用于本科教学的间接免疫荧光法观察玉簪保卫细胞微管骨架

细胞骨架的观察是生命科学类专业本科生一个很重要的实验。目前,在细胞生物学实验教学中常用考马斯亮蓝染色法观察植物细胞骨架,但存在诸多不足。该研究选取洋葱、大叶黄杨、玉簪三种植物材料,利用间接免疫荧光法对微管进行观察比较。结果表明,三种植物中,玉簪作为一种常见的绿化观赏花卉植物,取材方便、撕片容易,其气孔保卫细胞微管骨架免疫荧光图像显示清晰,在低温处理后也可观察到微管解聚现象。因此,玉簪可作为本科生实验教学中利用免疫荧光法观察微管骨架的一种易得、观察效果好的实验材料。     

应用Steedman's wax切片法观察植物细胞微管骨架

微管骨架在植物发育过程中起重要作用.由于植物细胞的特殊性,与动物细胞相比植物微管骨架的研究遇到更多的困难.简略地介绍了曾被国内外学者应用的植物微管骨架的各种研究方法及其局限性.Steedman's wax是一种多脂蜡.它熔点低(35～37℃),具有与石蜡相同的切片性质,能够切成不同厚度的连续切片,适合深埋于器官内部的组织或细胞的免疫细胞化学研究.介绍了应用Steedman's wax切片法观察植物细胞微管骨架的一般程序和方法以及经过作者检验且切实可行的一些技术改进.     

冰冻切片法在植物微管骨架研究中的应用

介绍了冰冻切片法研究植物微管骨架的一般程序和技术上的一些改进,结果证明,改进的冰冻切片技术,可以对植物不同类型的细胞进行很好的标记。实验结果表明,甘蔗正在迅速伸长的幼叶分布的微管类型主要是与细胞伸长轴方向垂直的周质微管,幼叶基部尤其是第三幼叶基部分布的主要是与细胞伸长轴方向平行的周质微管。表明冰冻切片法在植物微管骨架的研究中具有广阔的应用前景。     

应用Steedman's wax 切片法观察植物细胞微管骨架

微管骨架在植物发育过程中起重要作用。由于植物细胞的特殊性,与动物细胞相比植物微管骨 架的研究遇到更多的困难。简略地介绍了曾被国内外学者应用的植物微管骨架的各种研究方法及其局限 性。Steedman's wax是一种多脂蜡。它熔点低(35~37℃),具有与石蜡相同的切片性质,能够切成不同厚 度的连续切片,适合深埋于器官内部的组织或细胞的免疫细胞化学研究。介绍了应用Steedman's wax 切 片法观察植物细胞微管骨架的一般程序和方法以及经过作者检验且切实可行的一些技术改进。     

植物微管骨架参与下胚轴伸长调节机制研究进展

微管作为细胞骨架的重要成员,在植物生长发育过程中起重要作用。下胚轴作为研究细胞伸长的模式系统之一,其伸长受到多种信号的调节。该文综述了微管骨架在响应环境和生长发育信号调节下胚轴伸长过程中的作用及机制,旨在帮助读者深入理解微管骨架响应上游信号在植物下胚轴伸长中的作用机理。     

植物微管和微丝骨架研究的进展

从以上叙述的资料中可以看出,近年来在植物微管蛋白的分离及其化学性质、微管的组织中心、微管的异质性、微丝的分布,以及微管和微丝骨架的功能及基因调节等方面的研究取得不少新的进展;特别是从植物中直接分离微管蛋白取得成功、以及微管蛋白异型、微管冷稳定性与植物抗寒性的关系及微丝分布广泛性等的发现,对植物细胞骨架的进一步研究具有重要意义。     

绿豆根尖细胞微管骨架有丝分裂时相发育变化的研究

提纯猪脑微管蛋白,制备兔抗微管蛋白抗血清,以此抗体与羊抗兔ｌｇＧ－ＦＩＴＣ因清,对绿豆根尖细胞进行间接免疫荧光标记和荧光显微镜检,得到了绿豆根尖细胞有丝分裂微管骨架周期发育变化的时相,如：早前期带,纺棰体微管,成膜体微管等,结果证明了双子叶植物具有与单子叶植物相似的细胞分裂微管周期时相,表明了微管架周期时相变化在高等植物中具有普遍性和共同变化的规律,讨论了微管骨架时相发育变化与染色有丝分裂行为的关     

