苔藓植物孢子发生的研究进展

苔藓植物孢子发生的过程是一个复杂的形态建成的过程,在此过程中,孢子母细胞经过减数分裂的两次精确的核分裂以及细胞质分裂,形成单倍体的四分孢子,再经孢子壁的发育过程,形成成熟的孢子。本文重点介绍了苔藓植物孢子发生过程中细胞质裂片、质体及核的变化、微管系统及纺锤体、胞质分裂和孢子壁形成过程的特点及其研究进展。     

糜子的小孢子发育和雄配子体形成

1.小孢子四分体的排列方式为左右对称形、直列式和T形。2.花粉第一次右丝分裂前夕,部分细胞质定向集中并形成细胞质索。3.有丝分裂末期出现成膜体,而后形成分开营养细胞和生殖细胞的拱形壁。4.营养核移至萌发孔,拱形壁开始消失,生殖细胞经过变形变化并进入营养细胞的细胞质。当生殖细胞完成位移并和营养核紧密贴近后,它开始分裂。5.在生殖细胞的有丝分裂过程中,其纺锤体轴的方向不止一个;细胞质分裂是产生缢缩沟。6.由     

杜仲胚胎学的研究——Ⅱ.雄配子体的发育

杜仲(Eucommia ulmoides Oliv)小孢子母细胞减数分裂属同时型。小孢子阶段短暂,当细胞体积略增大,未形成液泡时,细胞核由中部移向边缘即进行第一次分裂。在分裂中期,多数纺锤体轴垂直于花粉壁,呈不对称形;少数平行于壁,其两极相似。分裂过程中细胞质内逐渐形成几个大液泡,并消耗贮藏淀粉。生殖细胞位于边缘时,与营养细胞间的拱形壁呈PAS正反应。随后当生殖细胞内移到营养细胞质内的过程中,液泡逐渐解体,贮藏物质重新累积,花粉体积增大。成熟花粉具三沟孔,二细胞型。花粉管单一无分枝,当生殖细胞在花粉管中分裂时,营养核由椭圆形变长,结构松散,并处于其近侧。二个精子一前一后相接近,营养核紧邻其前端,未见有在其后面的现象。     

温光敏核不育水稻N28S 无花粉败育的显微结构观察

采用石蜡切片和荧光显微技术观察了温光敏核不育水稻N28S 无花粉败育过程中的显微结构变化, 结果显示: N28S 的小孢子母细胞形成后细胞质变得稀薄, 一部分不能进行减数分裂, 一部分减数分裂阻滞在细线期或胞质分裂异常, 最终所有细胞液泡化解体消失。在此过程中, 还观察到小孢子母细胞在细线期胼胝质壁不产生或提早消失, 以及小孢子发育后期花药壁绒毡层的异常解体。认为N28S 的无花粉败育是由小孢子母细胞的细胞质异常引起的, 胼胝质壁和绒毡层的异常是结果而不是原因。     

温光敏核不育水稻N28S无花粉败育的显微结构观察

采用石蜡切片和荧光显微技术观察了温光敏核不育水稻N28S无花粉败育过程中的显微结构变化,结果显示：N28S的小孢子母细胞形成后细胞质变得稀薄,一部分不能进行减数分裂,一部分减数分裂阻滞在细线期或胞质分裂异常,最终所有细胞液泡化解体消失。在此过程中,还观察到小孢子母细胞在细线期胼胝质壁不产生或提早消失,以及小孢子发育后期花药壁绒毡层的异常解体。认为N28S的无花粉败育是由小孢子母细胞的细胞质异常引起的,胼胝质壁和绒毡层的异常是结果而不是原因。     

