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H Ris

Publications and source records attributed to H Ris.

At least 37 records · Page 2Linked to original sources
PubMed1980

Origin of kinetochore microtubules in Chinese hamster ovary cells.

We have attempted to determine whether chromosomal microtubules arise by kinetochore nucleation or by attachment of pre-existing microtubules. The appearance of new microtubules was investigated in vivo on kinetochores to which microtubules had not previously been attached. The mitotic apparatus of Chinese hamsters ovary cells was reconstructed in three dimensions from 0.25 micrometer thick serial sections, and the location of chromosomes, kinetochore outer disks, centrioles, virus-like particles and microtubules determined. Central to the interpretation of these data is a synchronization scheme in which cells entered Colcemid arrest without forming mitotic microtubules. Cells were synchronized by the excess thymidine method and exposed to 0.3 microgram/ml Colcemid for 8 h. Electron microscopic examination showed that this Colcemid concentration eliminated all microtubules. Mitotic cells were collected by shaking off, and cell counts showed that over 95% of the cells were in interphase when treatment began and thus were arrested without the kinetochores having been previously attached to microtubules. Cells were then incubated in fresh medium and fixed for high voltage electron microscopy at intervals during recovery. -- In early stages of recovery, short microtubules were observed near and in contact with kinetochores and surrounding centrioles. Microtubules were associated with kinetochores facing away from centrosomes and far from any centrosomal microtubules, and thus were not of centrosomal origin. At a later stage of recovery, long parallel bundles of microtubules, terminating in the kinetochore outer disk, extended from kinetochores both toward and away from centrosomes. Because microtubules had never been attached to kinetochores, the possibility that kinetochore microtubules were initiated by microtubule stubs resistant to Colcemid was eliminated. Therefore we conclude that mammalian kinetochores can initiate microtubules in vivo, thus serving as microtubule organizing centers for the mitotic spindle, and that formation of kinetochore-microtubule bundles is not dependent on centrosomal activity.

Animals↗
PubMed1979

Electron microscopy of the spindle in locally heated cells.

Individual living cells in metaphase were exposed to a steep temperature gradient by placing a microheater near one spindle pole. The cells were then fixed and the spindle was examined by electron microscopy. The structure of the warmer half-spindle differed from the cooler half-spindle in several ways. Kinetochore microtubules were nearly parallel in the warmer half-spindle but were divergent in the cooler. The total length of microtubules in the warmer half-spindle was 52 per cent greater and the number of kinetochore microtubules per kinetochore averaged 16 per cent higher than in the cooler half-spindle. The warmer half-spindle was longer than the cooler. These observations clearly demonstrate a locally enhanced assembly of microtubules in the warmer half-spindle. The electron microscope study makes still clearer the unusual character of chromosome movement in the differentially heated cells: the structure of the warmer half-spindle is hard to distinguish from that in normal cells, yet chromosome movement there is far slower than normal (Nicklas, 1979).

Animals↗
PubMed1978

Comparison of active zones in spinal cord of anesthetized and unanesthetized rats: a high voltage electron microscope study.

High voltage electronmicroscopy was applied to the study of presynaptic nerve terminals and specifically the arrangement and staining properties of synaptic vesicles in and around the active zone. The zinc iodideosmium tetroxide technique was used for the impregnation of neuropil derived from the spinal cord of anesthetized (40 mg/kg pentobarbital sodium) and unanesthetized rats. Under optimal conditions of section thickness and tilt angles stereoscopic views of a paracrystalline lattice arrangement of synaptic vesicles could be ascertained. The ZIO affinity was significantly higher in synaptic vesicles of the anesthetized as compared with the unanesthetized preparations.

Animals↗
PubMed1976

Electron-microscopic study of the spindle and chromosome movement in the yeast Saccharomyces cerevisiae.

Mitosis in yeast Saccharomyces cerevisiae was investigated in thick (0-25-I mum) serial sections with a high voltage electron microscope and in preparations of spheroplasts spread on a water surface. Spindle microtubules originate from a plaque-like structure called the spindle pole bosis the SPB duplicates and a set of long and short microtubules develops on each SPB. The spindle arises as the SPBs separate on the nuclear membrane adense and are not individually visible. Genetic studies, however, have indicated that there are 17 linkage groups. The number of microtubules was determined in diploid and haploid spindles on serial stereo micrographs. In diploid mitosis about 40 microtubules issue from a SPB. Most are non-continuous and often they are visibly associated with a chromatin fibre. The spindle in haploid cells is similar except that the number of microtubules is about half that in diploid cells and the SPB is smaller. The pole-to-pole microtubules vary in number from spindle to spindle, but in each case enough microtubules are present to account for each linkage group being associated with a single non-continuous microtubule. We conclude that mitosis in yeast is comparable in its general aspect to that observed in typical eukaryotes.

