Distribution of rDNA in the nucleus of Giardia lamblia: detection by Ag-I silver stain.

Previous investigations have proved that diplomonads have primitive cell nuclei and lack a nucleolus. We determined the distribution of ribosomal DNA (rDNA) in diplomonad nuclei that lacked a nucleolus. Giardia lamblia was used as the experimental organism with Euglena gracilis as the control. The distribution of rDNA was demonstrated indirectly by the modified Ag-I silver technique that can indicate specifically the nucleolus organizing region (NOR) by both light and electron microscopy. In ultrathin sections of silver stained Euglena cells, all silver grains were concentrated in the fibrosa of the nucleolus, while no silver grains were found in the cytoplasm, nucleoplasm, condensed chromosomes or pars granulosa of the nucleus. In the silver stained Giardia cells, no nucleolus was found, but a few silver grains were scattered in the nucleus. This suggests that the rDNA of Giardia does not form an NOR-like structure and that its nucleus is in a primitive state.     

Effect of external environmental factors on DNA content in Euglena]

The chromosome number and DNA content in Euglena gracilis cell have been shown by the number of investigators to vary essentially under the influence of some environmental factors (low phosphate and acetate concentrations, density of culture, temperature, light/dark regime, streptomycin treatment). Variability in the nuclear DNA quantity is supposed to be an important peculiarity of the structure and functional activity of the genome of Euglena gracilis.     

Isolation, fractionation and template activity of the continuously-condensed chromatin of Euglena gracilis

A technique for isolating whole chromatin from nuclei of the lower eukaryote Euglena gracilis is presented. This chromatin, which appears under the electron microscope as uniformly condensed fibers, can, nevertheless, be subfractionated into distinct heterochromatic and euchromatic fractions. The euchromatin, comprising about 14% of the total DNA of the nucleus, contains over 80 % of the total endogenous RNA polymerase activity measured. The K_m for this enzyme is higher than that found for prokaryotes, but falls in the range found for other eukaryotes. Stability constants, calculated from cation-chromatin binding data, suggest that internal carboxyl groups of chromosomal proteins, at least, are involved in the condensation of Euglena chromatin. The relationship between Euglena chromatin and that of higher eukaryotes is discussed.     

Evidence for the late origin of introns in chloroplast genes from an evolutionary analysis of the genus Euglena.

The origin of present day introns is a subject of spirited debate. Any intron evolution theory must account for not only nuclear spliceosomal introns but also their antecedents. The evolution of group II introns is fundamental to this debate, since group II introns are the proposed progenitors of nuclear spliceosomal introns and are found in ancient genes from modern organisms. We have studied the evolution of chloroplast introns and twintrons (introns within introns) in the genus Euglena. Our hypothesis is that Euglena chloroplast introns arose late in the evolution of this lineage and that twintrons were formed by the insertion of one or more introns into existing introns. In the present study we find that 22 out of 26 introns surveyed in six different photosynthesis-related genes from the plastid DNA of Euglena gracilis are not present in one or more basally branching Euglena spp. These results are supportive of a late origin for Euglena chloroplast group II introns. The psbT gene in Euglena viridis, a basally branching Euglena species, contains a single intron in the identical position to a psbT twintron from E.gracilis, a derived species. The E.viridis intron, when compared with 99 other Euglena group II introns, is most similar to the external intron of the E.gracilis psbT twintron. Based on these data, the addition of introns to the ancestral psbT intron in the common ancester of E.viridis and E.gracilis gave rise to the psbT twintron in E.gracilis.     

