ONTOGENETIC DEVELOPMENT OF SPINOUS EXINE IN HIBISCUS SYRIACUS (MALVACEAE)

Pollen development in Hibiscus syriacus L. (Malvaceae) was studied with light (LM), scanning (SEM) and transmission (TEM) electron microscopes, with special attention to the formation of extremely long spines of the pollen grains. At the early tetrad stage, probacules are initiated directly on the plasma membrane and grow in coincidence with the height of primexine matrix within a callosic wall. Subsequently, a pretectum appears at the top of the probacules and then a foot layer is formed by accumulation of white line centered lamellations. Before dissolution of the callosic wall, a reticulate patterned pretectum is established around the microspores. There is not, however, any morphological indication on the initiation of the spines during the tetrad period within a callosic wall. It is after dissolution of the callosic wall that the spines of exine begin to form by the apposition of lamellated sheets. The lamellated sheets show a concentric configuration around the developing supratectal spines. The mature pollen grain is spheroidal, polycolporate, 160–170 μm in diameter, with supratectal spines 20–25 μm long. The supratectal spines of Hibiscus pollen are not homologous with the other exinous protrusions which are determined within the callosic wall during tetrad stage.     

Tetrad pollen formation in quartet mutants of Arabidopsis thaliana is associated with persistence of pectic polysaccharides of the pollen mother cell wall

The quartet (qrt) mutants of Arabidopsis thaliana produce tetrad pollen in which microspores fail to separate during pollen development. Because the amount of callose deposition between microspores is correlated with tetrad pollen formation in other species, and because pectin is implicated as playing a role in cell adhesion, these cell-wall components in wild-type and mutant anthers were visualized by immunofluorescence microscopy at different stages of microsporogenesis. In wild-type, callose was detected around the pollen mother cell at the onset of meiosis and around the microspores during the tetrad stage. Microspores were released into the anther locule at the stage where callose was no longer detected. Deposition and degradation of callose during tetrad pollen formation in qrt1 and qrt2 mutants were indistinguishable from those in wild-type. Enzymatic removal of callose from wild-type microspores at the tetrad stage did not release the microspores, suggesting that callose removal is not sufficient to disperse the microspores in wild-type. Pectic components were detected in the primary wall of the pollen mother cell. This wall surrounded the callosic wall around the pollen mother cell and the microspores during the tetrad stage. In wild-type, pectic components of this wall were no longer detectable at the time of microspore release. However, in qrt1 and qrt2 mutants, pectic components of this wall persisted after callose degradation. This result suggests that failure of pectin degradation in the pollen mother cell wall is associated with tetrad pollen formation in qrt mutants, and indicates that QRT1 and QRT2 may be required for cell type-specific pectin degradation to separate microspores.     

LP28, a lily pollen-specific LEA-like protein, is located in the callosic cell wall during male gametogenesis

. LP28, a pollen-specific LEA-like protein identified in Lilium longiflorum purportedly related to the desiccation tolerance of pollen, was localized during male gametogenesis using immuno-electron microscopy. At premeiotic interphase, LP28 label is absent from the microsporocyte. LP28 label was first detected in the cell wall of the microsporocyte at meiotic prophase I. LP28 gradually increased as the cell wall thickened. In the dyad, after the first meiotic division, LP28 label also appeared in the septum. In the tetrad, after the second meiotic division, LP28 was detected throughout the cell wall, including the septa. Immunolabeling of callose during meiosis indicated that the appearance and localization of LP28 was very similar to that of callose. After the microspores were released from the tetrad by digesting the callosic cell wall, LP28 was not found in the microspores. In bicellular pollen, just after microspore mitosis, LP28 appeared in the generative cell wall, which also consisted of callose. After pollen germination, LP28 also accumulated in the callosic layer of the elongated pollen tube wall and the callose plug. Thus, LP28 colocalized with the callosic cell wall during male gametogenesis. The possible role of LP28 with respect to wall formation during meiosis and pollen development is discussed.     

Ultrastructural studies on plastids of generative and vegetative cells inLiliaceae 3. Plastid distribution during the pollen development inGasteria verrucosa (Mill.) Duval

Summary This paper describes the development of pollen grains ofGasteria verrucosa from the late microspore to the mature two-cellular pollen grain. Ultrastructural changes and the distribution of plastids as a result of the first pollen mitosis have been investigated using light and electron microscopy. The microspores as well as the generative and the vegetative cell contain mitochondria and other cytoplasmic organelles during all of the observed developmental stages. In contrast, the generative cell and the vegetative cell show a different plastid content. Plastids are randomly distributed within the microspores before pollen mitosis. During the prophase of the first pollen mitosis the plastids become clustered at the proximal pole of the microspore. The dividing nucleus of the microspore is located at the distal pole of the microspore. Therefore, the plastids are not equally distributed into both the generative and the vegetative cell. The possible reasons for the polarization of plastids within the microspore are briefly discussed. The lack of plastids in the generative cell causes a maternal inheritance of plastids inGasteria verrucosa.     