Chromatin↗
PubMed1975

Primitive mitotic mechanisms.

Unorthodox mitotic mechanisms are reviewed and their contribution to the understanding of evolution of the orthodox mitotic apparatus is considered. Dinoflagellates and hypermastigote flagellates are of particular significance because the microtubular mitotic apparatus is entirely extranuclear with the nuclear membrane persisting through mitosis. Chromosomes are attached to the nuclear membrane. In hypermastigole flagellates early kinetochore separation is on the nuclear membrane without any contribution from microtubules. In dinoflagellates the chromosomes are also attached to the nuclear membrane, but at least in some species cytoplasmic microtubules connect to the attachment site. In Syndinium the attachment site resembles a typical kinetochore, but is inserted in the nuclear membrane. A similar kinetochore is found in certain Radiolaria, but with an intranuclear spindle apparatus the association with the nuclear membrane is no longer necessary and has been lost. Mitosis in the yeast Saccharomyces is essentially orthodox, though chromosomes do not condense. No kinetochores are seen, but a single microtubule makes direct contact with the 20 nm chromatin fiber of each chromosome and shortens during anaphase. About 5-10 microtubules are continuous between the spindle pole bodies and form the elongating central spindle.

Biological Evolution↗
PubMed1974

An unusual mitotic mechanism in the parasitic protozoan Syndinium sp.

Syndinium and related organisms which parasitize a number of invertebrates have been classified with dinoflagellates on the basis of the morphology of their zoospores. We demonstrate here that with respect to chromosome structure and chemistry as well as nuclear division, they differ fundamentally from free-living dinoflagellates. Alkaline fast green staining indicates the presence of basic proteins in Syndinium chromosomes. Chromatin fibers are about 30 A thick and do not show the arrangement characteristic of dinoflagellate chromosomes. The four V-shaped chromosomes are permanently attached at their apexes to a specific area of the nuclear membrane through a kinetochore-like trilaminar disk inserted into an opening of the membrane. Microtubules connect the outer dense layer of each kinetochore to the bases of the two centrioles located in a pocket-shaped invagination of the nuclear envelope. During division kinetochores duplicate, and each sister kinetochore becomes attached to a different centriole. As the centrioles move apart, apparently pushed by a bundle of elongating microtubules (central spindle), the daughter chromosomes are passively pulled apart. During the process of elongation of the central spindle, the cytoplasmic groove on the nuclear surface which contains the central spindle sinks into the nuclear space and is transformed into a cylindrical cytoplasmic channel. A constriction in the persisting nuclear envelope leads to the formation of two daughter nuclei.

Animals↗
PubMed1972

Characterization of insoluble protein fractions of mitochondria from Saccharomyces cerevisiae.

Saccharomyces cerevisiae was grown in a chemostat in the presence of excess oxygen. Cells harvested from fully derepressed and strongly repressed steady states show typical promitochondria-like structures under conditions of strong repression. Insoluble membrane proteins were extracted from highly purified mitochondria and submitted to isoelectric focusing in 6% polyacrylamide gels. Some 20 protein bands were obtained from derepressed cells. The pattern was clearly different (quantitatively and possibly qualitatively) from repressed mitochondria. In contrast to ribosomal proteins, insoluble membrane protein fractions were found in the acid section (pH 4 to 6.8) of the ampholyte gels. It can be concluded that glucose repression plays a prominent role in the synthesis of the functional mitochondrial membranes.

Aerobiosis↗
PubMed1969

Mapping of deletions and substitutions in heteroduplex DNA molecules of bacteriophage lambda by electron microscopy.

