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A Komamine

Publications and source records attributed to A Komamine.

At least 37 records · Page 2Linked to original sources
PubMed1992

A gene family homologous to the S-phase specific gene in higher plants is essential for cell proliferation in Saccharomyces cerevisiae.

Previously we reported the isolation and characterization of the gene, cyc07, which was specifically expressed in the S phase during the cell cycle in synchronous cell division cultures of the higher plant, Catharanthus roseus. We found that the yeast Saccharomyces cerevisiae contains two closely related genes which show a high degree of similarity (about 64% at the amino acid level) to cyc07 of C. roseus. Site-directed disruption mutations demonstrated that the two yeast genes, homologous to cyc07, constitute an essential gene family for cell proliferation in yeast cells. Furthermore, the rate of cell proliferation varied with the gene copy number.

Amino Acid Sequence↗
PubMed1991

Molecular cloning of the gene for plant proliferating-cell nuclear antigen and expression of this gene during the cell cycle in synchronized cultures of Catharanthus roseus cells.

A cDNA library was screened for plant proliferating-cell nuclear antigen (PCNA) from Catharanthus roseus (periwinkle). A lambda gt11 cDNA library was constructed using poly(A)-rich RNA isolated from the cells in the S phase. A cDNA clone for PCNA was isolated by using a rice genomic clone, pCJ-1, which contains PCNA-related gene sequences. The cDNA contains an open reading frame of 804 nucleotides, encoding a protein of 268 amino acids with a molecular mass of 29,765 Da. When conservative substitutions were included, a high degree of similarity (about 85%) was observed between the predicted amino acid sequence of periwinkle PCNA and that of human PCNA. Expression of mRNA for periwinkle PCNA was undetectable or very weak in quiescent cells, such as phosphate-starved cells, auxin-starved cells and cells in the stationary phase. In the synchronous progression of the cell cycle induced by the addition of phosphate or auxin, the active accumulation of periwinkle PCNA mRNA was observed preferentially in the S phase. When an inhibitor of DNA synthesis, aphidicolin, was added to the cells at the G1 phase, an increase in the level of PCNA mRNA was observed. The partial inhibition of protein synthesis at the G1 phase by a protein inhibitor, anisomycin, caused the arrest of cells in the G1 phase. No increase of the level of periwinkle PCNA mRNA was observed in cells arrested at the G1 phase by the inhibition of protein synthesis. These results indicate that the induction of mRNA for periwinkle PCNA occurred independently of the initiation of DNA replication, but that synthesis of certain proteins at the G1 phase was required for the induction of periwinkle PCNA mRNA at the S phase.

Amino Acid Sequence↗
PubMed1991

Isolation of Genes that Are Preferentially Expressed at the G(1)/S Boundary during the Cell Cycle in Synchronized Cultures of Catharanthus roseus Cells.

A cDNA library was screened for genes that may be involved in the progression of the cell cycle of cells of higher plants. The Catharanthus roseus L. (G) Don. cells were synchronized by the double phosphate starvation method, and a lambdagt11 cDNA library was prepared using poly(A)(+) RNA from cells in the S phase of the cell cycle. Two independent sequences, cyc02 and cyc07, were identified by differential screening. The levels of cyc02 and cyc07 mRNAs increased dramatically, but transiently, at the G(1)/S boundary of the cell cycle. High levels of cyc02 mRNA, but not of cyc07 mRNA, were also present in cells arrested at the G(1) phase by phosphate starvation. In an asynchronous batch culture, cyc02 and cyc07 mRNAs accumulated transiently at different stages of the growth cycle, cyc02 mRNA early in the stationary phase, and cyc07 mRNA in the midlogarithmic phase. When the proliferation of cells was arrested by nutrient starvation, i.e. by sucrose or nitrogen starvation, the relative amounts of the cyc02 and cyc07 mRNAs decreased. These results indicate that cyc02 and cyc07 contain nucleotide sequences from growth-related genes. The analysis of nucleotide sequence of cyc02 shows that the predicted product of this gene is basic and is composed of 101 amino acids. No significant homology to other known proteins was detected.

Journal Article↗
PubMed1991

Polyamines and the Cell Cycle of Catharanthus roseus Cells in Culture.

