Ultrastructural Observation on the Intra- and Intercellular Microtrabecular Network of the Pollen Mother Cells in Onion (Allium cepa)

Using DGD embedment-free electron microscopy, ultrastructural observation on the intra- and intercellular microtrabecular network (MN) of the pollen mother cells (PMC) of the whole meiotic prophase Ⅰ in onion ( Allium cepa L.) was performed. Complex nuclear MN was observed in the nucleus of PMCs, spreading throughout the nuclear region. The nucleolus and chromosomes were connected with the MN filament network. The uniformity of nuclear MN changed with the development of the PMCs. A lamina-like structure surrounded the nucleus and joined the MN in nucleus and in cytoplasm, but it disassembled at the end of prophase Ⅰ. There was also a complex cytoplasmic MN in PMCs, without obvious variation during the prophase Ⅰ. Furthermore, MN in cytoplasmic connections (plasmodesmata and cytoplasmic channels) was noticed to link the frameworks in two neighboring PMCs into one entity. Cytomixis was observed at synizesis of prophase Ⅰ in this experiment, and MN in cytoplasm and in nucleus was noticed to distribute in these granules which migrated from one PMC into its neighboring cell. At this time the nucleus moved aside from center of the PMC, but the rest of the cell was still fulfilled with MN filaments. The relationships of nuclear MN with nucleolus and chromosomes, lamina with nucleus, as well as intra- and intercellular MN with cytomixis are discussed in this paper.     

百合花粉母细胞核骨架的超微结构观察

参照动物细胞核骨架的研究方法,用整装电镜技术和DGD包埋-去包埋技术研究了选择性抽提的和完整的百合(Lilium davidii var. willmottiae (Wilson) Roffill)花粉母细胞。结果表明,住减数分裂前期Ⅰ,百合花粉母细胞核内存在一个精细的非染色质纤维——核骨架。该网络由5～15nm的纤维交织而成,广泛地分布于细胞核内。这些核骨架有的分布于染色体间,有的分布于染色体周围,并与染色体和核仁相连。随着减数分裂时间的推移,染色体(质)间核骨架纤维逐渐减少,染色体(质)周围的核骨架纤维逐渐增多,并与染色体内部的纤维结构相连,表明核骨架一方面为染色体拓扑变化提供一个空间支架,另一方面也可能参与了染色体骨架的构建。     

东乡野生稻与栽培稻正反交种间杂种F1的雄配子发生与发育

该文以东乡野生稻与栽培稻(美国光壳稻P1003)的正反交种间杂种F₁(正交为光壳稻P1003×东乡野生稻; 反交为东乡野生稻×光壳稻P1003)为试材, 研究其各个减数分裂时期的染色体行为特征、染色体交换重组及雄配子发育特点。结果表明: 正反交杂种F₁花粉母细胞细胞核减数分裂的同步性较高, 细胞质为连续型胞质分裂类型。在细胞核分裂的过程中, 核仁在前期I到中期I逐渐消失。染色体在前期I到中期I逐渐收缩, 变得更加清晰可见。在终变期和中期I, 90.54%以上的花粉母细胞能形成12个二价体(含少数棒状二价体和8字型二价体), 部分花粉母细胞(正交9.46%, 反交7.65%)出现少量的单价体、三价体和四价体。后期I观察到1.27%–1.35%的花粉母细胞含有1至数条落后染色体。最终有92.6%–94.8%的小孢子能发育成形态正常、染色能力强的成熟花粉粒。另外, 在正交杂种F₁的粗线期中存在2个核仁, 而反交杂种F₁及其双亲的粗线期只有1个核仁。这些研究结果可为作物品种改良、种质资源创新以及种间亲缘关系研究提供细胞学证据。     

烟草花粉母细胞细胞融合过程中细胞骨架的超微结构观察

应用普通电镜和DGD去包埋技术 ,研究了烟草花粉母细胞中的细胞融合现象及细胞融合过程中细胞骨架的变化。观察发现 ,处于凝线期的花粉母细胞 ,其内含物 ,包括细胞器和染色质 ,主要通过胞质通道向相邻细胞发生转移。DGD去包埋观察发现 ,花粉母细胞中核骨架与细胞质内及胞间连丝和胞质通道内胞质骨架连接成一个整体。在整个细胞融合过程中 ,均有核骨架纤维与染色质相连。本文讨论了细胞骨架在细胞融合过程中的作用     

