Comparisons of tests for aneuploidy

The fundamental problems that face us in the development of suitable assay systems for the detection of potentially aneugenic (aneuploidy-inducing) chemicals include: (a) the diversity of cellular targets and mechanisms where perturbations of structure and function may give rise to changes in chromosome number, and (b) the phylogenetic differences that exist between species in their mechanism and kinetics of cell division and their metabolic profiles. A diverse range of assay systems have been developed, which have been shown to have potential for use in the detection of either changes in chromosome number or of perturbations of the events which may be causal in the induction of aneuploidy.

Chromosome number changes may be detected cytologically by karyotypic analysis, or by the use of specialised strains in which aneuploid progeny may be observed due to phenotypic differences with aneuploid parental cells or whole organisms. Techniques for the detection of cellular target modifications range from in vitro studies of tubulin polymerisation to observations of the behaviour of various cellular organelles and their fidelity of action during the division cycle.

The diversity of mechanisms which may give rise to aneuploidy and the qualitative relevance of events observed in experimental organisms compared to man make it unlikely that the detection and risk assessment of the aneugenic activity of chemicals will be possible using a single assay system. Optimal screening and assessment procedures will thus be dependent upon the selection of an appropriate battery of predictive tests for the measurement of the potentially damaging effects of aneuploidy induction.     

Auxin-induced rapid changes in translatable mRNAs in tobacco cell suspension

When 2,4-dichlorophenoxyacetic acid (2,4-D)-dependent tobacco cell suspensions, one normal and one transformed by Agrobacterium tumefaciens, were subcultured on hormone-lacking medium the stationary phase of the cell cycle was reached earlier than on medium containing 2,4-D. Addition of the auxin 2,4-D could restore cell division activity within 10–12 h for the most rapidly reacting cell line. The cell-division response was characterized as being auxin-specific and optimal with 2,4-D at 2.2 10^-6 M. Although the cell lines used showed different characteristics, both reacted with a rapid increase in at least three mRNA species within 1 or 2 h after 2,4-D application. Two, 2,4-D-induced protein spots, seen after in-vitro translation, had the same characteristics (MWs 35 kilodaltons (kDa) and 25 kDa with isoelectric points of 7.1 and 6.3, respectively) in both cell lines. Water-treated controls did not show alterations in the translatable mRNA populations. This indicates that the accumulation of the corresponding mRNAs is an early hormone-induced event. Since cell division is the only measurable reaction found after auxin application, cell systems as described here offer excellent possibilities for studying early auxin-induced changes at the molecular level preceding mitosis.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - IAA indole-3-acetic acid - kDa kilodalton     

A new gene controlling the frequency of cell division per round of DNA replication in Escherichia coli

Summary A novel mutant of Escherichia coli, named cfcA1, was isolated from a temperature-sensitive dnaB42 strain, and found to have the following characteristics. Division arrest and lethality induced by inhibition of DNA replication was reduced and delayed in the cfcA1 dnaB42 strain, as compared with the parental dnaB42 strain. Two types of inhibition of division induced by the addition of nalidixic acid or hydroxyurea were suppressed by the cfcA1 mutation. Under permissive conditions for DNA replication, the colony forming ability of cfcA1 cells was significantly reduced as compared with that of cfc ⁺ cells; conversely the division rate of cfcA1 cells was higher than that of cfc ⁺ cells. The cfcA1 mutation partially restored division arrest induced in the thermosensitive ftsZ84 mutant at the restrictive temperature and suppresed the UV sensitivity of the lon mutation. The mutation was mapped at 79.2 min on the E. coli chromosome. Taking these properties into account, it is hypothesized that the cfcA gene is involved in determining the frequency of cell division per round of DNA replication by interacting with the FtsZ protein which is essential for cell division.     

Ultrastructure of vegetative organization and cell division in the freshwater red algaCompsopogon

Summary Uniseriate filaments of the freshwater red algaCompsopogon coeruleus were examined by transmission electron microscopy for details of vegetative organization and cell division with the goal of providing useful taxonomic characters. Each cell's single, complex chloroplast contains a peripheral encircling thylakoid, and unlike the vast majority of red algae, the cis-regions of dictyosomes are not consistently juxtaposed with mitochondria. These subcellular features, which are present in all examined genera in theCompsopogonales, Erythropeltidales, andRhodochaetales, along with certain unique reproductive characteristics, unify these three orders. During mitosis in uncorticated axial cells, a small, ring-shaped nucleus associated organelle (NAO) is located at each division pole, an intranuclear spindle comes to a moderately acute focus at the flattened, fenestrated metaphase-anaphase division poles and perinuclear ER partially encloses dividing nuclei, including a well-developed interzonal midpiece. The cleavage furrow penetrates the large, central vacuolar region to separate daughter nuclei. These cell division features most closely resemble the pattern described for the orderCeramiales. Our observations of vegetative and dividing cells ofC. coeruleus supplement the growing volume of evidence in favour of uniting all red algae into a single class without subclass designations.Abbreviations ER endoplasmic reticulum - IZM interzonal midpiece - MT microtubule - MTOC microtubule organizing center - NAO nucleus associated organelle - NE nuclear envelope - PER perinuclear endoplasmic reticulum     