桔梗花粉母细胞减数分裂及雄性败育的细胞生理学研究

以桔梗不育系PA及其保持系PB为试验材料,采用石蜡切片和改良苯酚品红染色压片法对花粉母细胞减数分裂和雄配子体发育过程进行比较,探讨桔梗雄性不育系小孢子形成过程和败育发生的细胞生理学机理。结果表明:(1)桔梗保持系PB花粉母细胞减数分裂的细胞质分裂为同时型,同一花药减数分裂较同步;在中期Ⅰ和中期Ⅱ,少数细胞中可见赤道板外染色体;四分体以四面体为主,成熟花粉粒为二核花粉。(2)不育系PA花粉母细胞减数分裂后期Ⅰ开始出现异常,表现为细胞质形态改变,末期Ⅱ之后细胞质不能分裂,形成异常四分体,胼胝质壁不能溶解,四分体难以释放出游离小孢子而被降解,导致败育。(3)在发育过程中,桔梗不育系花蕾游离脯氨酸、可溶性蛋白含量低于保持系,而SOD活性、丙二醛含量均高于保持系。(4)桔梗不育系PA及其保持系PB花粉母细胞减数分裂和雄配子体发育过程存在明显差异,桔梗雄性败育过程大体可分为4个阶段,即后期Ⅰ细胞质异常、末期Ⅱ之后细胞质不能分裂、四分体难以释放游离小孢子、四分体被降解仅残留碎片。研究认为,桔梗不育花蕾(开花前)生长发育过程中,体内活性氧代谢紊乱、丙二醛积累及游离脯氨酸等"物质代谢损亏"可能是引起桔梗雄性败育的原因。     

白菜核雄性不育系可育和不育花药中Ca2+的分布

研究了白菜(Brassica campestris L. ssp.chinensis Makino)细胞核雄性不育系花药中Ca2 的分布特征.在可育花药发育过程中,减数分裂后花药壁细胞中钙颗粒明显增加.早期小孢子开始积累钙颗粒并特异性地附在小液泡膜上.小孢子分裂后,大液泡消失过程中又伴随着许多钙颗粒附在小液泡膜上,显示出Ca2 与花粉中液泡的形成和分解有关.在不育花药中,最早出现的钙颗粒异常分布是在小孢子母细胞的胼胝质壁中积累了较多的钙颗粒.然而,在小孢子细胞质中钙颗粒一直很少,也不形成大液泡,最后通过细胞质收缩的方式败育.这是首次发现Ca2 参与调控花药发育过程,其异常分布与花粉败育密切相关.     

核桃楸的胚胎学研究（Ⅰ）——小孢子发生及雄配子体发育

通过石蜡制片技术对小孢子的发生和雄配子体发育过程进行了系统的研究。结果表明：花药含有4个花粉囊;花粉囊壁包括表皮、纤维层、中层、绒毡层;花药壁发育属基本型,腺质绒毡层,中层和绒毡层在花粉发育过程中逐渐解体,成熟的花粉囊只保留表皮和纤维层。小孢子母细胞减数分裂中,细胞质分裂是同时型。四分体排列方式为四面体型,四分体解体产生单核花粉粒,成熟花粉为2-细胞型。     

玉竹雄配子的发育——着重阐明质体在生殖细胞和营养细胞中的分布和变化

玉竹(Polygonatum simizui Kitag)小孢子在分裂前,质体极性分布导致分裂后形成的生殖细胞不含质体,而营养细胞包含了小孢子中全部的质体。生殖细胞发育至成熟花粉时期,及在花粉管中分裂形成的两个精细胞中始终不含质体。虽然生殖细胞和精细胞中都存在线粒体,但细胞质中无DNA类核。玉竹雄性质体的遗传为单亲母本型。在雄配子体发育过程中,营养细胞中的质体发生明显的变化。在早期的营养细胞质中,造粉质体增殖和活跃地合成淀粉。后期,脂体增加而造粉质体消失。接近成熟时花粉富含油滴。对百合科的不同属植物质体被排除的机理及花粉中贮藏的淀粉与脂体的转变进行了讨论。     

烟草小孢子母细胞减数分裂过程中微管分布变化

应用间接免疫荧光标记技术和激光共聚焦扫描显微镜成像技术观察了烟草小孢子母细胞减数分裂过程中微管的分布变化。在减数分裂前期,小孢子母细胞中的微管较短,随机分散在细胞质中。在减数分裂中期,细胞质中微管形成纺锤体,控制染色体的分布。进入减数分裂I后期,部分纺锤体微管将两组染色体拉向两级。在减数分裂Ⅱ中期,细胞中的微管又形成两个纺锤体。在减数分裂Ⅱ后期,纺锤体微管解聚为微管蛋白分散在细胞质中。胞质分裂发生在四个细胞核形成之后,通过细胞核之间的质膜向内缢缩分隔四个细胞核,产生四个小孢子。     

