the micropylar end enlarging plasmalemma
surface and plying a role in reception of pollen tube. The cytoplasm contains abundant
organelles, such as mitochondria, endoplasmic reticulum, and plastids, which indicate that they
are metabolically active cells. No cell wall is present at the chalazal part of a synergid, and there
are some flocculent materials and vesicles in the spaces of cytoplasm membranes among
synergid, egg cell and central cell in embryo sacs. It is assumed that synergids besides
participation in double fertilization have trophic role supplying embryo sac with nutritive
substances (Higashiyama, 2002).
In the most flowering plants, both synergids show similar structure before fertilization
and contain haploid nuclei. The fact of synergid proliferation in some species of genus Allium,
however, should be indicative of different functional role, which it might play during seed
formation. So far, not much is known about a special role of such giant synergid. According to
literature data, synergids of A. ursinum undergo endomitosis direct after cell formation in the
embryo sac and become polyploid. At the fertilization time, the degenerating synergid receiving
pollen tube contains 4n, and the nucleus of persistent synergid 16n set of chromosomes
(Hasitschka-Jenschke, 1957). Endomitosis in one of the synergid of A. nutans lead to formation
of polytene chromosomes and cause proliferation of the nucleus, while the second synergid
remains haploid. Histochemical studies have revealed that the cytoplasm of the proliferated
synergid before fertilization when compare with the egg cell contains more storage substance,
such as polysaccharides, rRNA, proteins, fats (see Batygina, 1990). The nucleus is more
intensively stained for DNA and RNA. During endosperm development the storage substances
disappear from the synergid, which according to some authors might be indicative of trophic
function of them (Batygina, 1990). In spite of this data, in generall information about factors
determining synergid proliferation in the genus Allium is scarce and it is not known what the
function of this phenomenon is.
Throughout growth and development, eukaryotic organisms must co-ordinate DNA
synthesis, chromosome segregation, and cell division in order to generate differentiated cells
with the proper complement of chromosomes. The amount of DNA in plant nuclei has been
possible to estimate for over 50 years. Work on plants has played a leading part in research to
describe and understand the origin, extent and effects of variation in the DNA amount in the
unreplicated haploid nuclear chromosome complement defined as the 1C-value. Nowadays, C-
values are determined for over 4100 species of angiosperms. They vary c. 1000-fold from c. 0.11
pg in Fragaria viridis to 127.4 pg in Fritillaria assyriaca (Bennett, Leitch, 1995). Polyploids are
expected to have larger C-values than their diploid progenitors, increasing in direct proportion
with ploidy. Similarly, basic genome size is predicted to be the same at all levels of ploidy.
Endopolyploidization, i.e. the existence of different ploidy levels (labelled as 2C, 4C, 8C…) in
adjacent cells of a species, is a common phenomenon in seed plants. Nevertheless, the biological
significance of endopolyploidy is not yet clear.
In the present study we addressed the following questions: 1) What is the nature of
synergid proliferation in species of genus Allium; 2) what is biological function of synergid
proliferation; 3) what is DNA C-value in the nucleus of proliferated synergid.
Materials and Methods
The following species of genus Allium L. have been used in this study: Allium
atroviolaceum Boiss., A. rotundum L. (genus Allium L., subg. Allium, sect. Allium), A.
fistulosum L. (genus Allium L., subg. Rhizirideum (G.Don ex Koch) Wendelbo, sect. Cepa
(Mill.) Prokh., subsect. Phyllodolon (Salisb.) Kamelinand) and A. cepa L. (genus Allium L.,
subg. Rhizirideum (G.Don ex Koch) Wendelbo, sect. Cepa (Mill.) Prokh., subsect. Cepa (Mill.)
For light microscopy, buds, flowers and fruits have been collected at different stages of
development, fixed in FAA (formalin, acetic acid, 70% ethanol, 5:5:90) and embedded in
paraffin. 10-12 µm thick sections were prepared on microtome Reichert, Austria, and stained in
hematoxylin according to known method by Meier. Examination was carried out using light
microscope Polivar, Reichert, Austria.
The investigation of endopolyploidization pattern in the nucleus of giant synergid of A.
atroviolaceum (2n=16) was carried out by cytophotometric measurements of DNA content.
