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J D Berger

Publications and source records attributed to J D Berger.

At least 37 records · Page 2Linked to original sources
PubMed1978

Regulation of macronuclear DNA content in Paramecium tetraurelia.

The macronucleus of Paramecium divides amitotically, and daughter macronuclei with different DNA contents are frequently produced. If no regulatory mechanism were present, the variance of macronuclear DNA content would increase continuously. Analysis of variance within cell lines shows that macronuclear DNA content is regulated so that a constant variance is maintained from one cell generation to the next. Variation in macronuclear DNA content is removed from the cell population by the regulatory mechanism at the same rate at which it is introduced through inequality of macronuclear division. Half of the variation in macronuclear DNA content introduced into the population at a particular fission by inequality of division is compensated for during the subsequent period of DNA synthesis. Half of the remaining variation is removed during each subsequent cell cycle. The amount of variation removed in one cell cycle is proportional to the postfission variation. The cell's power to regulate DNA content is substantially greater than that required to compensate for the small differences that arise during division of wild-type cells. For example, a constant variance was still maintained when the mean difference between sister cells was increased to ten times its normal level in a mutant strain. The observations are consistent with a replication model that assumes that each cell synthesizes an approximately constant amount of DNA which is independent of the initial DNA content of the macronucleus. It is suggested that the amount of DNA synthesized may be largely determined by the mass of the cell.

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A cdc2-like kinase associated with commitment to division in Paramecium tetraurelia.

Cell division in higher eukaryotes is mainly controlled by p34cdc2 or related kinases and by other components of these kinase complexes. We present evidence that cdc2-like kinases also occur in Paramecium. Two polypeptides reacted with an antibody directed against the perfectly conserved PSTAIR region found in cdc2 kinases in other eukaryotes. Only the less abundant peptide bound to p13suc1 from Schizosaccharomyces pombe. Using centrifugal elutriation to select cells on the basis of size, we isolated highly synchronous Paramecium G1 cells. With this procedure, we demonstrated that the p13suc1-associated cdc2-like histone H1 kinase was activated before cell division at the point of commitment to division in Paramecium. Further, we show that Paramecium cdc2-like proteins occurred principally as monomers and that these monomers were active as histone H1 kinases in vitro.

Amino Acid Sequence↗
PubMedDate not supplied

Commitment to division in ciliate cell cycles.

Near the end of the cell cycle, ciliates commit irreversibly to cell division. The point of commitment occurs at the time of oral polykinetid assembly and micronuclear anaphase. The commitment is a checkpoint which requisites a threshold cell mass/DNA ratio and stomatogenesis. It is also a physiological transition point, involving cdk protein kinases similar to those of other eukaryotes. Both P34 kD and P36 kD kinases, similar to the S. pombe cdc2 kinases, have been described to have activity as monomers. Subsequent to commitment to division, dramatic cytoskeletal modifications occur for separation of organelles, cortex morphogenesis and cytokinesis. Numerous mutants affecting cytoskeletal function associated with the division process have been obtained in several species. Of these, only the cc1 mutant in Paramecium affects cell cycle progression prior to commitment to division. The material reviewed is used to speculate about the mechanisms of regulation of pre-fission morphogenesis and cell division related processes in ciliates.

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Two distinct classes of mitotic cyclin homologues, Cyc1 and Cyc2, are involved in cell cycle regulation in the ciliate Paramecium tetraurelia.

The eukaryotic cell cycle is regulated by the sequential activation of different CDK/cyclin complexes. Two distinct classes of mitotic cyclin homologues, CYC1 and CYC2, have been identified and cloned for the first time in the ciliate Paramecium. Cyc1 is 324 amino acids long with a predicted molecular mass of 38 kDa, whereas Cyc2 is 336 amino acids long with a predicted molecular mass of 40 kDa. They display 42-51% sequence identity to other eukaryotic mitotic cyclins within the 'cyclin box' region. The conserved 'cyclin box' and 'destruction box' elements can be identified within each of the sequences. Genomic Southern blot analysis indicated that the CYC1 gene has two isoforms, with 92.3% and 85.9% identify at the amino acid level and at the nucleotide level, respectively. Both Cyc1 and Cyc2 proteins showed characteristic patterns of accumulation and destruction during the vegetative cell cycle, with Cyc1 peaking at the point of commitment to division (PCD), and Cyc2 reaching the maximal level late in the cell cycle. Immunoprecipitation experiments with antibodies specific to Cyc1 and Cyc2 indicated that Cyc1 and Cyc2 associate with distinct CDK homologues. Both immunoprecipitates exhibited histone H1 kinase activity that oscillated in the cell cycle in parallel with the respective amount of cyclins present. Histone H1 kinase activity associated with Cyc1 reached a peak at PCD while Cyc2 showed maximal activity when about 75% cells have completed cytokinesis. We propose that Cyc1 may be involved in commitment to division, in association with the CDK that binds to p13suc1, Cdk3, and that the Cyc2/Cdk2 complex may regulate cytokinesis. PCR-amplification revealed similar sequences in Tetrahymena, Sterkiella, Colpoda and Blepharisma, suggesting the conservation of the cyclin genes within ciliates. Although cell cycle regulation in ciliates differs in some respects from that of other eukaryotes, the cyclin motifs have clearly been conserved during evolution.