Electron microscopy of heteroduplex DNA molecules, composed of one strand of Escherichia coli phage lambda(+) DNA annealed to the complementary DNA strand of a lambda deletion or substitution mutant, permits visualization, as well as precise measurements and mapping, of the unpaired single-stranded regions of nonhomology in the otherwise double-stranded molecules. In the lambdab2 mutant, the central segment (13 percent) of the lambda(+) DNA molecule is shown to be deleted. In the hybrid phages lambda(i434) and lambda(i21) a segment of the right arm of the lambda(+) genome (5.5 or 7.6 to 9 percent) is replaced by the corresponding immunity regions of phage 434 (3.3 percent or phage 21 (4 percent) DNA. The b5 region in the lambdab5 mutant appears to be identical to the i(21) segment. From these data it is possible to estimate the size and posiion of those lambda genes which are replaced by the i(434) and i(21) segments. The method permits preparing complete physical maps of viral genomes with a precision heretofore unattainable.

Centrifugation, Density Gradient↗
PubMed1969

Division in the dinoflagellate Gyrodinium cohnii (Schiller). A new type of nuclear reproduction.

Dinoflagellates are of interest because their chromosomes resemble the nucleoplasm of prokaryotes both chemically and ultrastructurally. We have studied nuclear division in the dinoflagellate Gyrodinium cohnii (Schiller), using cells obtained from cultures undergoing phasic growth. Electron micrographs of serial sections were used to prepare three-dimensional reconstructions of nuclei and chromosomes at various stages of nuclear division. During division, a complex process of invagination of the intact nuclear envelope takes place at one side of the nucleus and results in the formation of parallel cylindrical cytoplasmic channels through the nucleus. These invaginations contain bundles of microtubules, and each of the bundles comes to lie in the cytoplasm of a cylindrical channel. Nuclear constriction occurs perpendicular to these channels without displacement of the microtubules. There are no associations between chromosomes and the cytoplasmic microtubules. In dividing cells most chromosomes become V-shaped, and the apices of the V's make contact with the membrane surrounding cytoplasmic channels. It is proposed that the membrane surrounding cytoplasmic channels in the dividing nucleus may be involved in the separation of daughter chromosomes. Thus, dinoflagellates may resemble prokaryotes in the manner of genophore separation as well as in genophore chemistry and ultrastructure.

Cell Division↗
PubMed1967

Occurrence, isolation, and characterization of polyribosomes in yeast.

This report details the procedural requirements for preparing cell-free extracts of yeast rich in polyribosomes. This enabled us to demonstrate the occurrence of polyribosomes in yeast, to show their role in protein synthesis, and to devise methods for their resolution and isolation. When certain precautions are met (the use of log phase cells, rapidly halting cell growth, gentle methods of disruption, sedimentation through exponential density gradients, etc.), individual polyribosome size classes ranging up to the heptosome can be fractionated and separated from their nearest neighbors. Larger size classes are resolved partially among themselves, free of smaller polyribosomes. This was confirmed by extensive electron micrographic studies of material from the various fractions obtained upon density gradient centrifugation of yeast extracts. Modifications of the gradients and procedure should allow fractionation and isolation of the larger polyribosomes, including those containing polycistronic messages. Yeast polyribosomes are disaggregated to single ribosomes by longer term grinding, cell disruption by the French pressure cell, the Hughes press, or by incubation with dilute RNAse. Yeast polyribosomes are active in the incorporation of amino acids into polypeptide; the single ribosomes exhibit only slight activity. The latter activity is probably due to the presence of a small fraction of monosomes still containing mRNA. Poly-U stimulates amino acid incorporation only in the single ribosomes.

Cell Division↗
PubMed1967

How bacteriophage chi attacks motile bacteria.

Bacteriophage chi attaches to the filament of a bacterial flagellum by means of a tail fiber, but the ultimate receptor site for the phage is located at the base of the bacterial flagellum. Here, the phage injects its deoxyribonucleic acid into the bacterium, leaving the empty phage attached at the base. It is suggested that chi slides along the filament of the flagellum to the base, owing to the movement of the flagellum. The role of motility would thus be to provide for rapid adsorption of the phage by guiding the phage to the adsorption sites at the bases of the flagella. Bacteria whose motility has been strongly inhibited by cold or anaerobic conditions still adsorb chi at the filaments and bases of flagella if a high multiplicity is used. This indicates that direct collisions with the bases may also be possible. Bacteria must be flagellated in order for chi to attach, but only a short flagellum, perhaps only the flagellar base, is necessary.

Adsorption↗