中国科学家在植物细胞骨架研究领域取得突破性进展

微管是细胞骨架的重要组成部分,为真核细胞生命活动所必需。与其它生物体类似,微管不仅在植物生长发育中起重要作用,而且参与响应外界环境信号。近期,中国科学家在解析植物微管精准切割及微管骨架动态重构调控机制的研究中取得突破性进展。     

双子叶植物细胞周期微管骨架的时相变化的免疫荧光显微研究

本工作用两个循环的组装－去组装超速离心法提取和纯化猪脑微管蛋白,制备兔抗微管蛋白血清,并首次用免疫荧光显微术,对双子叶植物绿豆根尖细胞周期微管骨架各时相的排布进行了检测和分析。讨论了微管周期和染色体周期的细胞有丝分裂行为。     

慈姑根尖细胞微管骨架的免疫荧光观察

分别用考马斯亮蓝染色和间接免疫荧光标记,并运用荧光倒置显微镜和激光共聚焦显微镜,对慈姑根尖固定后酶解获得的去壁细胞和细胞团块以及根尖细胞分裂周期中微管骨架列阵进行详细观察,以探索高等植物微管周期的普遍性。结果表明:慈姑根尖固定后酶解可获得大量结构完整的去壁细胞与细胞团块;考马斯亮蓝染色观察可见,慈姑根尖细胞中丰富的蛋白物质以及处于不同分裂期的细胞核染色体;免疫荧光观察可见,慈姑根尖细胞周期中微管骨架保存较好,主要有周质微管、早前期带微管、纺缍体微管和成膜体微管4种循序变化的排列方式,构成了高等水生植物分裂细胞中典型的微管周期。实验结果证明,高等水生植物与陆生植物微管周期具有相似性,为植物微管周期概念提供了新的实例。     

Cell proliferation,cell shape,and microtubule and cellulose microfibril organization of tobacco BY-2 cells are not altered by exposure to near weightlessness in space

The microtubule cytoskeleton and the cell wall both play key roles in plant cell growth and division, determining the plant’s final stature. At near weightlessness, tubulin polymerizes into microtubules in vitro, but these microtubules do not self-organize in the ordered patterns observed at 1g. Likewise, at near weightlessness cortical microtubules in protoplasts have difficulty organizing into parallel arrays, which are required for proper plant cell elongation. However, intact plants do grow in space and therefore should have a normally functioning microtubule cytoskeleton. Since the main difference between protoplasts and plant cells in a tissue is the presence of a cell wall, we studied single, but walled, tobacco BY-2 suspension-cultured cells during an 8-day space-flight experiment on board of the Soyuz capsule and the International Space Station during the 12S mission (March–April 2006). We show that the cortical microtubule density, ordering and orientation in isolated walled plant cells are unaffected by near weightlessness, as are the orientation of the cellulose microfibrils, cell proliferation, and cell shape. Likely, tissue organization is not essential for the organization of these structures in space. When combined with the fact that many recovering protoplasts have an aberrant cortical microtubule cytoskeleton, the results suggest a role for the cell wall, or its production machinery, in structuring the microtubule cytoskeleton.     

Microtubule cytoskeleton: a track record

The plant microtubule cytoskeleton forms unique arrays during cell division and morphogenesis. Recent studies have addressed the biogenesis, turnover, spatio-temporal organisation and cellular function of microtubules. The results suggest that both conserved eukaryotic mechanisms and plant-specific modifications determine microtubule dynamics and function.     

植物激素对微管和纤维素微纤丝排向的调节

回顾了微管和纤维素微纤丝在细胞骨架构成和延展中的作用;综述了植物激素在微管和纤维素微纤丝排向中的调节功能,并对细胞扩大和伸长的机制进行了探讨。     

Plant microtubule cytoskeleton complexity: microtubule arrays as fractals

Biological systems are by nature complex and this complexity has been shown to be important in maintaining homeostasis. The plant microtubule cytoskeleton is a highly complex system, with contributing factors through interactions with microtubule-associated proteins (MAPs), expression of multiple tubulin isoforms, and post-translational modification of tubulin and MAPs. Some of this complexity is specific to microtubules, such as a redundancy in factors that regulate microtubule depolymerization. Plant microtubules form partial helical fractals that play a key role in development. It is suggested that, under certain cellular conditions, other categories of microtubule fractals may form including isotropic fractals, triangular fractals, and branched fractals. Helical fractal proteins including coiled-coil and armadillo/beta-catenin repeat proteins and the actin cytoskeleton are important here too. Either alone, or in combination, these fractals may drive much of plant development.     

