In vitro assembly of plant tubulin in the absence of microtubule-stabilizing reagents

The assembly of microtubules is essential for physiological functions of microtubules. Addition of microtubule-stabilizing reagents or microtubule “seeds” is usually necessary for plant tubulin assemblyin vitro, which hinders the investigation of plant microtubule dynamics. In the present note, highly purified plant tubulins have been obtained from lily pollen, a non-microtubule-stabilizing reagent or microtubule “seed” system for plant tubulin assembly has been established and the analysis of plant tubulin assembly performed. Experiment results showed that purified tubulin polymerizedin vitro, and a typical microtubule structure was observed with electron microscopy. The kinetics curve of tubulin assembly exhibited typical “parabola”. The presence of taxol significantly altered the character of plant tubulin assembly, including that abnormal microtubules were assembled and the critical concentration for plant tubulin assembly was decreased exceedingly from 3 mg/mL in the absence of taxol to 0.043 mg/mL in ihe presence of taxol.     

Progress in molecular parasitology

Summary Substantial progress has been made in the last ten years in understanding the structural and functional organization of parasitic protozoa and helminths and the complex physiological relationships that exist between these organisms and their hosts. By employing the new powerful techniques of biochemistry, molecular biology and immunology the genomic organization in parasites, the molecular basis of parasite's variation in surface antigens and the biosynthesis, processing, transport and membrane anchoring of these and other surface proteins were extensively investigated. Significant advances have also been made in our knowledge of the specific and often peculiar strategies of intermediary metabolism, cell compartmentation, the role of oxygen for parasites and the mechanisms of antiparasitic drug action. Further major fields of interest are currently the complex processes which enables parasites to evade the host's immune defense system and other mechanisms which have resulted in the specific adaptations which enabled parasites to survive within their host environments. Various approaches in molecular and biochemical parasitology and in immunoparasitology have been proven to be of high potential for serodiagnosis, immunoprophylaxis and drug design.This paper is based on a review presented at a workshop on Molecular Parasitology, organized by the Swiss Society of Tropical Medecine and Parasitology at the University of Neuchâtel, March 1985.     

包囊游仆虫包囊形成和解脱过程中微管蛋白基因表达的变化

采用干涉相差显微术和实时荧光定量PCR(Q-PCR)技术显示,腹毛类纤毛虫包囊游仆虫(Euplotes encysticus)包囊形成和解脱过程中,细胞微管胞器始终处于非装配及装配的功能活动状态,脱包囊过程中伸缩泡的运动对细胞充分吸水用于微管胞器的再装配及细胞运动起了关键的作用,细胞内α-、β-和γ-微管蛋白基因的表达量相伴发生变化.其中,微管胞器去分化时,细胞形成毛基体非吸收型包囊,细胞α-、β-和γ-微管蛋白基因的表达量整体呈现逐渐减少的趋势,其细胞微管蛋白基因的相对表达量从大到小依次是β-、α-和γ-微管蛋白基因,但α-、β-微管蛋白基因的起始拷贝数远大于后一种微管蛋白基因的表达量;微管胞器再分化时,伴随着伸缩泡的剧烈伸缩运动,口纤毛器和体纤毛器微管先后形成,细胞在包囊内做旋转运动并脱囊而出,迅速转化为营养细胞,期间细胞内α-、β-和γ-微管蛋白基因的表达量呈现从小到大增加的趋势,其微管蛋白基因的相对表达量从大到小依次是β-、α-和γ-微管蛋白基因,而前两种微管蛋白基因的起始拷贝数远大于后一种微管蛋白基因的表达量.结果表明,包囊游仆虫形成包囊和脱包囊过程中,伴随着皮层微管胞器的不同分化,细胞内α-、β-和γ-微管蛋白基因始终处于不同的功能活动状态,其中作为微管组织中心主要组分的γ-微管蛋白也始终处于不同的功能状态.     

马达蛋白运动轨道的静电特性

在微管结构特点的基础上,介绍了马达蛋白运动轨道的静电特性,如电荷分布、电偶极以及相互作用势,为构造马达蛋白的物理模型提供较为真实的物质基础．     

In vitro assembly of plant tubulin in the absence of microtubule-stabilizing reagents

The assembly of microtubules is essential for physiological functions of microtubules. Addition of microtubule-stabilizing reagents or microtubule "seeds" is usually necessary for plant tubulin assembly in vitro, which hinders the investigation of plant microtubule dynamics. In the present note, highly purified plant tubulins have been obtained from lily pollen, a non-microtubule-stabilizing reagent or microtubule "seed" system for plant tubulin assembly has been established and the analysis of plant tubulin assembly performed. Experiment results showed that purified tubulin polymerized in vitro, and a typical microtubule structure was observed with electron microscopy. The kinetics curve of tubulin assembly exhibited typical "parabola". The presence of taxol significantly altered the character of plant tubulin assembly, including that abnormal microtubules were assembled and the critical concentration for plant tubulin assembly was decreased exceedingly from 3 mg/mL in the absence of taxol to 0.043 mg/mL in the presence of taxol.     

A basic protein from bovine brain that co-precipitates with tubulin in vitro

A 193 kDa protein consisting of 58 kDa subunits, which has pI values of 8.50 and 8.65, was purified from bovine brain cytosol. It formed heavy precipitates with tubulin, and the molar ratio of tubulin dimer to this protein in the precipitate was 3.2. In contrast to microtubules containing ordinary microtubule-associated proteins, these complexes remained stable against cold and 1 mM CaCl₂.Acknowledgments. This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan.     

八肋游仆虫一个新β-微管蛋白基因的克隆与序列分析

从八肋游仆虫细胞中克隆到一种新的β-微管蛋白基因.序列分析结果表明:在大核中,该基因全长1561bp.与已经报道过的β-微管蛋白基因不同,它的5′端基因上游调控序列有49 bp,AT含量为75.5%.上游调控序列中仅有一个TATAA框,没有CCAAT框;该基因的3′端的下游调控序列有121 bp,富含AT碱基,达到86.8%,其中有明显的反向重复序列.上下游调控序列中各有一个断裂信号TTGAA和TTCAA,负责从小核到大核发育过程中大核染色体的形成.从小核基因组中克隆到相应的基因片段,比大核中的序列多7个碱基5′-CATGCTC-3′,其功能尚不清楚.序列比对表明,新的β-微管蛋白基因与已经报道的β-微管蛋白基因的同源性92.3%,阅读框中有2个氨基酸的变化,将它命名为β2-微管蛋白基因.