Embryological studies of Oxalis debilis Kunth

The study of pollen and female gametophyte development of Oxalis debilis was carried out to elucidate the reasons for the absence of seed production in this species. The formation of an incomplete callosic wall separating the microspores of the tetrad was observed; therefore, cytoplasm connections were present between the microspores. Microspores of different sizes and with different grades of vacuolation were observed in the mature anther. Only few microspores divide mitotically and form the generative and vegetative cell. The cytoplasm of the vegetative cell may accumulate different substances in reserve. Pollen grains are 8–12?colpate, and the morphology and electron density of the exine varies. Because flowers in full anthesis have all ovules at the megasporocyte stage with the nucellar epidermis intact, fecundation could not occur. The production of only a few viable pollen grains and the absence of successful megagametophyte development prevent fertilization and account for the absence of fruits and seeds formation.     

Reproduction in seagrasses: pollen wall morphogenesis in Amphibolis antarctica and wall structure in filiform grains

The pre–meiotic anther of the marine angiosperm Amphibolis antarctica contains microsporocytes and sterile cells. The microsporocytes divide conventionally to produce tetrads, but the sterile cells degenerate and contribute to the future pe–riplasmodium. Each tetrad of young microspores is contained within a vesicle defined by a membrane. After release from the tetrad, the microspores increase in length and rapidly become filiform. The microspore nucleus soon divides and partitioning of the cytoplasm delimits the generative cell from the vegetative cell of the binucleate pollen grain. The division and the early pollen growth occurs while the grains are segregated within vesicles in the periplasmodium. These compartments, established at microspore release, remain structurally intact throughout the vacuolate period of pollen development, when pollen wall assembly begins. This process is initiated as particles migrate from the inner face of the vesicle membrane into the lumen of the vesicle and microfibrillar elements form between adjacent particles. The particles and microfibrils form a loose, three–dimensional network. The vesicle membrane then disappears and the binuclate grains become immersed in the tapetal residuum. Additional wall components are now deposited upon the primary fibrillar stratum. Short lamellae, resembling fragments of membrane, frequently associated with electron–opaque globuli, are found intermixed with the surface microfibrils. Apparently, granular material originating in the degenerating periplasmodium may be the precursor of the globuli, and contact with the lamellae brings about an alteration in state. At this stage the pollen wall is resolved as two distinct fibrillar strata and the lamellae and globuli are incorporated as inclusions into the superficial zone of the outer stratum. The mature pollen wall exhibits faint stratification and the presence of the subsurface inclusions is readily demonstrated in germinating grains by section staining with phosphotungstic acid. The pollen wall in A. antarctica is compared with that in filiform grains of other seagrasses.     

Microspore development inBrassica napus and the effect of high temperature on division in vivo and in vitro

Summary Brassica napus cv. Topas microspores isolated and cultured near the first pollen mitosis and subjected to a heat treatment develop into haploid embryos at a frequency of about 20%. In order to obtain a greater understanding of the induction process and embryogenesis, transmission electron microscopy was used to study the development of pollen from the mid-uninucleate to the bicellular microspore stage. The effect of 24 h of high temperature (32.5 °C) on microspore development was examined by heat treating microspore cultures or entire plants. Mid-uninucleate microspores contained small vacuoles. Late-uninucleate vacuolate microspores contained a large vacuole. The large vacuole of the vacuolate stage was fragmented into numerous small vacuoles in the late-uninucleate stage. The late-uninucleate stage contained an increased number of ribosomes, a pollen coat covering the exine and a laterally positioned nucleus. Prior to the first pollen mitosis the nucleus of the lateuninucleate microspore appeared to be appressed to the plasma membrane; numerous perinuclear microtubules were observed. Microspores developing into pollen divided asymmetrically to form a large vegetative cell with amyloplasts and a small generative cell without plastids. The cells were separated by a lens-shaped cell wall which later diminished. At the late-bicellular stage the generative cell was observed within the vegetative cell. Starch and lipid reserves were present in the vegetative cell and the rough endoplasmic reticulum and Golgi were abundant. The microspore isolation procedure removed the pollen coat, but did not redistribute or alter the morphology of the organelles. Microspores cultured at 25 °C for 24 h resembled late-bicellular microspores except more starch and a thicker intine were present. A more equal division of microspores occurred during the 24 h heat treatment (32.5 °C) of the entire plant or of cultures. A planar wall separated the cells of the bicellular microspores. Both daughter cells contained plastids and the nuclei were of similar size. Cultured embryogenie microspores contained electron-dense deposits at the plasma membrane/cell wall interface, vesicle-like structures in the cell walls and organelle-free regions in the cytoplasm. The results are related to embryogenesis and a possible mechanism of induction is discussed.Abbreviations B binucleate - LU late uninucleate - LUV late uninucleate vacuolate - M mitotic - MU mid-uninucleate - RER rough endoplasmic reticulum - TEM transmission electron micrograph     