Cytoskeletal organization during xylem cell differentiation

The water and mineral conductive tube, the xylem vessel and tracheid, is a highly conspicuous tissue due to its elaborately patterned secondary-wall deposition. One constituent of the xylem vessel and tracheid, the tracheary element, is an empty dead cell that develops secondary walls in the elaborate patterns. The wall pattern is appropriately regulated according to the developmental stage of the plant. The cytoskeleton is an essential component of this regulation. In fact, the cortical microtubule is well known to participate in patterned secondary cell wall formation. The dynamic rearrangement of the microtubules and actin filaments have also been recognized in the cultured cells differentiating into tracheary elements in vitro. There has recently been considerable progress in our understanding of the dynamics and regulation of cortical microtubules, and several plant microtubule associated proteins have been identified and characterized. The microtubules have been observed during tracheary element differentiation in living Arabidopsis thaliana cells. Based on this recently acquired information on the plant cytoskeleton and tracheary element differentiation, this review discusses the role of the cytoskeleton in secondary cell wall formation.     

Putative microtubule-associated proteins from the Arabidopsis genome

Summary. Plant microtubule-associated proteins (MAPs) are important in modulating the function of the microtubule cytoskeleton. Various plant MAPs have already been described. However, because of the complexity of the plant microtubule cytoskeleton and its responses to developmental and environmental stimuli, there are undoubtedly many more MAPs to be discovered. We have used a literature search and the BLAST protein comparison program to identify which model MAPs from other taxa have close homologues in Arabidopsis thaliana. The search revealed Arabidopsis homologues of 14 model MAPs, with E values (numbers of proteins that will match the model protein merely by chance) of <1×10^–10 and homologous domains spanning 98–599 amino acid residues, representing 57.1–97.0% of the model MAP sequence, as well as 22.5–72.8% amino acid identities and 76.3–96.2% conservation of secondary structure in the homologous domain. All of the Arabidopsis homologues have either a full cDNA clone or an expressed sequence tag in the GenBank database and therefore are expressed. The proteins are likely to regulate a variety of functions, including tubulin folding, microtubule nucleation and polymerisation dynamics, microtubule-dependent cell cycle control, organisation of microtubule arrays, interaction of microtubules with plasma-membrane-associated protein complexes, and interactions with various other proteins. The exact functions of these putative MAPs in the plant cell remain to be elucidated empirically. The identification of these putative MAPs opens new avenues for the investigation of the complexities of the plant microtubule cytoskeleton.Present address: School of Biological Sciences, University of Manchester, Manchester, United Kingdom.Correspondence and reprints: School of Biological Sciences A12, University of Sydney, NSW 2006, Australia.Received October 21, 2002; accepted December 30, 2002; published online September 23, 2003     

The plant cytoskeleton: recent advances in the study of the plant microtubule-associated proteins MAP-65, MAP-190 and the Xenopus MAP215-like protein,MOR1

The microtubule cytoskeleton is a dynamic filamentous structure involved in many key processes in plant cell morphogenesis including nuclear and cell division, deposition of cell wall, cell expansion, organelle movement and secretion. The principal microtubule protein is tubulin, which associates to form the wall of the tubule. In addition, various associated proteins bind microtubules either to anchor, cross-link or regulate the microtubule network within cells. Biochemical, molecular biological and genetic approaches are being successfully used to identify these microtubule-associated proteins (MAPs) in plants, and we describe recent progress on three of these proteins.     

The POLARIS peptide of Arabidopsis regulates auxin transport and root growth via effects on ethylene signaling

The rate and plane of cell division and anisotropic cell growth are critical for plant development and are regulated by diverse mechanisms involving several hormone signaling pathways. Little is known about peptide signaling in plant growth; however, Arabidopsis thaliana POLARIS (PLS), encoding a 36-amino acid peptide, is required for correct root growth and vascular development. Mutational analysis implicates a role for the peptide in hormone responses, but the basis of PLS action is obscure. Using the Arabidopsis root as a model to study PLS action in plant development, we discovered a link between PLS, ethylene signaling, auxin homeostasis, and microtubule cytoskeleton dynamics. Mutation of PLS results in an enhanced ethylene-response phenotype, defective auxin transport and homeostasis, and altered microtubule sensitivity to inhibitors. These defects, along with the short-root phenotype, are suppressed by genetic and pharmacological inhibition of ethylene action. PLS expression is repressed by ethylene and induced by auxin. Our results suggest a mechanism whereby PLS negatively regulates ethylene responses to modulate cell division and expansion via downstream effects on microtubule cytoskeleton dynamics and auxin signaling, thereby influencing root growth and lateral root development. This mechanism involves a regulatory loop of auxin-ethylene interactions.