Investigation was carried out in the Laboratory of Cytology at the Institute of Zoology of the
Georgian Academy of Sciences. DNA content was determined by scanning cytophotometer,
Reichert, Austria, with one-wave cytophotometric method (wave length was 550 µm) on sections
after Feulgen staining. Probe diameter was 0,63 µm. All nuclei were scanned with the same
magnification (ocular x15, objective x40). DNA content was determined using the formula
C=VA, where C is DNA content in a single nucleus, V – volume of a nucleus, A – DNA
concentration on the section of a nucleus. Error of the method is 5%. Obtained data were
processed statistically (Brodskii, 1956). DNA content of the integument cell nucleus (number of
measured nuclei x=30) in the ovule of A. atroviolaceum was used as reference for species-
specific diploid DNA content. DNA content in synergid nuclei was measured at different stages
after cell formation in the embryo sac up to late globular stage of the embryo development
The ovule in all studied species is ortho-campylotropous, crassinucellate, with funicular
obturator covering micropyle (Fig. 1 A, B C). It is bitegmic. Embryo sac in investigated species
develops according to the bisporic Allium type. The megaspore nuclei in the diade undergo two
successive mitotic divisions and form 8-nucleate embryo sac with 3-celled egg apparatus,
containing egg cell and two symmetrically located synergids, the central cell with two polar
nuclei, and 3 antipodals. During maturation of the embryo sac, one of the synergids enlarges in
size considerably and becomes giant (Fig.1 B,D,E,F). It has much bigger sizes than the second
synergid (Fig.1 G,H,I). Filiform apparatus persists in both bigger and smaller cells, but, it is
more prominent in the hypertrophic one. Polar nuclei in the central cell do not fuse before
fertilization. However, they form close contact with each other (Fig.1 D). Antipodals, as usual,
are ephemeral and degenerate during maturation of the embryo sac.
The morphology of the giant synergid changes in different developmental stages. Soon
after formation it contains larger nucleus than the egg cell and the second synergid (Fig.1 D).
The form of the nucleus is lobed and irregular. At early stages of the development, the synergid
nucleus possess heavily condensed chromatin region, which by position and structure
corresponds to polytene chromosomes. The condensation increases during fertilization and
afterwards during embryogenesis (Fig.1 H,I). Simultaneously enlarges cell volumes of the
synergid. In A. cepa we have observed that the second synergid, which receives the pollen tube,
enlarges together with the giant synergid. It might have lesser size or in some cases be equal to
the giant one (Fig.1 I).
Before fertilization both synergis are intact. Shortly before fertilization, cytoplasm and
nuclear content of the smaller synergid become more condensed. The pollen tube penetrates into
the smaller synergid cell and releases its contents. After this process the synergid degenerates.
The persistent giant synergid remains intact until later stages and degenerates at late globular
stage when the embryo becomes pear shaped.
Cytologically, the structure of the somatic interphase nuclei of the integument tissue is
euchromatic and contains more or less uniformly distributed chromocenters, which should
correspond to the constitutive heterochromatin. The chromatin is condensed and distributed
uniformly in the nucleus.
Cytophotometric study of various Feulgen-stained synergid and integument nuclei
revealed clear difference in DNA content both in different types of the cells and in different
stages of synergid development. The amount of DNA measured in newly formed synergid was
already equal to 2C value found in the integument cell nucleus. The increase of DNA content in
the synergid nucleus starts before fertilization and continues until late globular stage of embryo
development, when degeneration processes start (Fig. 2). The following dynamic was observed.
Newly formed synergid in A. atroviolaceum contains 2C DNA. At the time of fertilization this
amounts to 4C. At the stage of proembryo it is already 6C and at the late globular stage of
embryo development 8C. No sign of mitotic cell division or formation of metaphase plate have
been observed in any of investigated materials. Intermediate amounts of DNA were found only
in tissues presumably undergoing an interphase synthesis of DNA preceding endoreduplication.
Mitosis and cytokinesis are the major cytological events associated with progression
through the cell cycle. A common way for a nucleus to differentiate in development is by
undergoing many rounds of DNA synthesis without an accompanied cell division, i.e., a process
of nuclear polyploidization. Nuclear polyploidy is commonly encountered in eukaryotic tissues,
and its occurrence in plants and animals is well reviewed (Leitch, 2000). Nuclear polyploidy can
hugely increase the DNA content of a cell. Nagl (1978) reviewed maximum levels of polyploidy
and reported values as high as 8,192C in the suspensor cells of the plant Phaseolus coccineus
and 524,288C (i.e., 219C) in silk glands of the insect Bombyx mori. There are several
mechanisms that give rise to polyploidy. The first is endoreduplication, where genomes replicate
without cell division. In many organisms the chromatids remain tightly associated, forming
polytene chromosomes, and these have been found in a diverse range of tissues and taxa in
vascular plants, they regularly occur in synergid and tapetal cells (Hasitschka-Jenske, 1957;
Leitch, 2000). Another mechanism is endomitosis, where replicated chromosomes condense as if
entering mitosis but then do not segregate; instead, they remain together within a single nucleus.
Polyploidy may also be accompanied by genome reorganization via DNA splicing, as occurs in
the ciliate Oxytricha and by polytene chromosome breakage, as occurs in some cells of
Drosophila melanogaster (Leitch, 2000).
The results of the present study have revealed that the nucleus of the synergid cell in A.
atroviolaceum undergoes endoreduplication up to 8C. The increase in DNA amount occurs by
DNA doubling as it is usual in polyteny (1C, 2C, 4C, 8C; Leitch, 2000). The volume of the
nucleus increases considerably and changes in patterns of nuclear morphology. Thereafter, the
nucleus becomes lobed and fragmented, and amount and sizes of dense chromatin masses
increase considerably. These changes have to be indicative that nuclear polyploidy is involved in
cell development. Furthermore, once DNA replication is complete, the nucleus can continue to
change and undergo further differentiation.
