纤毛虫大核和小核的形态及其发育过程

纤毛虫细胞内同时含有大核(macronucleus)和小核(micronucleus)两种类型的细胞核。大核和小核在结构和功能等诸方面有明显差异。但一些进化程度较低的纤毛虫大核和小核没有进一步的分化,甚至极少数纤毛虫仅含一种类型的细胞核。本文就纤毛虫大、小核的形态及其发育过程的基本特征,以及某些低等纤毛虫核器的特殊情况概述如下。     

纤毛虫大核分化过程中DNA的重组

纤毛虫大核分化过程中DNA的重组,包括小核染色体的断裂、间隙片段的删除、大核特定序列的扩增以及在大核基因的两端加上端粒。本文重点介绍和讨论了DNA片段删除与重排的类型、间隙DNA片段的序列特点,以及DNA删除与重排的机理。     

八种纤毛虫减数分裂基因的分子进化研究

减数分裂是真核生物适应性进化的重要机制,以8种纤毛虫作为实验对象,通过生物信息学方法对其14个减数分裂基因进行了鉴定及分子进化研究。结果表明:(1)不同的纤毛虫种类存在一些特异性的减数分裂基因的丢失与复制现象;(2)减数分裂相关基因在纤毛虫中很保守;(3)纤毛虫减数分裂重要的同源重组过程是在真核生物中不常见的Ⅱ型。本研究表明,纤毛虫减数分裂可能代表了真核生物较原始的减数分裂方式,在进化的过程中很保守,为研究真核生物减数分裂起源与进化提供了重要线索。     

纤毛虫大核提取方法上的改进及其扫描电镜观察

以包囊游仆虫(Euplotes encysticus)为材料,对Arikawa等报道的纤毛虫大核提取方法进行了改进,提出了一种更为简便有效的方法,即：用去垢剂Triton X-100处理细胞,结合玻璃微吸管的物理破碎,得到完整的大核后,应用扫描电镜观察。     

银杏小孢子中的微核形成及其在进化过程中的意义

通过ＤＡＰＩ（４’,６－ｄｉａｍｉｄｉｎｏ－２－ｐｈｅｎｙｌｉｎｄｏｌｅ）染色的压片,对银杏小孢子的形成过程进行了观察,发现小孢子母细胞减数分裂过程中,有的细胞在中期Ｉ、中期Ⅱ时出现染色体排列异常和后期染色体桥等畸变。所形成的四分体中,具有一定数量的微核;个别植株的四分体中可有高达６．５％的微核;具微核的小孢子很可能败育。结合古植物方面的研究结果,银杏小孢子形成过程的这些异常规象为认识银杏的演化趋     

纤毛虫大核发育过程中程序化的DNA消除

纤毛虫大核发育过程中发生程序性的ＤＮＡ消除,被消除的序列为内在删除顺序非编码区。消除以后,大核胚基中的ＤＮＡ发生片段化,在腹毛目纤毛虫中,ＤＮＡ断裂成为只有基因大小的ＤＮＡ分子;在四膜虫中,ＤＮＡ断裂成为平均长度约６００ｋｂ的片段,在断裂位点发现一个１５ｂｐ的顺序,为断裂位点的标志信号。通过对四膜虫小核基因组某一内在删除顺序的研究,发现此序列的删除受两套顺式作用顺序的控制。     

基因组功能预测的进化印记方法

改善基因组功能预测方案是目前功能基因组学的迫切问题,生物进化历程会在分子序列上留下相应进化印记－直系同源簇的特异模体,在这一生物学事实的基础上,提出了一个新的基因缚功能预测方法,首先利用进化分析方法构建直系同源簇,再找到各直系同源簇的功能模体,这样可以形成特异的功能模体库,未知基因的功能预测可望通过搜索该功能模体库而得以高效,准确地完成,对５个家族的检验初步证实该方案是可行的。     

RNA干涉在纤毛虫中的研究进展

RNA干涉是dsRNA介导的基因沉默现象,本文简要介绍了其作用的机制和生物学意义,重点阐述了RNA干涉在原生动物纤毛虫中的发现与应用,比较了RNA干涉与纤毛虫大核基因组重排机理的异同,并对RNA干涉在纤毛虫中传输的技术途径-RNAi喂饲法的原理也做了详细的介绍。     

