INDUCTION OF PROPHASE IN INTERPHASE NUCLEI BY FUSION WITH METAPHASE CELLS

Fusion of an interphase cell with a metaphase cell results in profound changes in the interphase chromatin that have been called "chromosome pulverization" or "premature chromosome condensation" In addition to the usual light microscopy, the nature of the changes has been investigated in the present study with electron microscopy and biochemical techniques Metaphase and interphase cells were mixed and fused at 37°C by means of ultraviolet-inactivated Sendai virus. After cell fusion, morphological changes in interphase nuclei occurred only in binucleate cells which contained one intact set of metaphase chromosomes Irrespective of the nuclear stage at the time of cell fusion, the morphologic changes that occurred 5–20 min later simulated very closely a sequence of events that characterizes the normal G2-prophase transition. Radioautography revealed that, late in the process, substantial amounts of RNA and probably protein were transferred from the interphase nucleus into the cytoplasm of fused cells. Thus, the findings indicate the existence in metaphase cells of factor(s) which are capable of initiating biochemical and morphological events in interphase nuclei intrinsic to the normal mitotic process.

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INDUCTION OF NUCLEAR ENVELOPES AROUND METAPHASE CHROMOSOMES AFTER FUSION WITH INTERPHASE CELLS

The Journal of Cell Biology ◽

10.1083/jcb.51.1.104 ◽

1971 ◽

Vol 51 (1) ◽

pp. 104-115 ◽

Cited By ~ 15

Author(s):

Tatsuro Ikeuchi ◽

Mitsuyoshi Sanbe ◽

Herbert Weinfeld ◽

Avery A. Sandberg

Keyword(s):

Nuclear Envelope ◽

Sendai Virus ◽

Chinese Hamster ◽

Formation Factor ◽

Electron Microscopic ◽

Metaphase Chromosomes ◽

Interphase Nuclei ◽

Multinucleate Cells ◽

Interphase Cells ◽

Nuclear Envelope Formation

The process of cellular fusion induced by Sendai virus in Chinese hamster cells (Don line) afforded us the opportunity to study nuclear envelope formation around metaphase sets in the presence of interphase nuclei, when chromosome pulverization failed to occur in such multinucleate cells. Morphologically, the enveloped metaphase chromosomes resembled a normal telophase nucleus, though minor differences prompted us to call it telophase-like. Electron microscopic observations demonstrated that the membranes enveloping the chromosomes appeared to be identical with a normal nuclear envelope. The longer the cells were incubated with Colcemid before fusion, the higher was the number of cells with telophase-like nuclei and the lower the percentage of cells with pulverizations. Furthermore, the number of pulverizations bore a somewhat direct relationship to the ratio of metaphase to interphase nuclei in multinucleate cells, and the number of telophase-like nuclei was inversely proportional to this ratio. A hypothesis is advanced in which a balance between the activities of a chromosome pulverization factor and a nuclear envelope formation factor, the former in metaphase cells and the latter in interphase cells, is decisive as to the nature of morphologic events observed in virus-induced fused cells.

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Morphological changes in Ehrlich ascites tumor cells during the cell fusion reaction with HVJ (Sendai virus)

Experimental Cell Research ◽

10.1016/0014-4827(81)90089-6 ◽

1981 ◽

Vol 132 (1) ◽

pp. 125-136 ◽

Cited By ~ 15

Author(s):

Jeman Kim ◽

Yoshio Okada

Keyword(s):

Tumor Cells ◽

Cell Fusion ◽

Sendai Virus ◽

Morphological Changes ◽

Fusion Reaction ◽

Ehrlich Ascites Tumor ◽

Ehrlich Ascites Tumor Cells ◽

Ascites Tumor ◽

Ehrlich Ascites

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PROPHASING OF INTERPHASE NUCLEI AND INDUCTION OF NUCLEAR ENVELOPES AROUND METAPHASE CHROMOSOMES IN HELA AND CHINESE HAMSTER HOMO- AND HETEROKARYONS

The Journal of Cell Biology ◽

10.1083/jcb.62.1.104 ◽

1974 ◽

Vol 62 (1) ◽

pp. 104-113 ◽

Cited By ~ 22

Author(s):

Yoshitaka Obara ◽

Lee S. Chai ◽

Herbert Weinfeld ◽

Avery A. Sandberg

Keyword(s):