DEVELOPMENT OF WHEAT (TRITICUM AESTIVUM) POLLEN. II. HISTOCHEMICAL DIFFERENTIATION OF WALL AND UBISCH BODIES DURING DEVELOPMENT

The histochemistry of different developmental stages of the pollen wall, aperture, and Ubisch bodies of Triticum aestivum is examined with light and transmission electron microscopy. Various parts of the callosic envelope of the tetrad spores stain differentially. At the late tetrad stage, the probacules and the coat of pro-Ubisch bodies are densely stained for acidic polysaccharides, protein, and neutral polysaccharides. The protectum and the core of pro-Ubisch bodies are moderately stained. Upon release of microspores from the callosic cell envelope, the stainability for acidic polysaccharides increases in the exine and in the wall of Ubisch bodies, becoming very intense in the wall of mature pollen grains and Ubisch bodies. The stainability for neutral polysaccharides is decreased in the mature pollen wall and in the Ubisch bodies, while the stainability for protein increases. The results also indicate the probability of the presence of unsaturated lipids and the absence of free aldehydes in the pollen wall and Ubisch bodies.     

The cytoskeleton and polarization during pollen development in Carex blanda (Cyperaceae)

Patterns of cytoskeletal organization during distinct polarizations that characterize pollen development in the sedge Carex blanda (Cyperaceae) were studied by correlated methods of immunohistochemistry and confocal and transmission electron microscopy. As is typical of the family Cyperaceae, Carex produces a unique pollen type known as a pseudomonad in which all four microspores of the tetrad are enclosed within the wall of a single pollen grain. Only one member of the tetrad is functional and the other three abort. The pseudomonads are precisely oriented in the locule with the functional microspore in the wide abaxial portion of the wedge-shaped cytoplasm adjacent to the tapetum, and the degenerative microspores are packed tightly in the pointed adaxial portion. A unique sequence of post-meiotic developmental events reflects both intracellular and intercellular polarity. Development includes: (1) random placement of tetrad nuclei in the coenocytic sporocyte after meiosis, (2) interrupted cytokinesis resulting in a tetrad of nuclei that migrates as a unit into the narrow adaxial tip, (3) completion of unequal cytokinesis and centering of the functional nucleus in the wide abaxial portion of the microsporocyte via a radial array of microtubules and microfilaments, (4) unequal mitosis resulting in a small generative cell at the proximal surface of the functional microspore (adjacent to the abortive microspores), and (5) recentering of the vegetative nucleus in the abaxial cytoplasm via a radial cytoskeletal array.     

Nuclear fusion in cultured microspores of barley

Fusion of the generative and vegetative nuclei physically separated by a wall has been observed in cultured microspores of barley. The generative cell appears to play an active role in fusion as it elongates toward the vegetative nucleus, becomes detached from the microspore wall, and finally completely encloses the vegetative nucleus. The generative cell wall disappears before nuclear fusion takes place. Since these events have been known to occur during pollen development in vivo, it is hypothesized that the occurrence of nuclear fusion in cultured microspores is the result of continued expression of the genes for gametophytic development.     

Development of the pollen grain wall in Canna

During the tetrad period spinules form on the Canna L. plasma membrane at intervals of 1–2 μm on a microspore surface of ca. 100 μm². The isolated spinules represent all that there is of a primexine-like nature. Immediately following loss of the callosic tetrad envelope a channeled, oncus-like zone forms on the plasma membrane over the entire microspore surface, elevating the spinules. The oncus-like zone becomes ca. 4 μm thick by microspore mitosis. Intine introduction during the pollen grain period coincides with substantial thinning of the oncus-like zone and pollen grain enlargement. In the final phases of maturation grains increase further in diameter and become packed with starch and lipoidal material. The oncus-like zone more than doubles in height necessitating a migration of the boundary of the oncus-like zone and intine. At pollen grain maturity the thin (ca. 200 nm) surface layer of the oncus-like zone appears to be compacted or filled in.     