DNA进化马尔可夫过程模型的评价与推广

本文对ＤＮＡ序列进化过程中核苷酸替代的随机模型进行了评价,对替代速率在时间和空间上不恒定的情形进行了考察和推广。Ｌａｎａｖｅ等（１９８４）曾提出一个模型,宣称对替代的模式未做任何假定,但事实上我们证明它假定替代过程是可逆的。运用２－ｐ、４－ｐ和６－ｐ模型进行的计算表明替代速度在位点间的差异会造成估计的替代数严重偏低,并且替代数越大,偏差也越大。替代模式在位点间的差异也会造成估计值偏低,但偏差不严重     

筛选冠突伪尾柱虫有小核细胞系和无小核细胞系饥饿期差异表达的ESTs

利用显微切割的方法建立大型腹毛目纤毛虫---冠突伪尾柱虫(Pseudourostyla cristata)的无小核细胞系,分别提取有小核和无小核细胞系的总RNA,用SMART-PCR方法直接合成dscDNA,进而运用抑制性消减杂交技术(Sup-pression subtractive hybridization,SSH)对冠突伪尾柱虫去小核前后的差异表达基因进行研究,构建了有与无小核细胞系在饥饿期差异表达基因的消减文库,随机挑取了284个克隆,应用cDNA阵列技术鉴定阳性克隆。将鉴定出的17个阳性克隆进行Blastx分析,结果表明17个EST(Expressed sequence tag)均可能与小核体功能密切相关,此为进一步阐明小核的遗传机理奠定了基础。       

Characteristics of the membranes of the pellicle of the ciliatePseudomicrothorax dubius

Summary The membranes of the pellicle of the ciliatePseudomicrothorax dubius are investigated using thin section electron microscopy and freeze-fracture replicas. The plasma membrane is covered by a surface coat and is connected to the outer alveolar membrane by short, sometimes branched, bridges. The inner alveolar membrane is coated on both sides. The epiplasm lies in intimate contact with the cytoplasmic surface of this membrane, and there is a corresponding deposit on the other surface. This deposit is regularly striated.The epiplasmic layer and the alveoli are interrupted at sites of cytotic activity,e.g., the attachment sites of trichocysts, the cytoproct, and the parasomal sacs. The striated deposit ends where the epiplasm ends, indicating a direct relationship between these two epimembranous layers.There is a deposit along the sides of the first part of the tip of the trichocysts, and in this region the trichocyst membrane is free of intramembranous particles.The membrane of the parasomal sacs has a coat on both surfaces. That on the extraplasmic surface is similar to the surface coat of the plasma membrane. The origin of the cytoplasmic coat is unknown. The cytotic activity of these sacs is indicated by their highly irregular profiles.     

Biochemical and Cytological Evidence for an Overabundance of Mucocysts in the bcd Pattern Mutant of Tetrahymena thermophila

ABSTRACT. Three acidic proteins (42 kD, 43 kD and 50 kD) were present in unusually high concentrations in cortical preparations of the Tetrahymena pattern mutant broadened cortical domains (bcd). Antisera to the 42-kD and 50-kD proteins bound to discharging mucocysts and food vacuole contents in both wild-type and mutant cells. Subsequent analysis revealed that bcd mutant cell pellicles possess five times more "docked" mucocysts than their wild-type counterparts.     

The 44-kDa Regulatory Subunit of the Paramecium cAMP-Dependent Protein Kinase Lacks a Dimerization Domain and May Have a Unique Autophosphorylation Site Sequence

ABSTRACT. The 44-kDa regulatory subunit (R₄₄) of one form of cAMP-dependent protein kinase of Paramecium was purified, and two partial internal amino acid sequences from it were used to clone the corresponding cDNA. This R₄₄ cDNA clone was 1022-bp long, including 978 bp of coding sequence and 7 bp and 37 bp of 5' and 3' untranslated sequences, respectively. A 1.1-kb mRNA was labeled on a Northern blot. The deduced R₄₄ amino acid sequence had 31%–38% positional identity to the sequences of other cloned cAMP-dependent protein kinase regulatory subunits. R₄₄ sequence showed equal sequence similarity to mammalian types I and II regulatory subunits. The N -terminal sequence encoding the regulatory subunit dimerization domain found in most regulatory subunits is not present in the R₄₄ clone, confirming the lack of regulatory subunit dimer formation previously reported for the Paramecium cAMP-dependent protein kinase. The putative autophosphorylation site of R₄₄ contains the amino acid sequence TRTS, distinct from the consensus sequence RRXS, where X is any residue, found in other autophosphorylated cAMP-dependent protein kinase regulatory subunits and many cAMP-dependent protein kinase substrates.     