Nuclear compartmentalization is abolished during fission yeast meiosis

In eukaryotic cells, the nuclear envelope partitions the nucleus from the cytoplasm. The fission yeast Schizosaccharomyces pombe undergoes closed mitosis in which the nuclear envelope persists rather than being broken down, as in higher eukaryotic cells. It is therefore assumed that nucleocytoplasmic transport continues during the cell cycle. Here we show that nuclear transport is, in fact, abolished specifically during anaphase of the second meiotic nuclear division. During that time, both nucleoplasmic and cytoplasmic proteins disperse throughout the cell, reminiscent of the open mitosis of higher eukaryotes, but the architecture of the S. pombe nuclear envelope itself persists. This functional alteration of the nucleocytoplasmic barrier is likely induced by spore wall formation, because ectopic induction of sporulation signaling leads to premature dispersion of nucleoplasmic proteins. A photobleaching assay demonstrated that nuclear envelope permeability increases abruptly at the onset of anaphase of the second meiotic division. The permeability was not altered when sporulation was inhibited by blocking the trafficking of forespore-membrane vesicles from the endoplasmic reticulum to the Golgi. The evidence indicates that yeast gametogenesis produces vesicle transport-mediated forespore membranes by inducing nuclear envelope permeabilization.     

ZYGOSPORE GERMINATION IN CHLAMYDOMONAS MONOICA (CHLOROPHYTA): TIMING AND PATTERN OF SECONDARY ZYGOSPORE WALL DEGRADATION IN RELATION TO CYTOPLASMIC EVENTS

The Chlamydomonas monoica Strehlow zygospore is a dormant heavily walled spore adapted to survive extreme environmental conditions. The zygospore wall is multilayered and includes an acetolysis-resistant component related to the general class of compounds referred to as sporopollenin. Germination of the zygospore requires induction and completion of nuclear meiotic divisions, cytokineses to produce the four vegetative progeny cells, and breakdown of the zygospore wall to allow progeny release. Analysis of zygospore wall breakdown by transmission electron microscopy of synchronously germinating zygospores revealed differences in the timing and nature of disintegration of the various wall layers. Breakdown of the outer trilamellar sheath occurred within 6 h after light induction, concomitant with the onset of prophase I. At the same time, stored lipid bodies were consumed and replaced by large cytoplasmic vacuoles. Degradation of the inner more massive wall layer was initiated several hours later at about the time of the second meiotic division. In areas beneath breaks in the trilamellar sheath, a fibrous electron opaque bridge of wall material was retained whereas degradation of the remainder of the inner layer progressed. Finally, disintegration of this bridge material, after the completion of the meiotic divisions and synthesis of progeny cell walls, resulted in the opening of large slits in the trilamellar sheath, allowing escape of the flagellated vegetative progeny. The chloroform resistance typical of mature zygospores was lost at approximately the same time that the initial breaks in the trilamellar sheath were detected but before disintegration of the inner wall layer(s).     

<Emphasis Type="Italic">SSP2</Emphasis>, a sporulation-specific gene necessary for outer spore wall assembly in the yeast<Emphasis Type="Italic"> Saccharomyces cerevisiae</Emphasis>

Sporulation in yeast consists of two highly coordinated processes. First, a diploid cell that is heterozygous at the mating-type locus undergoes meiosis, in which one round of DNA replication is followed by two rounds of nuclear division. Second, the meiotic products are packaged into spore cells that remain within the mother cell. A large number of genes are induced specifically during sporulation, and their products carry out different sporulation-specific events. Expression of these sporulation-specific genes is controlled by several regulators which function at different stages of the sporulation program, resulting in a cascade of gene expression following induction of meiosis. Here we describe one sporulation-specific gene, SSP2, which is induced midway through meiosis. Ssp2 shows significant homology to the predicted product of a hypothetical ORF in Candida albicans. Homozygous mutant ssp2 diploid cells fail to sporulate. In the mutant background, meiotic recombination and nuclear divisions remain normal; however, viability declines rapidly. Following meiosis, ssp2 cells form the prospore membrane, but fail to form the outer layer of the spore wall. The Ssp2 protein localizes to the spore wall after meiosis II. In addition, the ssp2 defect is also associated with delayed and reduced expression of late sporulation-specific genes. Our results suggest that SSP2 function is required after meiosis II and during spore wall formation.     