Anatomical differences on development of fertile and sterile pollen grains of Prunus salicina Lindl.

Anatomical changes occurring during the microsporogenic development of P. salicina Lindl. were studied in male fertile and male sterile genotypes. Male fertile pollen grains showed three well determined pore regions, without ektexine. Intine was thick and surrounded the vegetative cell. Vegetative cells enclosed the generative cells; their cytoplasm was rich in plastids, abundant RER and active mitochondria. Development of sterile pollen was different from the meiosis step. Microspores did not show germination pores and ektexine was continuous around the whole grain. Pollen grains showed an atypical shape. The tapetum persisted after the tetrad stage and showed hypertrophy and vacuole development, resulting in abnormal microspore development. Only a few pollen grains and rudiments of collapsed microspores close to the anther wall were formed at anthesis.     

含笑花药发育的超微结构研究

对含笑花药发育中的超微结构变化进行观察,结果显示:(1)花粉发育中有三次液泡变化过程——第一次是小孢子母细胞在形成时内部出现了液泡,这可能与胼胝质壁的形成有关;第二次是在小孢子母细胞减数分裂之前,细胞内壁纤维素降解区域形成液泡,它的功能可能是消化原有的纤维素细胞壁;第三次是在小孢子液泡化时期,形成的大液泡将细胞核挤到边缘,产生极性。(2)含笑花粉在小孢子早期形成花粉外壁外层,花粉外壁内层在小孢子晚期形成,而花粉内壁是在二胞花粉早期形成;花粉成熟时,表面上沉积了绒毡层细胞的降解物而形成了花粉覆盖物。研究认为,含笑花粉原外壁的形成可能与母细胞胼胝质壁有关,而由绒毡层细胞提供的孢粉素物质按一定结构建成了花粉覆盖物。     

凤仙花花药发育中多糖和脂滴组织化学研究

以不同发育时期的凤仙花花药为实验材料,采用组织化学方法,对花药发育中的结构变化及多糖和脂滴物质分布进行观察。结果表明:(1)凤仙花的花药壁由6层细胞组成,包括1层表皮细胞,2层药室内壁细胞,2层中层细胞和1层绒毡层细胞。其中绒毡层细胞的形态不明显,很难与造孢细胞区分,且在小孢子母细胞时期退化。(2)在小孢子母细胞中出现了一些淀粉粒,但减数分裂后,早期小孢子中的淀粉粒消失,又出现了一些小的脂滴;随着花粉的发育,小孢子形成大液泡,晚期小孢子中的脂滴也消失;小孢子分裂形成二胞花粉后,营养细胞中的大液泡降解、消失,二胞花粉中又开始积累淀粉;接近开花时,成熟花粉中充满细胞质,其中包含了较多的淀粉粒和脂滴。(3)在凤仙花的花药发育中,绒毡层细胞很早退化,为小孢子母细胞和四分体小孢子提供了营养物质;其后的中层细胞退化则为后期花粉发育提供了营养物质。     

莴苣花粉发育过程中ATP酶的分布

对莴苣花粉发育过程中ATPase的分布特征做了研究。四分体早期的小孢子细胞质中开始出现ATPase反应颗粒。之后,小孢子在发育过程中,花粉内壁聚集大量体积较大的ATPase反应颗粒,并一直保持到花粉即将成熟。在小孢子发育晚期,在花粉萌发孔处和小孢子大液泡中也特异性地聚集了较多ATPase颗粒。二胞花粉刚形成的生殖细胞表面呈现大量的ATPase反应颗粒,当生殖细胞脱离花粉内壁移入营养细胞,ATPase反应颗粒基本消失。生殖细胞分化过程中生殖细胞的ATPase反应颗粒逐渐低于营养细胞中的。在成熟花粉中,精细胞中的ATPase反应颗粒比营养细胞中的少,且主要集中在细胞核中。结果显示花粉发育过程中ATPase的特异分布与花粉发育的一些生物学事件密切相关。     

Pollen ontogeny in Magnolia liliflora Desr.