Amino Acid Sequence↗
PubMedDate not supplied

A novel member of the cyclin-dependent kinase family in Paramecium tetraurelia.

Passage through the cell cycle in eukaryotes requires the successive activation of different cyclin-dependent protein kinases. Here, we describe the identification and characterization of a novel class of cyclin-dependent protein kinase, termed Cdk2, in the ciliate Paramecium tetraurelia. It is 301 amino acids long, 7 amino acids shorter than Cdk1, the CDK that is associated with macronuclear DNA synthesis. All the catalytic domains typical of protein kinases can be located within the sequence and putative regulatory phosphorylation sites equivalent to Thr14, Tyr15, and Thr161 in human CDK1 are also conserved. The 'PSTAIRE' region characteristic of most CDKs is perfectly conserved. Cdk2 shares only 48% homology to Cdk1 at the amino acid level, suggesting that the evolutionary separation of Cdk1 and Cdk2 is ancient, and implying that they have different roles in cell cycle regulation. Like Cdk1, Cdk2 does not bind to yeast p13suc1, even though it has better conservation of p13suc1 binding sites than Cdk1 does. The Cdk2 protein level is relatively constant throughout the vegetative cell cycle. Cdk2 exhibits kinase activity towards bovine histone H1 in vitro with the maximal level late in the cell cycle, suggesting it may be involved in the regulation of cytokinesis. Our results further support the view that an analogue of the cyclin-dependent kinase cell cycle regulatory system like that of yeast and higher eukaryotic cells operates in Paramecium and that a family of cyclin-dependent kinases may control different aspects of the Paramecium cell cycle.

Amino Acid Sequence↗
PubMedDate not supplied

Timing of life cycle morphogenesis in synchronous samples of Sterkiella histriomuscorum. II. The sexual pathway.

Isolates of Sterkiella (Oxytrichidae, Stichotrichia, Ciliata) are commonly used to study macronuclear development. These organisms respond to changes in food abundance variably by encystment-excystment, conjugation, cannibalism or rescaling cell size. An isolate of Sterkiella histriomuscorum (previously Oxytricha fallax and O. trifallax) is used because two complementary mating types are available. We provide observations on conjugation in cultures of this isolate. Using synchronous samples of conjugants, the timing of stages of nuclear divisions during conjugation was determined. Following ex-conjugant cultures over time, the onset of clonal aging and senescence is described. Cells become sexually mature after a brief period of "adolescence", during which time selfing is possible. Senescent cultures are less vigorous, unable to conjugate and encyst more readily. Excystment survival decreases with clonal age. These results can serve as reference for long-term cultures of this species and for analysing particular stages of developmental processes during conjugation.

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Riding the ciliate cell cycle--a thirty-five-year prospective.

Studies of the ciliate cell cycle have moved from early examination of its biochemistry with heat-synchronized Tetrahymena through descriptive studies of Paramecium using small synchronous cell samples. These studies described what happens during the cell cycle and provided some initial insights into control, especially the idea that there was a point at which cells became committed to division. This early work was followed by an analytical phase in which the same small sample techniques, combined with gene mutations, were used to tease apart some major features of the regulation of cell growth kinetics, including regulation of macronuclear DNA content and regulation of cell size, the control of timing of initiation of macronuclear DNA synthesis, and the control of commitment to division in Paramecium. The availability of new molecular genetic approaches and new means of manipulating cells en masse made it possible to map out some of the basic features of the molecular biology of cell cycle regulation in ciliates. The challenge before us is to move beyond the 'me-too-ism' of validating the presence of basic molecular regulative machinery underlying the cell cycle in ciliates to a deeper analysis of the role of specific molecules in processes unique to ciliates or to analysis of the role of regulatory molecules in the control of cell process that can be uniquely well studied in ciliates.

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