Nuclear Envelope ◽

Hela Cells ◽

Interphase Nucleus ◽

Chinese Hamster ◽

Binucleate Cell ◽

Metaphase Chromosomes ◽

Interphase Nuclei ◽

Multinucleate Cells ◽

Interphase Cells ◽

Nuclear Envelope Formation

Fusing human HeLa metaphase cells with HeLa interphase cells resulted within 30 min in either of two phenomena in the resultant binucleate cell: either prophasing of the interphase nucleus or formation of a normal-appearing nuclear envelope around the metaphase chromosomes. The frequency of either occurrence was strongly dependent on environmental pH. At pH's of 6.6–8.0, prophasing predominated; at pH 8.5 nuclear envelope formation predominated. Additionally, the frequencies of the two events in multinucleate cells depended on the metaphase/interphase ratio. When the ratio was 0.33 nuclear envelope formation predominated; when it was 2.0 prophasing predominated. In their general features, the results with fused HeLa cells resembled those reported earlier with fused Chinese hamster Don cells. However, the results provided an indication that between pH 6.6 and 8.0 the HeLa metaphase cells possessed a much greater capacity than the Don metaphase cells to induce prophasing. Fusion of Don metaphase cells with HeLa interphase cells or of Don interphase cells with HeLa metaphase cells at pH 8.0 resulted in nuclear envelope formation or prophasing in each kind of heterokaryon. As in the homokaryons, the frequencies of the two events in the heterokaryons depended on the metaphase/interphase ratio. The statistics of prophasing and nuclear envelope formation in the homo- and heterokaryon populations were consistent with the notion that disruption or formation of the nuclear envelope depends on the balance attained between disruptive and formative processes.

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Morphological changes in Ehrlich ascites tumor cells during the cell fusion reaction with HVJ (Sendai virus)

Experimental Cell Research ◽

10.1016/0014-4827(80)90056-7 ◽

1980 ◽

Vol 130 (1) ◽

pp. 191-202 ◽

Cited By ~ 7

Author(s):

Jeman Kim ◽

Yoshio Okada

Keyword(s):

Tumor Cells ◽

Cell Fusion ◽

Sendai Virus ◽

Morphological Changes ◽

Fusion Reaction ◽

Ehrlich Ascites Tumor ◽

Ehrlich Ascites Tumor Cells ◽

Ascites Tumor ◽

Ehrlich Ascites

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DNA Synthesis and Mitosis in Fused Cells

Journal of Cell Science ◽

10.1242/jcs.5.3.603 ◽

1969 ◽

Vol 5 (3) ◽

pp. 603-624

Author(s):

R. T. JOHNSON ◽

H. HARRIS

Keyword(s):

Cell Cycle ◽

Dna Synthesis ◽

Cell Fusion ◽

Sendai Virus ◽

Single Cells ◽

Multinucleate Cell ◽

Multinucleate Cells ◽

Binucleate Cells ◽

The Third ◽

Nuclear Events

When HeLa cells growing in asynchronous culture are fused together by inactivated Sendai virus, synchrony of DNA synthesis and mitosis is rapidly imposed in the resultant multinucleate cells, even though the single cells which fuse to form these multinucleate cells are at different stages of the cell cycle. Some measure of nuclear co-ordination can already be observed within a few hours of cell fusion, and maximal synchrony is achieved within 2 days. By the end of the second day after fusion, asynchronous DNA synthesis or mitosis is rare. In binucleate cells this high order of synchrony is maintained for at least 5 days, but in cells containing greater numbers of nuclei some loss of synchrony begins to appear after the third day. The results indicate that the ability of the multinucleate cell to co-ordinate nuclear events is not impaired by the use of virus to facilitate cell fusion.

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Simultaneous genotypic and immunophenotypic analysis of interphase cells using dual-color fluorescence: a demonstration of lineage involvement in polycythemia vera

Blood ◽

10.1182/blood.v80.4.1033.1033 ◽

1992 ◽

Vol 80 (4) ◽

pp. 1033-1038 ◽

Cited By ~ 53

Author(s):

CM Price ◽

EJ Kanfer ◽

SM Colman ◽

N Westwood ◽

AJ Barrett ◽

...

Keyword(s):

Polycythemia Vera ◽

Fluorescent In Situ Hybridization ◽

Chromosomal Abnormalities ◽

Myeloproliferative Disorders ◽

Chromosome 8 ◽

Dual Fluorescence ◽

Interphase Cell ◽

Molecular Alterations ◽

Interphase Cells

Abstract Fluorescent in situ hybridization has become a useful technique by which chromosomal abnormalities may be shown in interphase cells. We present a dual-fluorescence method whereby a chromosomal and immunophenotypic marker can be visualized simultaneously in the same interphase cell. Two patients with the myeloproliferative disorder polycythemia vera and trisomy for chromosome 8 have been studied using this technique and selective involvement of the myeloid and erythrocyte lineages has been shown by the detection of the trisomy in immunophenotyped cells. Simultaneous analysis of genotype and immunophenotype in individual cells from patients with myeloproliferative disorders or leukemia may help identify the developmental and lineage status of cells in which molecular alterations have resulted in clonal advantage.