ULTRASTRUCTURE OF SPOROGENESIS IN THE MOSS,AMBLYSTEGIUM RIPARIUM I. MEIOSIS AND CYTOKINESIS

Emphasis is placed on three aspects of meiosis in the moss Amblystegium riparium (Hedw.) BSG: 1***) nature of the sporogenous layer; 2) prophasic microtubules and polarity; and 3) cleavage pattern. Spore tetrads develop while still encased by archesporial cell walls. The cellular nature of the sporogenous layer differs from the more usual occurrence of free sporocytes released into a common spore sac. Two important events mark the establishment of sporocyte polarity during meiotic prophase: 1) migration of the four plastids to the distal tetrad poles (telophase II poles); and 2) ingrowth of the sporocyte wall in eventual cleavage planes between the tetrad poles. An extensive, plastid-based microtubule system is associated with organelle migration during the establishment of sporocyte polarity in meiotic prophase. Disruption of the nuclear envelope in prometaphase I occurs at sites opposite the four plastids where microtubules extend from plastid envelope to nuclear envelope. Formation of a cell plate following the first meiotic division results in a dyad, whereas in many mosses meiosis is completed in the undivided sporocyte and is followed by simultaneous cleavage into a spore tetrad. Spore cleavage is accomplished by vesicular coalescence resulting in septa that coincide with the prophasic wall ingrowths.     

The fission yeast spore is coated by a proteinaceous surface layer comprising mainly Isp3

The spore is a dormant cell that is resistant to various environmental stresses. As compared with the vegetative cell wall, the spore wall has a more extensive structure that confers resistance on spores. In the fission yeast Schizosaccharomyces pombe, the polysaccharides glucan and chitosan are major components of the spore wall; however, the structure of the spore surface remains unknown. We identify the spore coat protein Isp3/Meu4. The isp3 disruptant is viable and executes meiotic nuclear divisions as efficiently as the wild type, but isp3∆ spores show decreased tolerance to heat, digestive enzymes, and ethanol. Electron microscopy shows that an electron-dense layer is formed at the outermost region of the wild-type spore wall. This layer is not observed in isp3∆ spores. Furthermore, Isp3 is abundantly detected in this layer by immunoelectron microscopy. Thus Isp3 constitutes the spore coat, thereby conferring resistance to various environmental stresses.     

SPORE GERMINATION AND DEVELOPMENT OF THE YOUNG GAMETOPHYTE OF THE OSTRICH FERN (MATTEUCCIA STRUTHIOPTERIS)

Light is required for the germination of spores of Matteuccia struthiopteris. Histochemical studies show that dormant spores contain no starch, but have an abundance of storage protein granules. Starch accumulates in the numerous chloroplasts of the spore on exposure to light and becomes gradually more extensive. Protein granules disappear as germination progresses. Following this, the centrally located nucleus migrates toward the proximal spore face. Concomitant with the nuclear migration, an increase of cytoplasmic RNA surrounding the nucleus occurs. An equal nuclear division and unequal cell division give rise to a 2-celled gametophyte consisting of a large prothallial cell and smaller rhizoidal cell. A new peripheral wall forms around the entire protoplast at the time of nuclear migration, while a transverse wall forms after nuclear division. The rhizoid emerges through the split raphe along the proximal spore face; it is rich in cytoplasmic RNA but contains very few chloroplasts and little starch. Electron microscopy of the 2-celled stage revealed a greater concentration of mitochondria, Golgi bodies, and a more extensive endoplasmic reticulum in the rhizoid than was found in the prothallial cell, which, however, was far richer in chloroplasts and lipid bodies. As the rhizoid elongates and becomes more vacuolated, cytoplasmic RNA decreases as cytoplasmic protein increases. The rhizoid undergoes no cell divisions, while the prothallial cell retains the potential for further cell division. The possible significance of the distribution of storage products, cell organelles, and other cell components were considered in relation to the non-equational cell division and differentiation of the 2 cells.     