Pollen ontogeny contributes significantly to the evolutionary analysis and the understanding of the reproductive biology of seed plants. Although much research on basal angiosperms is being carried out there are still many important features about which little is known in these taxa, such as the sporophytic structures related to pollen development and morphology. In this study, pollen development of Magnolia liliflora was analyzed by optical microscopy and transmission electron microscopy. The aim of this paper was to supply data that will help characterize basal angiosperms. Microsporogenesis is of the successive type, so that tetrads are decussate or isobilateral. The callosic walls form by the centripetal growth of furrows. The secretory tapetum develops orbicules, which start to form in the microspore tetrad stage. Pollen grains are shed at the bicellular stage. The exine wall has a granular infratectum. Ultrastructural changes observed in the cytoplasm of microspores and tapetal cells are related to the development of the pollen grain wall and orbicules. Centrifugal cell plates are more usual for the successive type of microsporogenesis. The presence of the successive type of microsporogenesis with callosic walls formed by the centripetal growth of furrows could reflect the fact that the successive type in Magnoliaceae is derived from the simultaneous type. The granular infratectum of the ectexine and the presence of orbicules could indicate that this species is one of the most evolved of the genus.     

POLLEN GRAIN DEVELOPMENT AND TAPETAL CHANGES IN STRELITZIA REGINAE (STRELITZIACEAE)

The proexine that forms within the callosic envelope before the end of the microspore tetrad period is thick (about 1 μm) and exceptionally complex. It has components equatable with tectum, columellae, and a nexine that includes lamellar zones. All these components persist in the exine although late in development they become difficult to recognize because this exine is reduced in thickness, apparently by stretching, to a maximum of 0.2 μm. Strelitzia is an example of an exine template, with receptors for sporopollenin, that is not maintained during development. The Strelitzia microspore surface changes from an exine like that on an interaperture sector to the channeled intinelike system common for the apertures of pollen grains. The exine on sterile grains gives what may be a rare view of a stabilized immature exine. The mature exine on viable pollen grains resembles this early exine only in the most impressionistic way. Tapetal cells go through at least one cycle of hyperactivity, dedifferentiation, mitosis, and then again hyperactivity before they finally decline.     

洋葱花粉发育过程中ATP酶的分布特征(简报)

在真核细胞中,除了线粒体和叶绿体ATPase的功能是合成ATP外,其余部位ATPase是水解ATP以获取生物能量的代谢酶,在生物体细胞内广泛存在。探索ATPase在细胞中的分布状态是研究细胞生理状态的一种重要手段。ATPase在细胞中的多少可反映出细胞当时的生活状态,这一特征已被初步用于探索小麦和水稻雄性不育的细胞生物学研究中,希望通过比较可育花药和不育花药中ATPase的分布差异寻找雄性不育的机理,发现     

芝麻核雄性不育系ms86-1微粉发生的细胞学观察

芝麻(Sesamum indicum)核雄性不育系ms86-1姊妹交后代表现为可育、部分不育(即微粉)及完全不育(简称不育)3种类型。不同育性类型的花药及花粉粒形态差异明显。Alexander染色实验显示微粉植株花粉粒外壁为蓝绿色, 内部为不均一洋红色, 与可育株及不育株花粉粒的染色特征均不相同。为探明芝麻微粉发生机理, 在电子显微镜下比较观察了可育、微粉、不育类型的小孢子发育过程。结果表明, 可育株小孢子母细胞减数分裂时期代谢旺盛, 胞质中出现大量脂质小球; 四分体时期绒毡层细胞开始降解, 单核小孢子时期开始出现乌氏体, 成熟花粉时期花粉囊腔内及花粉粒周围分布着大量乌氏体, 花粉粒外壁有11–13个棱状凸起, 表面存在大量基粒棒, 形成紧密的覆盖层。不育株小孢子发育异常显现于减数分裂时期, 此时胞质中无脂质小球出现, 细胞壁开始积累胼胝质; 四分体时期绒毡层细胞未见降解; 单核小孢子时期无乌氏体出现; 成熟花粉时期花粉囊腔中未发现正常的乌氏体, 存在大量空瘪的败育小孢子, 外壁积累胼胝质, 缺乏基粒棒。微粉株小孢子在减数分裂时期可见胞质内有大量脂质小球, 四分体时期部分绒毡层发生变形, 单核小孢子时期有部分绒毡层开始降解; 绒毡层细胞降解滞后为少量发育进程迟缓的小孢子提供了营养物质, 部分小孢子发育为正常花粉粒; 这些花粉粒比较饱满, 表面有少量颗粒状突起, 但未能形成覆盖层, 花粉囊腔中及小孢子周围存在少量的乌氏体。小孢子形成的育性类型与绒毡层降解是否正常有关。