东乡野生稻与栽培稻正反交种间杂种F_1的雄配子发生与发育

该文以东乡野生稻与栽培稻（美国光壳稻P1003）的正反交种间杂种F1（正交为光壳稻P1003×东乡野生稻;反交为东乡野生稻×光壳稻P1003）为试材,研究其各个减数分裂时期的染色体行为特征、染色体交换重组及雄配子发育特点。结果表明：正反交杂种F1花粉母细胞细胞核减数分裂的同步性较高,细胞质为连续型胞质分裂类型。在细胞核分裂的过程中,核仁在前期I到中期I逐渐消失。染色体在前期I到中期I逐渐收缩,变得更加清晰可见。在终变期和中期I,90.54%以上的花粉母细胞能形成12个二价体（含少数棒状二价体和8字型二价体）,部分花粉母细胞（正交9.46%,反交7.65%）出现少量的单价体、三价体和四价体。后期I观察到1.27%–1.35%的花粉母细胞含有1至数条落后染色体。最终有92.6%–94.8%的小孢子能发育成形态正常、染色能力强的成熟花粉粒。另外,在正交杂种F1的粗线期中存在2个核仁,而反交杂种F1及其双亲的粗线期只有1个核仁。这些研究结果可为作物品种改良、种质资源创新以及种间亲缘关系研究提供细胞学证据。     

甜瓜花粉母细胞减数分裂及雄配子体的细胞学研究

以薄皮和厚皮类型甜瓜为试材,采用改良的染色体制片方法,系统观察了甜瓜花粉母细胞的减数分裂及雄配子体发育的过程,结果表明:(1)甜瓜细胞核减数分裂的同步性较高,细胞质是同时型分裂,在细胞核分裂的过程中,核仁在前期Ⅰ到中期Ⅰ逐渐消失,在前期Ⅱ再次出现,随后消失,染色体在前期Ⅰ到中期Ⅰ逐渐收缩,变得清晰,至末期Ⅰ变得模糊,在前期Ⅱ再次清晰;(2)2种类型甜瓜终变期的染色体构型都以环状二价体为主;(3)在后期Ⅱ,观察到染色体的垂直和平行2种分离方式;(4)在前期Ⅰ和前期Ⅱ,伽师瓜"形成了多个较小的核仁,呈现一定的特殊性;(5)雄配子体发育经历了单核期和双核期,最后形成了成熟的花粉粒.研究表明,薄皮和厚皮类型甜瓜减数分裂的染色体行为基本一致,没有明显差异;伽师瓜"的核仁数量表现特殊可能与其长期的生态适应性有关.     

Electron Microscopic Studies on the Silver-stained Nucleolar Cycle of Physarum polycephalum

The dynamic changes of nucleolar ultrastructure in the cell cycle of Physarum polycephalum Schw. were studied by an en bloc silver-staining method. The results showed that the nucleolus was large in size and situated in the center of the nucleus in late G2-phase, and the fibrillar centers, dense fibrillar components and granular components could be observed in the nucleolus. During prophase, the nucleolus moved towards the periphery of the nucleus and in late prophase disintegrated near the nuclear envelope. In metaphase, the disintegrated nucleolar components were dispersed in masses and located at the periphery of the chromosomal region of the nucleus. No specifically silver-stained area and argentophilic protein sheath were observed on the chromosomes, but there were some big dispersed silver particles within the chromosomes. During telophase the nucleolar components moved towards the two poles along with the chromosomes and co-existed with the decondensing chromatin in daughter nuclei. The nucleolar components then gradually converged with one another and separated from the chromatin. A big nucleolus was formed in the nucleus about 120 min after the completion of mitosis.     