Sporogenesis in Bryophytes: Patterns and Diversity in Meiosis

Meiosis in bryophytes retains unusual features that provide clues to the innovation of sporogenesis in early land plants. Sporocytes are typically quadrilobed before nuclear division and the meiotic spindle is quadripolar with poles in the four future spore domains. Whereas seed plants consistently have anastral spindles arising from γ-tubulin in the perinuclear area, bryophytes have spindles organized at POs, plastids, or nuclear envelope. All of these MTOCs are significantly different from centrosomes of the algal ancestors. Mosses and hornworts have quadrilobed sporocytes with meiotic spindles organized at plastids. Meiosis in liverworts is extremely varied. Sporocytes of Jungermanniopsida are deeply quadrilobed and have microtubule bands marking division planes prior to cytoplasmic shaping. Spindles are organized at POs or nuclear envelope. Sporocytes of Marchantiopsida are quadrilobed to apolar with spindles organized at plastids, POs, or nuclear envelope. Pre-meiotic bands have been reported in only one marchantiod, the early divergent Blasia. An atlas of cytological data on 13 liverworts, 3 mosses and 2 hornworts is presented and analyzed.     

End3p-mediated endocytosis is required for spore wall formation in Saccharomyces cerevisiae

During sporulation in Saccharomyces cerevisiae, vesicles transported to the vicinity of spindle pole bodies are fused to each other to generate bilayered prospore membranes (PSMs). PSMs encapsulate the haploid nuclei that arise from the meiotic divisions and serve as platforms for spore wall deposition. Membrane trafficking plays an important role in supplying vesicles for these processes. The endocytosis-deficient mutant, end3Delta, sporulated poorly and the spores produced lost resistance to ether vapor, suggesting that END3-mediated endocytosis is important for sporulation. End3p-GFP localized to cell and spore peripheries in vegetative and sporulating cells and colocalized with actin structures. Correspondingly, the actin cytoskeleton appeared aberrant during sporulation in end3Delta. Analysis of meiosis in end3Delta mutants revealed that the meiotic divisions occurred with wild-type kinetics. Furthermore, PSMs were assembled normally. However, the levels of proteins required for spore wall synthesis and components of the spore wall layers at spores were reduced, indicating that end3Delta mutants are defective in spore wall synthesis. Thus, END3-mediated endocytosis is important for spore wall formation. Additionally, cytological analyses suggest that trafficking between the plasma membrane and PSMs is important earlier during sporulation.     

Cytological and genetic studies of the life cycle of Saccharomycopsis lipolytica

Summary In the alkane yeast Saccharomycopsis lipolytica (formerly: Candida lipolytica) the variability in the ascospore number is caused by the absence of a correlation between the meiotic divisions and spore wall formation. In four spored yeasts, after meiosis II, a spore wall is formed around each of the four nuclei produced by meiosis II. However, in the most frequently occurring two spored asci of S. lipolytica, the two nuclei are already enveloped by the spore wall after meiosis I due to a delay of meiosis II. This division takes place within the spore during the maturation of the ascus. In this case germination of the binucleate ascospore is not preceded by a mitosis. It follows that the cells of the new haploid clones are mononucleate. In the three spored asci, which occur rarely, only one nucleus is surrounded by a spore wall after meiosis I; the other nucleus undergoes meosis II before the onset of spore wall formation. The result is one binucleate and two mononucleate spores. In the one spored asci the two meiotic divisions occur within the young ascospore, i.e. spore wall formation starts immediately after development of the ascus. These cytological observations were substantiated by genetic data, which in addition confirmed the prediction that binucleate spores may be heterokaryotic. This occurs when there is a postreduction of at least one of the genes by which the parents of the cross differ. This also explains the high frequency of prototrophs in the progeny on non-allelic auxotrophs since random spore isolates are made without distinguishing between mono-and binucleate spores. The possibility of analysing offspring of binucleate spores by tetrad analysis is discussed. These findings enable us to understand the life cycle of S. lipolytica in detail and we are now in a position to start concerted breeding for strain improvement especially with respect to single cell protein production.