Investigation was made on the effect of partial depletion of polyamines (PAs), induced by treatment with inhibitors of the biosynthesis of PAs, on the distribution of cells at each phase of the cell cycle in Catharanthus roseus (L.) G. Don. cells in suspension cultures, using flow cytometry. More cells treated with inhibitors of arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) were accumulated in the G(1) phase than those in the control, while the treatment with an inhibitor of spermidine (SPD) synthase showed no effect on the distribution of cells. The endogenous levels of the PAs, putrescine (PUT), SPD, and spermine (SPM), were determined during the cell cycle in synchronous cultures of C. roseus. Two peaks of endogenous level of PAs, in particular, of PUT and SPD, were observed during the cell cycle. Levels of PAs increased markedly prior to synthesis of DNA in the S phase and prior to cytokinesis. Activities of ADC and ODC were also assayed during the cell cycle. Activities of ADC was much higher than that of ODC throughout the cell cycle, but both activities of ODC and ADC changed in concert with changes in levels of PAs. Therefore, it is suggested that these enzymes may regulate PA levels during the cell cycle. These results indicate that inhibitors of PUT biosynthesis caused the suppression of cell proliferation by prevention of the progression of the cell cycle, probably from the G(1) to the S phase, and PUT may play more important roles in the progression of the cell cycle than other PAs.

Journal Article↗
PubMed1991

Identification of a novel S-phase-specific gene during the cell cycle in synchronous cultures of Catharanthus roseus cells.

The cell-cycle specific cDNAs were isolated from a cDNA library prepared from cells in the S phase in the synchronous cultures of Catharanthus roseus. One of the isolated genes, which we refer to as cyc07, was analyzed in detail. The full-length cDNA of cyc07 contains an open reading frame of 735 nucleotides, encoding a protein of 245 amino acids with a molecular weight of 28,356 Da. The protein predicted from the nucleotide sequence is highly basic, as are mammalian histones. cyc07 mRNA was detected specifically in cells at the S phase in synchronous cultures. The induction and accumulation of mRNA in the S phase were suppressed when DNA synthesis was inhibited by aphidicolin. In the intact plant, cyc07 mRNA was found preferentially in root tips that contained meristematic tissue. A databank search revealed that a sequence homologous to the nucleotide sequence of cyc07 cDNA is present in the downstream region of the SIR3 gene in the yeast genome. The amino acid sequence predicted from the corresponding region of the yeast genome exhibited significant homology with that of cyc07 protein. These similarities between cyc07 and the corresponding region in yeast suggest that the homologous sequence in yeast is a novel gene that is functionally homologous to cyc07. Our results presented here suggest the possibility that cyc07 may play a role in the proliferation of higher plant cells, in particular in the entry into or progression of the S phase of the cell cycle.

Amino Acid Sequence↗
PubMed1989

Phase-Specific Polypeptides and Poly(A) RNAs during the Cell Cycle in Synchronous Cultures of Catharanthus roseus Cells.

This study shows an overall analysis of gene expression during the cell cycle in synchronous suspension cultures of Catharanthus roseus cells. First, the cellular cytoplasmic proteins were fractionated by two-dimensional gel electrophoresis and visualized by staining with silver. Seventeen polypeptides showed qualitative or quantitative changes during the cell cycle. Second, the rates of synthesis of cytoplasmic proteins were also investigated by autoradiography by labeling cells with [(35)S]methionine at each phase of the cell cycle. The rates of synthesis of 13 polypeptides were found to vary during the cell cycle. The silverstained electrophoretic pattern of proteins in the G(2) phase in particular showed characteristic changes in levels of polypeptides, while the rates of synthesis of polypeptides synthesized during the G(2) phase did not show such phase-specific changes. This result suggests that posttranslational processing of polypeptides occurs during or prior to the G(2) phase. In the G(1) and S phases and during cytokinesis, several other polypeptides were specifically synthesized. Finally, the variation of mRNAs was analyzed from the autoradiograms of in vitro translation products of poly(A)(+) RNA isolated at each phase. Three poly(A)(+) RNAs increased in amount from the G(1) to the S phase and one poly (A)(+) RNA increased preferentially from the G(2) phase to cytokinesis.

Journal Article↗
PubMed1985

Purification and some properties of chalcone synthase from a carrot suspension culture induced for anthocyanin synthesis and preparation of its specific antiserum.

Chalcone synthase was purified to homogeneity by polyacrylamide gel electrophoresis from cell suspension cultures of carrot in which anthocyanin synthesis was induced by transferring the cells from a medium containing 2,4-dichlorophenoxy-acetic acid (2,4-D) to one lacking it. A molecular weight of 80,000-85,000 for the enzyme was determined by gel filtration and disc-gel polyacrylamide electrophoresis, and one of about 40,600 for the subunit by SDS slab-gel electrophoresis. The primary reaction product was chalcone and the pH optimum of the reaction was 8.0. The Km values for 4-coumaroyl-CoA and malonyl-CoA were 5.7 microM and 18 microM, respectively. These properties of carrot chalcone synthase were discussed in comparison to those of that from cell cultures of parsley reported previously. Antiserum against chalcone synthase from carrot was obtained from mice bred under specific pathogen free conditions. Crossreactivity was examined by Western-blotting, and the high specificity of the antiserum against chalcone synthase was demonstrated.