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Quantitative cell fusion: the fusion sensitivity (FS) potential

Journal of Cell Science ◽

10.1242/jcs.49.1.87 ◽

1981 ◽

Vol 49 (1) ◽

pp. 87-97

Author(s):

D. Rohme

Keyword(s):

Cell Line ◽

Cell Fusion ◽

Cell Lines ◽

Dose Response ◽

Sendai Virus ◽

Biological Significance ◽

Diploid Cell ◽

Mammalian Cell Lines ◽

Virus Dose ◽

A Cell

The dose response of Sendai virus-induced cell fusion was studied in 10 mammalian cell lines, comprising 5 continuous and 5 diploid cell lines originating from 5 species. The extent of fusion was calculated using a parameter directly proportional to the number of fusion events (t-parameter). At lower levels of fusion the dose response was found to be based on the same simple kinetic rules in all cell lines and was defined by the formula: t = FS. FAU/(I + FS. FAU), where FS (fusion sensitivity) is a cell-specific constant of the fusion rate and FAU (fusion activity units) is the virus dose. The FS potential of a cell line was determined as the linear regression coefficient of the fusion index (t/(I - t)) on the virus dose. At higher levels of fusion, when the fusion extent reached cell-line-specific maximal levels, the dose response was not as uniform. In general, and particularly in the cases of the diploid cell lines, these maximal levels were directly proportional to the FS potentials. Thus, it was concluded that the FS potential is the basic quantitative feature, which expresses the cellular fusion efficiency. The fact that FS varied extensively between cell lines, but at the same time apparently followed certain patterns (being higher in continuous compared to diploid cell lines and being related to the species of origin of the cells), emphasizes it biological significance as well as its possible usefulness in studies of the efficiency of various molecular interactions in the cell membrane/cytoskeleton system.

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The Synthesis and Migration of Nuclear Proteins during Mitosis and differentiation of Cells in Rats

Journal of Cell Science ◽

10.1242/jcs.s3-106.75.229 ◽

1965 ◽

Vol s3-106 (75) ◽

pp. 229-240

Author(s):

R. T. SIMS

Keyword(s):

Granular Cell ◽

Nuclear Proteins ◽

Photographic Emulsion ◽

Metaphase Chromosomes ◽

Interphase Nuclei ◽

Cell Layers ◽

The Difference ◽

Mitotic Figures ◽

And Migration ◽

Silver Grains

Hooded rats were given an intraperitoneal injection of 3H-tyrosine, and killed in pairs 10 min, 30 min, 12 h, 36 h, 7 days, and 30 days later. A piece of skin with white growing hair, and the tongue, were taken from each animal and radioautographs were prepared. Silver grains were counted over whole nuclei and whole mitotic figures of the germinal cells and whole nuclei of differentiating cells of both tissues. It was found that the interphase nuclei have significantly more silver grains over them than the chromosomes at all stages of mitosis and there are virtually no grains over metaphase, anaphase, and early telophase chromosomes in both tissues of all the animals killed up to 36 h after the injection. The difference between the grain counts over the interphase nuclei and the chromosomes of dividing cells is at least 20-fold at 30 min in the hair matrix, at least 5-fold at 30 min in the tongue and at 36 h in both tissues. It was established that the differences observed between the radioactivities of the nuclei and chromosomes of mitotic figures are real from estimates of: the radioactivity of the cell cytoplasm, volumes of the metaphase chromosomes and interphase nuclei within 1µ of the photographic emulsion, and the volumes of cytoplasm separating the photographic emulsion and these structures. No protein synthesis was demonstrable in the chromosomes during metaphase, anaphase, and early telophase. Nuclear proteins leave the chromosomes during prophase and prometaphase and return to the nucleus during late telophase. The cells in the matrix and upper bulb of the growing hair follicle and those in the germinal, prickle, and granular cell layers of the tongue are in different functional states; 30 min after injection of 3H-tyrosine they have different amounts of it in their nuclear proteins. It is suggested that the amount incorporated into each nucleus is related to the rate at which proteins are being synthesized by the cell.

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Studies of membrane fusion. VI. Mechanism of the membrane fusion and cell swelling stages of Sendai virus-mediated cell fusion

Journal of Cell Science ◽

10.1242/jcs.43.1.103 ◽

1980 ◽

Vol 43 (1) ◽

pp. 103-118

Author(s):

S. Knutton

Keyword(s):

Membrane Fusion ◽

Cell Fusion ◽

Sendai Virus ◽

Freeze Fracture ◽

Viral Envelope ◽

Cell Swelling ◽

Fusion Event ◽

Virus Envelope ◽

Membrane Rupture ◽

Membrane Tubules

The membrane fusion and cell swelling stages of Sendai virus-mediated cell-cell fusion have been studied by thin-section and freeze-fracture electron microscopy. Sites of membrane fusion have been detected in human erythrocytes arrested at the membrane fusion stage of cell fusion and in virtually all cases a fused viral envelope or envelope components has been identified thus providing further direct evidence that cell-viral envelope-cell bridge formation is the membrane fusion event in Sendai virus-induced cell fusion. Radial expansion of a single virus bridge connecting 2 cells is sufficient to produce a fused cell. Membrane redistribution which occurs during this cell swelling stage of the fusion process is often accompanied by the formation of a system of membrane tubules in the plane of expansion of the virus bridge. The tubules originate from points of fusion between the bridging virus envelope and the erythrocyte membrane and also expand radially as cells swell. Ultimately membrane rupture occurs and the tubules appear to break down as small vesicles. When previously observed in cross-sectioned cells these membrane tubules were interpreted as sites of direct membrane fusion. The present study indicates that this interpretation is incorrect and shows that the tubules are generated subsequent to membrane fusion when 2 cells connected by a virus bridge are induced to swell. A mechanism to explain the formation of this system of membrane tubules is proposed.

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