特殊涡鞭毛虫——尖尾虫染色体与典型涡鞭毛虫染色体的比较研究

我们实验室与上海细胞生物学研究所的有关同志多年来在特殊涡鞭毛虫──尖尾虫（Ｏｘｙｒｒｈｉｓｍａｒｉｎａ）的细胞生物学和生物化学上作了一系列的研究,本文所报道的是这些工作中的一部分。对尖尾虫（Ｏｘｙｒｒｈｉｓｍａｒｉｎａ）的永久浓聚染色体的精细形象以及不同固定方法对其精细形象的影响作了观察,并与人肠道细菌的类核体、典型涡鞭毛虫原甲藻（Ｐｒｏｒｏｃｅｎｔｒｕｍｍｉｃａｎｓ）和鞭毛虫眼虫（Ｅｎｇｌｅｎａｓｐ．）的永久浓聚染色体作了比较。结果表明,细菌的类核体的精细形象受固定方法的影响极大。反之,不同的固定方法对于眼虫染色体的精细形象则看不出有任何显著的影响。至于典型涡鞭毛虫类的染色体,单用ＯｓＯ４或用戊二醛－ＯｓＯ４双固定法固定,都会得到典型涡鞭毛虫类染色体的横带样结构,然而单纯用戊二醛固定,却会得到大不相同的形象。在这方面尖角虫的染色体与一般的涡鞭毛虫类的染色体相距甚远,其染色体的精细形象本身也与眼虫类的染色体较为相似,而与一般涡鞭毛虫类的大不相同。本工作所得到的结果与以前我们在不同方面所得到的结果一致。研究结果全都表明,失足虫与典型涡鞭毛虫有着一系列重大的差异,实际上代表着一个介于一般鞭毛虫类与典型涡鞭毛虫类     

The nuclear matrix of Euglena gracilis (Euglenophyta): A stage of nuclear matrix evolution?

Euglena gracilis cell was extracted sequentially with CSK-Triton buffer, RSB-Magik solution and DNase-As solution. DGD embedment-free electron microscopy showed that in the extracted nucleus there was a residual non-chromatin fibrous network. That it could not be removed by hot trichloroacetic acid further supported the idea that it was a non-histone, non-chromatin fibrous protein network, and should be the internal network of the nuclear matrix. After the sequential extraction, the nuclear membrane was removed, leaving behind a layer of lamina; the chromatin was digested and eluted from the dense chromosomes and residual chromosomal structures that should be chromosomal scaffold were revealed. Western blot analysis with antiserum against rat lamins showed that nuclear lamina of the cell possessed two positive polypeptides, a major one and a minor one, which had molecular masses similar to lamin B and lamin A, respectively. Comparing these data with those of the most primitive eukaryote Archezoa and of higher eukaryotes, it was suggested that the lower unicellular eukaryote E. gracilis already had the nuclear matrix structure, and its nuclear matrix (especially the lamina) might represent a stage of evolutionary history of the nuclear matrix.     

Morphological changes of sperm nuclei during spermatogenesis in the brown alga Cystoseira hakodatensis (Fucales, Phaeophyceae)

Morphological changes and chromatin condensation of sperm nuclei were observed during spermatogenesis in the fucalean brown alga Cystoseira hakodatensis (Yendo) Fensholt. Ultrastructural studies have shown that the mature spermatozoid has an elongated and concave nucleus with condensed chromatin. The morphological changes and the chromatin condensation process during spermatogenesis was observed. Nuclear size decreased in two stages during spermatogenesis. During the first stage, spherical nuclei decreased in size as they were undergoing meiotic divisions and the subsequent mitoses within the antheridium. During the second stage, the morphological transformation from a spherical into an elongated nucleus occurred. Afterwards, chromatin condensed at the periphery in each nucleus, and chromatin‐free regions were observed in the center of the nucleus. These chromatin‐free regions in the center of nucleus were compressed by the peripheral chromatin‐condensed region. As the result, the elongated and concave nucleus of the mature sperm consisted of uniformly well‐condensed chromatin.     