多头绒泡菌银染核仁周期的电镜研究

以同步化培养的多头绒泡菌（Physarum poldycephalum Schw.)原生质团为材料,应用整体银染技术,电镜下研究了核仁在细胞周期中的超微结构变化。结果变化：核仁成熟时比较大,位于细胞核中央,核仁内可区分出纤维中心、密集纤维成分和颗粒成分等。前期时,核仁向边缘移动,前期末在近核膜处解体,解体的核仁物质主要呈团块状散开。中期时,解体的核仁物质位于细胞核中央染色体区域的周围,染色体上没有特异的银染区域,染色体周边也看不到银染的“鞘”状结构,但在染色体中可见一些散在的银染大颗粒。末期时,核仁物质与染色体一起到达两极,在子细胞核中与正在解集缩的染色质共存一起,以后核仁物质逐渐汇合并与染色质分开。大约在有丝分裂结束120min后,在细胞核中形成一候 中央位置的大核仁,结果提示,低等真核生物的核仁结构和周期变化与高等真核生物的不完全相同。     

POLLEN AND TAPETUM DEVELOPMENT IN DESMODIUM GLUTINOSUM AND D. ILLINOENSE (PAPILIONOIDEAE; LEGUMINOSAE)

Each of the four microsporangia has three or four wall layers, a uninucleate tapetum of various cell shapes with nuclei that remain in prophase, and 12-24 pollen mother cells (PMCs). A sterile transverse septum sometimes bisects the microsporangium. PMCs secrete callose but not uniformly, and contact among them continues through meiosis. Simultaneous cytokinesis by furrowing isolates each microspore in callose, which later disperses. The separated microspores become vacuolate, undergo mitosis to become pollen, and later become filled with food reserves. Endothecial wall thickening and tapetal dissolution occur after pollen engorgement. Calcium oxalate crystals form in tapetal cells during the sporogenous stage, reach maximum size during early meiosis, and remain prominent until tapetal dissolution.     

Early meiosis in Rhynchospora pubera L. (Cyperaceae) is marked by uncommon ultrastructural features

The family Cyperaceae has an unusual microsporogenesis in which tetrad formation does not occur. In addition, other cytological features are important, such as the occurrence of holokinetic chromosomes and post-reductional meiosis. We have examined the ultrastructural features of the pollen mother cell (PMC) of Rhynchospora pubera. Anthers of several sizes were analyzed using light and transmission electron microscopy. The PMC before meiosis presented a central nucleus and a regular profile of the nuclear envelope. During prophase I, the nucleus was in the abaxial region of the cell. This cellular polarization was accompanied by other marked ultrastructural features in the nuclear envelope. Morphological changes involved dilations of perinuclear cisterns and polarization of the nuclear pore complexes. The results show that polarization occurs in the initial phases of microsporogenesis in R. pubera, unlike other plant species.     

Cellular changes during the acquisition of embryogenic potential in isolated pollen grains ofNicotiana tabacum

Summary We used electron microscopical techniques to study ultrastructural changes during the acquisition of embryogenic competence in immature pollen grains ofNicotiana tabacum, isolated at the early- or mid-bicellular stage and cultured in vitro under starvation conditions. Cytoplasmic and nuclear changes during the starvation treatment are reported. Dedifferentiation of plastids, dilation of the wall of the generative cell, the appearance of a large vacuole, loss of nuclear pores in the vegetative nucleus, changes in chromatin and nucleolar structure, and a decrease in the size of the nucleolus were observed. We suggest that these events are the first step in the switch from generative to vegetative generation during pollen embryogenesis.     

Dynamics of antimonate-precipitated calcium and degeneration in unpollinated pearl millet synergids after maturity

