Cytological features of oogenesis and their evolutionary significance in the fern Osmunda japonica.

The development of the egg and canal cells in the fern Osmunda japonica Thunb. was studied during oogenesis by transmission electron microscopy. The mature egg possesses no fertilization pore and no typical egg envelope. In addition, an extra wall formed around the canal cells during oogenesis and apparently blocked protoplasmic connections between the egg and the canal cells. The periodic acid Schiff (PAS) reaction revealed that the extra wall was most likely composed of polysaccharides. Maturation of the egg was accompanied by the formation of a separation cavity above the egg and by some changes in the morphology of the nucleus and cytoplasmic organelles. The chromatin of the nucleus becomes condensed and the upper surface of the nucleus becomes closely associated with the plasmalemma. Amyloplasts in the egg cytoplasm were numerous and conspicuous, with most in close proximity to the nucleus. Finally, the cytoplasm on one side of the egg became vesiculated and the overlying plasmalemma was easily disrupted. These cytological features of the egg and the canal cells during oogenesis in O. japonica are markedly different from those of the leptosporangiate ferns and suggest a significant evolutionary divergence in reproductive cellular features between Osmundaceae and leptosporangiate ferns.

Ultrastructural and cytochemical studies on oogenesis of the fern Pteridium aquilinum.

Egg development in Pteridium aquilinum var. latiusculum was studied using ultrastructural and cytochemical methods to examine structural features influencing fertilization in leptosporangiate ferns. Ultrastructural observations indicate a separation cavity is first formed above the egg during oogenesis with a pore region persistently connecting the egg and the ventral canal cell. The egg envelope is formed by deposition of amorphous materials in the separation cavity on the outer surface of plasmalemma. The egg envelope was not formed across the pore region; instead, a fertilization pore was formed. During oogenesis, the egg nucleus produced extensive evaginations containing osmiophilic bodies. Cytochemical experiments revealed that the egg envelope displays strong periodic acid-Schiff reaction indicative of polysaccharides, with negligible Sudan black B staining for lipids, suggesting that the egg envelope is composed principally of polysaccharides, and not lipids. The present manuscript provides new insights into egg structure and development of Pteridium, including discovery and characterization of the fertilization pore and observations on the chemical nature of the egg envelope, thus contributing to the understanding of the cytological mechanism of the sexual reproduction of ferns.

Formation of the fertilization pore during oogenesis of the fern Ceratopteris thalictroides.

The development of the fertilization pore during oogenesis of the fern Ceratopteris thalictroides was followed using transmission electron microscopy. The newly formed egg is appressed closely to the adjacent cells. There are well-developed plasmodesmata between the egg and the ventral canal cell, but none between the egg and the jacket cells of the archegonium. During maturation, a separation cavity is formed around the egg. However, a pore region persistently connects the egg and the ventral canal cell. The extra egg membrane is formed by deposition of sheets of endoplasmic reticulum (ER), but no ER is deposited on the inner surface of the pore region. Thus, a fertilization pore, covered by a layer of plasmalemma, is formed. The ventral canal cell undoubtedly participates the formation of the fertilization pore, probably by absorbing the sheets of ER beneath the pore region. The functional significance of the ventral canal cell in formation of the fertilization pore is discussed. The features of the mature egg include that abundant concentric membranes and osmiophilic vesicles occur in the cytoplasm of the mature egg. The initial, round nucleus of the egg eventually becomes cup-shaped. This investigation gives some new insights about the cells participating oogenesis in ferns.

Cytological events during zygote formation of the fern Ceratopteris thalictroides.

The cytological events, including nuclear fusion, digestion of male organelles and rebuilding of the plasmalemma and cell wall, during zygote formation of the fern Ceratopteris thalictroides (L.) Brongn. are described based on the observations of transmission electron microscopy. When the spermatozoid enters the egg and contacts the cytoplasm, the male chromatin relaxes continually. The microtubular ribbon (MTr) is separated from the male nucleus and then an envelope reappears around the male nucleus. During nuclear fusion, the egg nucleus becomes highly irregular and extends some nuclear protrusions. It is proposed that the protrusions fuse with the male nucleus actively. After nuclear fusion the irregular zygotic nucleus contracts gradually. It becomes spherical before the zygote divides. The male chromatin is identifiable as fibrous structure in the zygotic nucleus in the beginning, but it gradually becomes diffused completely. The male organelles, including the MTr, multilayered structure, flagella and the male mitochondria are finally digested in the zygotic cytoplasm. Finally a new plasmalemma and cell wall are formed outside the protoplast. The organelles in the zygote are rearranged, which produces a horizontal polarity zygote. The zygote divides with an oblique-vertical cell plate facing the apical notch of the gametophyte.

Ultrastructure of the mature egg and fertilization in the fern Ceratopteris thalictroides.

The ultrastructure of the mature egg and fertilization in the fern Ceratopteris thalictroides (L.) Brongn. were observed by transmission electron microscopy. The results revealed that the mature egg possesses an obvious egg membrane at the periphery of the egg. Furthermore, a fertilization pore was identified in the upper egg membrane of the mature egg. The structure of the pore is described for the first time. The fertilization experiment indicated that spermatozoids crowd into the cavity above the egg through the neck canal of the archegonium; however, only one of these can penetrate into the egg through the fertilization pore. Immediately on penetration of the spermatozoid, the egg begins to shrink. The volume of the fertilized egg decreases to almost one-half that of the unfertilized egg. As a result, the protoplasm of the fertilized egg becomes dense and opaque, which may lead to a situation where the organelles of both the egg and the fertilizing spermatozoid become indistinguishable. Simultaneously, abundant vesicles containing concentric membranes or opaque materials appear near the fertilization pore in the cytoplasm of the fertilized egg. These vesicles are considered to act as a barrier that prevents polyspermy. The present study provides a new insight into the ultrastructure of the mature egg and the cytological mechanism of fertilization in ferns.

Transcription factor AtMYB103 is required for anther development by regulating tapetum development, callose dissolution and exine formation in Arabidopsis.

Downregulation of the transcription factor AtMYB103 using transgenic technology results in early tapetal degeneration and pollen aberration during anther development in Arabidopsis thaliana. This paper describes the functional analysis of the AtMYB103 gene in three knock-out mutants. Two male sterile mutants, ms188-1 and ms188-2, were generated by ethyl-methane sulfonate (EMS) mutagenesis. A map-based cloning approach was used, and ms188 was mapped to a 95.8-kb region on chromosome 5 containing an AtMYB103 transcription factor. Sequence analysis revealed that ms188-1 had a pre-mature stop codon in the AtMYB103 coding region, whereas ms188-2 had a CCT-->CTT base-pair change in the first exon of AtMYB103, which resulted in the replacement of a proline by a leucine residue in the R2R3 domain. The third mutant, an AtMYB103 transposon-tagging line, also showed a male sterile phenotype. Allelism tests indicated that MS188 and AtMYB103 belong to the same locus. Cytological observation revealed defective tapetum development and altered callose dissolution in ms188 plants. Additionally, most of the microspores in mature anthers were degraded and surviving microspores lacked exine. AtMYB103 encoded an R2R3 MYB protein that is predominantly located in the nucleus. Real-time RT-PCR analysis indicated that the callase-related gene A6 was regulated by AtMYB103. Expression of the exine formation gene MS2 was not detected in mutant anthers. These results implicate that AtMYB103 plays an important role in tapetum development, callose dissolution and exine formation in A. thaliana anthers.