An Ultrastructural Comparison of Euglena gracilis Klebs, Bleached Euglena and Astasia longa Pringsheim

SYNOPSIS. Some of the cytologic features of Euglena gracilis Klebs, Euglena gracilis var. bacillaris Gross (colorless), and Astasia longa Pringsheim were compared in an electron-microscope study. E. gracilis was readily differentiated from the colorless forms by the presence of chloroplasts. The 2 colorless forms were structurally similar; however, it was possible to differentiate bleached Euglena from Astasia longa by the presence of irregular plate-like aggregates of paramylum in the former; similar forms of paramylum were not seen in chloroplast-containing E. gracilis or in A. longa .     

Evolution of the Chloroplast Genome in Photosynthetic Euglenoids: A Comparison of Eutreptia viridis and Euglena gracilis (Euglenophyta)

The chloroplast genome of Eutreptia viridis Perty, a basal taxon in the photosynthetic euglenoid lineage, was sequenced and compared with that of Euglena gracilis Ehrenberg, a crown species. Several common gene clusters were identified and gene order, conservation, and sequence similarity was assessed through comparisons with Euglena gracilis. Significant gene rearrangements were present between Eutreptia viridis and Euglena gracilis chloroplast genomes. In addition, major expansion has occurred in the Euglena gracilis chloroplast accounting for its larger size. However, the key chloroplast genes are present and differ only in the absence of psaM and roaA in Eutreptia viridis, and psaI in Euglena gracilis, suggesting a high level of gene conservation within the euglenoid lineage. Further comparisons with the plastid genomes of closely related green algal taxa have provided additional support for the hypothesis that a Pyramimonas-like alga was the euglenoid chloroplast donor via secondary endosymbiosis.     

Electron microscope study of chromatin in hepatocyte nuclei during the first hours after partial hepatectomy. V. Changes in the relative area of condensed chromatin and the density of chromatin fibril packing in the ultrathin sections

The degree of chromatin condensation was studied on ultrathin cell sections of guinea pig hepatocytes during the prereplicative period after partial hepatectomy. Three time points were chosen for analysis namely 2,5, 5 and 9 hrs after operation since they show marked increasing (2.5 hrs), decreasing (5 hrs) and repeated increasing (9 hrs) of the amount of ethidium bromide binding to chromatin. The degree of chromatin condensation was determined by measuring the area occupied by condensed chromatin and also by measuring the number of chromatin fibrils per a certain length. The condensed chromatin with varying localization in the nucleus were studied separately. The changes of nucleoplasmic chromatin were most pronounced: at 2.5 and 9 hrs after operation the decrease of the relative area and of the density of chromatin fibrils package was observed; these parameters were near to control at 5 hrs after operation. In general the changes in nucleoplasmic chromatin were correlated with the changes of the activity of the chromatin in the whole nucleus. The decondensation of the perimembranous chromatin was manifested in the decrease of its area and was expressed only at 9 hrs after operation. The perinucleolar chromatin was found to show the gradual decondensation which was manifested mainly by the decrease of its relative area. Thus the condensed chromatin seems to be a labile structure which undergoes essential changes in the process of the exit of the hepatocytes from G0-stage of the cell cycle, during the prereplicative period.     

Guanine Metabolism in Carbon-Limited Euglena gracilis

SYNOPSIS. Guanine-8-³H is not a specific label for nucleic acids in Euglena gracilis (strain SM-L1, streptomycin-bleached), but is incorporated into all biochemical fractions. Autoradiographic results show that guanine is metabolized in, both the nucleus and the cytoplasm of these Eugelna , but nuclear metabolism of guanine occurs only at a very low level in carbon-limited cells. Further, carbon-limited Euglena incorporate less total guanine than do control Euglena in a comparable time period. However, the majority of label incorporated into the nucleic acid fraction of the carbon-limited cells is incorporated into RNA.     

Actin Gene Sequence from Euglena gracilis

ABSTRACT The full length coding sequence of the Euglena gracilis actin gene was determined by RT-PCR of Euglena gracilis mRNA. Conserved regions in the actin amino acid sequence were used as guides for the synthesis of degenerate primers. Sequence was obtained for 1.238 nucleotides, of which 1.131 were coding for 377 amino acids. Sequence comparisons showed a similarity with other actins of 56% to 80%. Even though most of the actin amino acid sequence was conserved, some regions showed high divergence, i.e. the DNase I-binding loop at the N-terminal region. The construction of a phylogenetic tree based on actin sequences from different organisms placed Euglena gracilis in a cluster with Trypanosoma brucei and Leishmania major.     