Pearl millet synergids from unpollinated pistils at 1.5–2 and 2.5–3 days postmaturity (dpm) were examined using transmission electron microscopy following antimonate fixation to precipitate loosely-bound calcium (Ca). With increasing age of synergids, the gap above filiform apparatus (FA) and the coalesced vacuoles in midchalazal core extended and merged. The FAs became compressed and precipitates along their common wall were dispersed. The matrix material in numerous small chalazal vacuoles changed from dense to flocculent. Precipitates in vacuoles appeared mainly as clumps without or with a halo in the dense matrix and mostly finely distributed in the flocculent matrix. Eventually, vacuoles became free of both matrix and precipitates. Precipitates bound to nucleus, nucleolus, and micropylar cytoplasm increased initially, but then seemed to decrease, while the nucleus became disorganized and the nucleolus disappeared. Precipitates in the embryo sac wall and nucellar cells also increased initially, but then decreased. At very late stages, egg apparatus and ES lost structural integrity and lacked precipitates. Differences in degeneration and Ca levels of sister synergids were smaller at 1.5–2 dpm than at 2.5–3 dpm. A logical and correlative scheme of degenerative events and Ca distributional changes occurring in pearl millet synergids from maturity to 2.5–3 dpm is presented. The significance of results in pollen tube/sperm cell interactions with synergids and Ca is addressed.On Specific Cooperative Agreement 58-6612-8-002 with the Department of Biochemistry, University of Georgia, Athens, GA 30602, USA     

Cytochemical localization of cellulase activity in pollen mother cells of david lily during meiotic prophase I and its relation to secondary formation of plasmodesmata

Summary Cellulase activity was localized at the ultrastructural level in pollen mother cells (PMCs) of David lily [Lilium davidii var.willmottiae (Wilson) Roffill] at different stages of meiotic prophase I. The enzyme was observed to appear at the early leptotene stage and reached its highest level at the subsequent zygotene stage, and its subcellular distribution revealed by the presence of electron-dense deposits of reaction product was found to be restricted exclusively to the endoplasmic reticulum (ER), the vesicles derived from that, and the cell wall, especially at the sites of secondary plasmodesmata and cytoplasmic channels where the wall was being digested. Other cytoplasmic organelles, such as dictyosomes and Golgi vesicles, lacked such deposits of reaction product. After zygotene the enzyme activity decreased abruptly, and at the pachytene stage only very few deposits could be observed in the cell wall. Our results indicate that cellulase is synthesized on rough ER and secreted directly via the smooth ER and ER-derived vesicles into the cell wall by exocytosis, where it brings about local wall breakdown, leading to the secondary formation of plasmodesmata and cytoplasmic channels.     

CELL DIVISION IN COSMARIUM BOTRYTIS1

Cell division in Cosmarium is described. Premitotic cells are very dense; the semicells, previously appressed to one another, separate slightly during entry into prophase. This separation coincides with deposition of a girdle of new wall material around the isthmus, where the 2 semicells are joined. Micro-tubules, abundant around the isthmus wall during interphase, all disappear during prophase; meanwhile, other microtubules proliferate outside the nuclear envelope. By metaphase, the nucleolus has dispersed, although remnants of it persist. The nuclear envelope breaks up, but some membranes coat metaphase chromosomes. The spindle, while being typical, is somewhat multipolar; microtubules, usually associated with elements of endoplasmic reticulum, are oriented toward numerous regions in the poles. During anaphase and telophase, these spindle tubules become increasingly directed toward a few discrete foci, and they persist after telophase. Meanwhile, the septum grows from the girdle of wall material to bisect the cell. After cytokinesis, some microtubules reappear near the isthmus, but only adjacent to the older, non-expanding semicell wall. Cell expansion then takes place, during which the nucleus, ensheathed in a complex microtubular system, moves into the forming semicell. Later, the chloroplast follows the nucleus and its 2 pyrenoids elongate and divide. When semicell expansion is complete, the chloroplast cleaves adjacent to the isthmus. The nucleus, now apparently not associated with microtubules, concurrently moves back into the isthmus. Continuous deposition of primary wall material accompanies cell expansion. Wall materials are apparently secreted as aggregates (perhaps derived from the contents of vesicles) adjacent to the plasmalemma, whose fibrous components become increasingly oriented in the outer layers of the wall by stretching. Late in semicell formation, this deposition ceases and during further expansion, the semi-cell develops a pattern of warts and ridges. Secondary wall deposition under the primary wall then follows, matching this pattern of ornamentation. In addition, numerous plugs of amorphous material capped by specialized regions of the plasmalemma, traverse the entire thickness of the secondary wall which becomes further thickened at these particular sites. The amorphous plugs presumably are eroded away later to form the mucilage pores of the vegetative cells. The wall microtubules gradually become more symmetrically arrayed around the isthmus as this new secondary wall thickens. These observations are discussed in comparison with other work on morphogenesis in desmids.     