Acyltransferases↗
PubMed1985

Identification and Isolation of Single Cells that Produce Somatic Embryos at a High Frequency in a Carrot Suspension Culture.

A system was established in which single cells differentiated to embryos at a high frequency. Small spherical single cells from a carrot (Daucus carota L. cv "Kurodagosun") cell suspension culture were obtained by fractionation through sieving, using nylon screens and then density gradient centrifugation in Percoll solutions. Eighty-five to 90% of these small single cells differentiated to embryos when they were cultured in a medium containing 2,4-dichlorophenoxyacetic acid (5 x 10(-8) molar), zeatin (10(-6) molar), and mannitol (0.2 molar) for 7 days, followed by transfer to a medium containing zeatin (10(-7) molar) but no auxin. This indicates that there are at least two phases in the differentiation of embryos from single cells. The progression of the first phase required exogenous auxin, whereas that of the second phase was inhibited by the same growth regulator. The relationship between the morphology and potency for embryogenesis from single cells was discussed. The system established here is a useful one for investigation of differentiation process from a single cell to a whole plant via embryogenesis, especially in its early stage.

Journal Article↗
PubMed1980

Establishment of an Experimental System for the Study of Tracheary Element Differentiation from Single Cells Isolated from the Mesophyll of Zinnia elegans.

Single cells were isolated mechanically from the mesophyll of adult plants and of seedlings of Zinnia elegans L. cv. Canary bird. When single cells isolated from the first leaves of seedlings were cultured in a liquid medium in the dark with rotation, they differentiated to tracheary elements with a reasonable degree of synchrony in the 24-hour period between days 2 and 3 after culture. The proportion of tracheary elements as a percentage of total cells reached nearly 30% 3 days after culture. Factors favoring cytodifferentiation were certain optimum levels of both alpha-naphthalene-acetic acid (0.1 milligram per liter) and benzyladenine (1 milligram per liter), a low concentration of ammonium chloride (0 to 1 millimolar), and an initial cell population density in the range 0.4 to 3.8 x 10(5) cells/ml. It was possible to follow analytically the sequence of cytodifferentiation in individual cells in this system.

Journal Article↗
PubMed1980

Direct Evidence for Cytodifferentiation to Tracheary Elements without Intervening Mitosis in a Culture of Single Cells Isolated from the Mesophyll of Zinnia elegans.

A serial observation of the process of tracheary element differentiation from single cells isolated from the mesophyll of Zinnia elegans L. cv. Canary bird provided the first direct evidence for the cytodifferentiation without intervening mitosis. Percentage of the tracheary elements formed without cell division was about 60% of total tracheary elements formed on the 4th day of culture. The number of tracheary elements formed without intervening mitosis was not reduced in the presence of colchicine at the concentrations blocking cell division. These facts clearly indicate that cell division is not a prerequisite for tracheary element differentiation in this system.

Journal Article↗
PubMed1979

Synchronization of somatic embryogenesis in a carrot cell suspension culture.

Synchronization of somatic embryogenesis was achieved in a carrot (Daucus carota L. cv. "Kurodagosun") suspension culture by sieving the initial heterogeneous cell population, by density gradient centrifugation in Ficoll solutions, and by subsequent repeated centrifugations at a low speed (50g) for a short time (5 seconds), followed by transferring the cell clusters obtained, which were composed of 3 to 10 cells, to a medium containing zeatin (0.1 micromolar) but no auxin. The frequency of embryo formation reached more than 90%, and synchrony of the embryogenetic process was observed at least in the early stages of the process. The system established in the present work provides a useful system for biochemical research into the mechanisms of somatic embryogenesis.

Journal Article↗
PubMed1978

Change in the proportion of two aspartokinases in carrot root tissue in response to in vitro culture.

Two isofunctional aspartokinases (EC 2.7.2.4) exist in fresh root tissue of carrot (Daucus carota, cv. Oogata sanzun). The threoninesensitive portion constitutes about 70% of the activity; the lysinesensitive, less than 20%. Culture of slices of carrot tissue for 3 days reversed the ratio as the lysine-sensitive activity preferentially increased. Inhibition by threonine and lysine was additive in both enzyme preparations from fresh and cultured tissues. The activities were resolved into two distinct fractions of different sensitivity to threonine and lysine by DEAE-Sephadex A-50 column chromatography.

Journal Article↗