It is generally assumed that polyploidy occurs to amplify genes without the energetically
demanding process of cell division (Leitch, 2000). Thus, many secretory cell types are polyploid.
A gene amplification-expression argument to explain polyploidy does not explain the several
types of polyploidy found in a single organism or why polyploid nuclei themselves undergo
developmental changes. It is possible that the polyploidization process in the synergid of genus
Allium increases metabolic activity of the cell, which might be involved in the nutrition of the
embryo in early stages of endosperm development.
There are other potential roles for polyploidy. In polyploid nuclei, chromosome arms
might be able to associate in a manner that is impossible without multiple copies of each
chromosome. Such interactions might be important for chromosome trans-sensing (Leitch,
2000). Alternatively, nuclear polyploidy could amplify the genetic component of a cell which is
destined to be long-lived and perhaps vulnerable to mutation. In so doing, nuclear polyploidy
might extend the duration of cell viability. The full significance of polyploidy is unknown.
However, it could play roles in gene amplification, genome restructuring, chromosome
interactions, and cell longevity. It is not excluded that the polyploidization process in the
persistent synergid in genus Allium determines longer functioning of this cell degenerating
usually soon after fertilization, i.e. after completion of the main task of this type of cell. However
the trophic function might be good reason for organism to maintain a structure participating in
crucial developmental process, such as supply the embryo with nutrients.
One interesting point is the negative correlation between genome size and extent of
endopolyploidization in animals and seed plants Nagl (1978). It is assumed that occurrence of
endopolyploidy in seed plants is determined genetically. Moreover, endopolyploidization
behaviour seems to be typical for some seed plant families (Tschermak-Woess 1950; Nagl
1978). In some taxonomic groups many species contain highly endopolyploidized tissues
whereas this is not the case in other taxa. The degree of endopolyploidization differs between the
different organs of a species. Additionally, endopolyploidization is related to life cycle.
Endopolyploidy is very frequently found in annual and biennial species and also in some
perennial herbs whereas it seems to be absent in wooden species. At the same time, it was
assumed that a minimum amount of nuclear DNA in species with small genomes is frequently
realized by endoreduplication, to be required to maintain the regulatory and functional state of
certain cells (Kasahara et al., 2005). All these data are indicative of importance of
endopolyploidization for maintenance of successful life conditions for living organisms. As well,
it seems that some taxonomic groups are more depending on the increase of genome size than
Variation in DNA content is common for vegetative parts of different species of genus
Allium. It was estimated (Ohri et al., 1998) that 4C DNA amounts of 86 species from genus
Allium show a 8.35-fold difference ranging from 35.60 pg (A. ledebourianum, 2n = 16) to 297.13
pg (A. validum, 2n = 56). At diploid level the difference was 3.57-fold between A.
ledebourianum (35.60 pg) and A. ursinum (127.14 pg). Strong variation in genome size has been
obtained for 28 species and altogether 57 accessions or cultivars by different authors (Baranyi,
Greilhuber, 1999). These results have shown that a significant loss and/or gain of DNA have to
occur during evolution of these taxa. Therefore, it should be expected that the genome size
variation might occur within different cell types and at different developmental stages in the
species of this genus.
We thank Dr. G. Kvinikhidze for providing kind help to conduct cytophotometric investigation.
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Fig. 1. A – Ovule and embryo sac of Allium fistulosum, x200; B – Embryo sac with giant
synergid in the ovule of A. cepa, x 250; C – Ovule of A. rotundum, x 180; D – Giant synergid,
egg cell and polar nuclei before fertilization in A. cepa, x 390; E – Giant synergid at the moment
of fertilization in A. atroviolaceum, x 410; F – Giant synergid and nuclear endosperm in A.
fistulosum, x380; G – Condensed chromatin in the polyploid nucleus of hypertrophic synergid in
A. atroviolaceum, x 550; Structure of nucleus of the giant synergid, degenerating synergid and
egg cell in A. fistulosum before fertilization, x 550; I- Proliferation of both synergids in A. cepa,
x 510. EG- egg cell; EN – endospem nucleus; ES – embryo sac; P – polar nucleus; S – synergid.
Fig. 2. DNA amount (C) in conditional optical units and number of nuclei measured in the giant
synergid of A. atroviolaceum at different stages of development. The DNA amount, 2C, 4C, 6C
and 8C corresponding to the optical units are shown at the top of the graphic together with the
stages of development: EMS – embryo sac maturation, F – fertilization, EN – nuclear endosperm
formation, EM – embryogenesis.
Number of nuclei
30 60 90 120 150 180 210 240 270 300 330 360 390 420 450 480 510 540
2C ESM4C F
6C EN8C EM