Genotoxic activity of four metabolites of the soy isoflavone daidzein

Products containing phytoestrogens are increasingly promoted as the “natural” alternative to estrogen replacement therapy. In the present study, we have used the in vitro micronucleus assay in L5178Y mouse lymphoma cells to investigate the genotoxic potential of the isoflavone daidzein, and of four daidzein metabolites known to be formed in humans. Whereas no induction of micronuclei was observed with daidzein up to the limit of solubility (100 μM), all four daidzein metabolites, i.e. equol (2.3-fold induction at 100 μM), O-desmethylangolensin (6.2-fold induction at 10 μM), 4′,6,7-isoflavone (6.7-fold induction at 100 μM) and 3′,4′,7-isoflavone (8.2-fold induction at 100 μM) induced micronuclei in a concentration-dependent manner. Thus, both reductive and oxidative metabolites of the soy isoflavone daidzein exhibit genotoxic potential in vitro.     

Temperature-induced modifications in size and pattern of microtubular organelles in a ciliate,Dileptus II. Formation and spatial arrangement of the microtubular skeleton of the oral parts

Summary Microtubular organelles formed during continuous exposure to high or low temperature were studied. Neither heat nor cold prevents formation of the microtubular skeleton in the oral parts of the ciliateDileptus. Elevated temperature causes the formation of short microtubular fibres, while the size of the oral structure is not diminished. This leads to failure in sculpturing of the cytostome. Heat-treatment may also alter the localization of the anchor site of fibres around the circumference of the basal bodies, and the orientation of the fibres. Cold-treatment evokes the formation of small mouthparts containing a lower number of organelles, although these are properly shaped and there are no deviations in the position or orientation of fibres. It seems that low temperature may suppress the rate of formation of microtubular organelles, while elevated temperature affects their patterning.Abbreviations MTOC microtubule organizing centre - T transverse fibres - B basal body - Cy cytostome - K kinetodesma - P postciliary fibre - C compound fibre - L lamina - F filamentous bundle - M monokinetid     

SINE Retrotransposition During the Evolution of the Pecoran Ruminants

SINE retrotransposition events have proven their value as phylogenetic markers in several eukaryotic taxa at different taxonomic levels. The genomes of ruminants contain three related SINE elements, Bov-tA, Bov-A2, and Bov-B. To estimate the time points of retrotransposition of individual copies of these SINEs, we designed PCR primers on database sequences containing SINE insertions in cattle, sheep, or goat genomes and tested for the presence of these copies in the genomes of other ruminants. It was checked by sequencing whether length variation of the PCR products reflected a SINE retrotransposition. One Bov-B and nine Bov-tA insertions were shared by cattle, sheep, goat, and giraffe, indicating an early retrotransposition event before the radiation of the Pecora, while three other Bov-tA and two Bov-B elements were apparently inserted later. The ruminant α-lactalbumine gene contains a hotspot of early and more recent Bov-tA insertions, a Bov-tA replacement as well as a recent Bov-B insertion. Three Bov-A2 insertions were found to be shared only by the Bovidae, the Bovini, and the Bos and Bison species, respectively, indicating that most Bov-A2 insertions are relatively recent. The time elapsed since the retrotransposition was also reflected in the degeneration of the direct repeats that flank SINE inserts. We suggest that retrotransposition of SINEs may serve as phylogenetic markers in the ruminant families, subfamilies, and even tribes. In addition, sequencing of SINE insertions revealed several other unique deletions/insertions that also may be informative for phylogenetic reconstructions of ruminants. Received: 19 January 2001 / Accepted: 6 June 2001     

Recombination During In Vitro Evolution

Recombination, the swapping of large portions of genetic information between and among parental genotypes, can be applied to in vitro evolution experiments on functional nucleic acids. Both homologous and heterologous recombination can be achieved using standard laboratory techniques. In many cases, recombination can allow for the discovery of a ribozyme or DNAzyme phenotype that would not likely be encountered by reliance on point mutations alone. In addition, recombination can often aid in the discovery of global optima in sequence space and/or lessen the number of generations it would take to reach optima. Recombination is most efficiently used in combination with point mutations and applied after the first couple of rounds of selection but before high-fitness genotypes dominate the selection. The “recombination zone” describes that region of sequence space—defined by the residues that will ultimately participate in the function of the winning nucleic acid(s)—where recombination is expected to be the most beneficial in the search for high-fitness genotypes.[Reviewing Editor: Martin Kreitman]Author order determined by a single Bernoulli trial as implemented by RPS.