Cortical cytoskeletal ring in prophase II leads to correction of abnormalities of the first meiotic division and to meiotic restitution of pollen mother cell nucleus

The deviation of prophase cytoskeletal ring formation was determined during meiotic division in 50% of pollen mother cells (PMCs) in maize haploid No 1498 (Zea mays). At prophase in both meiotic divisions the cytoskeletal ring is formed in cortical region of cytoplasm instead of perinuclear. Sometimes formation of both perinuclear and cortical rings is observed in the same cell. It has been shown that in multinucleate PMCs the cortical ring leads to the consolidation of chromosomes into common spindle and to meiotic restitution.     

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。     

A comparative light and electron microscopic analysis of microspore and tapetum development in fertile and cytoplasmic male sterile radish

To gain further insight into the abortive stages and ultrastructural changes leading to pollen degeneration of a novel cytoplasmic male sterile radish 805A, we compared differences of cellular and subcellular structure of sterile anther with fertile anther by light and electron microscopy analysis. Two types of locule degeneration in sterile anther were detected, of which the time of degeneration occurred and completed was different. In type I, abnormality of pollen mother cells (PMCs) and tapetal cells, including condensation of cytoplasm and large vacuoles within tapetal cells, was shown at PMC stage. In type II, meiosis and early tetrad stage progressed normally except for large vacuoles that appeared in tapetal cells. Ultrastructural alterations of the cellular organization were observed in the type II locules, such as chromatin condensation at the periphery of the nucleus and degeneration of the karyotheca, compared with normal pollen development. The results suggested that the cytoplasmic male sterility anther degeneration was probably caused by dysfunctions of tapetum and vacuolation of tapetum, PMCs, and microspores. Thus, the identical factors, which induced CMS in the same cytoplasmic and nuclear genetic background, might affect development of tapetum and microspore at different stages during the cytoplasmic male sterile 805A anther development.     

CELL DIVISION IN SPIROGYRA. I. MITOSIS

Dividing cells of Spirogyra sp. were examined with both the light and electron microscopes. By preprophase many of the typical transverse wall micro-tubules disappeared while others were seen in the thickened cytoplasmic strands. Microtubules appeared in the polar cytoplasm at prophase and by prometaphase they penetrated the nucleus. They were attached to chromosomes at metaphase and early anaphase, and formed a sheath surrounding the spindle during anaphase; they were seen in the interzonal strands and cytoplasmic strands at telophase. The interphase nucleolus, containing 2 distinct zones and chromatinlike material, fragmented at prophase; at metaphase and anaphase nucleolar material coated the chromosomes, obscuring them by late anaphase. The chromosomes condensed in the nucleoplasm at prophase, moving into the nucleolus at prometaphase. The nuclear envelope was finally disrupted at anaphase during spindle elongation; at telophase membrane profiles coated the reforming nuclei. During anaphase and early telophase the interzonal region contained vacuoles, a few micro-tubules, and sometimes eliminated n ucleolar material; most small organelles, including swollen endoplasmic reticulum and tubular membranes, were concentrated in the polar cytoplasm. Quantitative and qualitative cytological observations strongly suggest movement of intact wall rnicrotubules to the spindle at preprophase and then back again at telophase.     

Cytokinesis in plant male meiosis

In somatic cell division, cytokinesis is the final step of the cell cycle and physically divides the mother cytoplasm into two daughter cells. In the meiotic cell division, however, pollen mother cells (PMCs) undergo two successive nuclear divisions without an intervening S-phase and consequently generate four haploid daughter nuclei out of one parental cell. In line with this, the physical separation of meiotic nuclei does not follow the conventional cytokinesis pathway, but instead is mediated by alternative processes, including polar-based phragmoplast outgrowth and RMA-mediated cell wall positioning. In this review, we outline the different cytological mechanisms of cell plate formation operating in different types of PMCs and additionally focus on some important features associated with male meiotic cytokinesis, including cytoskeletal dynamics and callose deposition. We also provide an up-to-date overview of the main molecular actors involved in PMC wall formation and additionally highlight some recent advances on the effect of cold stress on meiotic cytokinesis in plants.