Gravitaxis in the flagellate Euglena gracilis--results from NiZeMi, clinostat and sounding rocket flights

Many motile microorganisms including flagellates such as the green Euglena gracilis move up and down within the water column and use a number of external clues for their orientation, the most important of which may be light and gravity. The cells use positive phototaxis and negative gravitaxis to move closer to the surface of the water column which for energetic reasons is vital for their survival. However, most phytoplankton organisms cannot tolerate the bright irradiance of unfiltered solar radiation at the surface which also bleaches the photosynthetic pigments, disables the photosynthetic apparatus and impairs phototaxis, gravitaxis and motility in Euglena. Thus, it is not surprising that at higher irradiances negative phototaxis operates antagonistically to the responses described above to guide the cells into deeper water where they are protected from excessive radiation. Phototaxis and gravitaxis are not independent from one another: in a vertically positioned cuvette negative gravitaxis can be "titrated" by light impinging from above and is compensated at about 30 W m-2. While the photoreceptor for phototaxis has been identified in Euglena gracilis biochemically and spectroscopically, the gravireceptor is not yet known. Young cultures of Euglena gracilis show a positive gravitaxis, the ecological signficance of which is not yet understood while older cultures show negative gravitaxis. One hypothesis concerning the nature of graviperception is based on a passive physical process such as an asymmetric distribution of the mass within the cell. However, the observation that short term UV irradiation decreases the precision of negative gravitaxis rather indicates the involvement of an active physiological gravireceptor. Furthermore, some heavy metal ions have been found to change the direction of movement from positive to negative gravitaxis in young cells.     

Effect of glucose on carbohydrate synthesis from alanine or lactate in hepatocytes from starved rats.

Euglena gracilis was found to contain a peroxidase that specifically require L-ascorbic acid as the natural electron donor in the cytosol. The presence of an oxidation-reduction system metabolizing L-ascorbic acid was demonstrated in Euglena cells. Oxidation of L-ascorbic acid by the peroxidase, and the absence of ascorbic acid oxidase activity, suggests that the system functions to remove H2O2 in E. gracilis, which lacks catalase.     

The fractionation of histones isolated from Euglena gracilis.

1. The histones of Euglena gracilis were separated by gel filtration into five fractions. 2. Each fraction was characterized in terms of its electrophoretic, solubility and compositional properties. 3. Euglena gracilis clearly contains histones corresponding to vertebrate H1, H2B, H3 and H4 fractions, although they all differ in containing more lysine. 4. The remaining Euglena histone is considered to be homologous to vertebrate histone H2A, but it differs in having a much higher ratio of lysine to arginine. 5. The Euglena histone H1 appears to be lacking in aspartic acid. 6. Electrophoresis in the presence of sodium dodecyl sulphate indicates that the molecular weights of the Euglena histones are close to those of the homologous vertebrate histones.     

Amino acid sequence of calmodulin from Euglena gracilis.

The complete amino acid sequence of calmodulin from Euglena gracilis was determined by isolation and sequence analyses of peptides derived from calmodulin by digestion with trypsin and Staphylococcus aureus V8 protease. Euglena calmodulin consists of 148 amino acid residues; it lacks tryptophan and cysteine and contains one tyrosine, three histidine and two NE-trimethyllysine residues/molecule of the protein. Its N-terminus was blocked with an acetyl group and C-terminal lysine was trimethylated. Euglena calmodulin is the first calmodulin so far examined in which the C-terminal lysine is trimethylated. The comparison of amino acid sequences between Euglena and human brain calmodulins indicated 17 amino acid substitutions in Euglena calmodulin.