Changes of Escherichia coli cell cycle parameters during fast growth and throughout growth with limiting amounts of thymine

It is generally accepted that during fast growth of Escherichia coli, the time (D) between the end of a round of DNA replication and cell division is constant. This concept is not consistent with the fact that average cell mass of a culture is an exponential function of the growth rate, if it is also accepted that average cell mass per origin of DNA replication (M_i) changes with growth rate and negative exponential cell age distribution is taken into account. Data obtained from cell composition analysis of E. coli OV-2 have shown that not only (M_i) but also D varied with growth rate at generation times () between 54 and 30 min. E. coli OV-2 is a thymine auxotroph in which the replication time (C) can be lengthened, without inducing changes in , by growth with limiting amounts of thymine. This property has been used to study the relationship between cell size and division from cell composition measurements during growth with different amounts of thymine. When C increased, average cell mass at the end of a round of DNA replication also increased while D decreased, but only the time lapse (d) between the end of a replication round and cell constriction initiation appeared to be affected because the constriction period remained fairly constant. We propose that the rate at which cells proceed to constriction initiation from the end of replication is regulated by cell mass at this event, big cells having shorter d times than small cells.Abbreviations OD₄₅₀ and OD₆₃₀ Optical density at a given wavelength in nm Dedicated to Dr. John Ingraham to honor him for his many contributions to Science     

Regulation of brood size in a burying beetle,Nicrophorus tomentosus (Silphidae)

Regulation of brood size in a biparental burying beetle, Nicrophorus tomentosusWeber, was studied by providing pairs with one of two sizes of mouse carcasses in the laboratory. For a given carcass size, there was an inverse relationship between number and mass offspring in a brood. The requirement for regulation was that brood size was adjusted such that mean mass of individual larvae was constant for carcasses of different size. Brood size was regulated if parents were present but regulation did not occur if parents were removed prior to hatching of larvae. Pairs bred in quick succession on two carcasses raised fewer than the regulated number of young in the second reproductive attempt. Reasons for regulation of brood size in this genus are discussed.     

Growth and reproduction of the mosquitofish,Gambusia affinis,in relation to temperature and ration level: consequences for life history

The allocation of energy to growth and reproduction, in relation to temperature and food availability, was investigated in laboratory experiments with the mosquitofish,Gambusia affinis. At constant temperature of 20, 25 and 30°C and ad libitum feeding, specific growth rates increased with increasing temperature at 1.7, 3.1 and 3.4% dry mass day⁻¹, respectively. Growth rates in a cycling temperature regime (20–30°C, ) were faster than in a 25°C constant temperature. As temperature increased from 20 to 30°C, mean age at first reproduction decreased from 191 to 56 days and brood size and mass of offspring increased significantly. Interbrood interval was also temperature dependent; estimates at 25 and 30°C for females >1000 mg were 22.6 and 18.6 days, respectively. Interbrood interval could not be calculated at 20°C. Although fitness was highest at 30°C, females at 25°C invested a greater proportion of surplus energy (growth and reproduction) to reproduction (38%) than at 20 (17%) or 30°C (36%) during the 32-week study. Fish at cooler temperatures began reproduction at a smaller size. Where rations were controlled at low, medium, and ad libitum levels, somatic and gonadal growth increased with increasing temperatures and food availability. The proportion of energy invested in reproduction was highest at 25°C for each comparable ration level. Calculated energy budgets indicated that over the 10-week study, 17–22% of the food energy was invested in growth, 0–7% in reproduction, and 75–83% in respiration and excretory losses, depending on feeding and temperature conditions.     

A variation in the structure of the protein-coding region of the human p53 gene

An extensive analysis of genomic DNA preparations from a number of normal and malignant tissues revealed BglII site polymorphism of the human p53 gene. Approximately 10% of p53 gene alleles were found to contain an additional BglII site localized in a region of intron I. This allelic form of p53 gene was also responsible for p53 protein having altered electrophoretic mobility. Molecular cloning and sequencing of both the alleles of p53 gene revealed a base-pair change in codon 72 causing arginine → proline substitution in the allele with the additional BglII site. Both variants of the p53 gene may occur in homozygous state and are therefore functional.     

A nuclear protein associated with cell divisions in plants

A nuclear protein, present in carrot meristems and rapidly proliferating cultured cells of carrot (Daucus carota L.) has been identified by the use of a monoclonal antibody (MAb 21D7). By combining the techniques of two-dimensional polyacrylamide gel analysis and blotting separated proteins onto nitrocellulose sheets, it was shown that the antibody detected a single polypeptide of apparent molecular mass (M _r) of 45000 and an isoelectric focusing point (pI) of 6.7. This protein was found by subcellular fractionation and immunofluorescence to be highly concentrated in the nucleoli of somatic and zygotic embryos of a wide range of plants. It was not detectable in logarthmically growing cells ofEscherichia coli, yeast, embryos ofDrosophila melanogaster or cultured C3H mouse cells. These data indicate that this protein is a highly conserved non-histone protein associated with nuclei of rapidly dividing plant cells.Abbreviations M _r apparent molecular mass - Da dalton - Ig immunoglobulins - MAb monoclonal antibody - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis - 2-D gel two-dimensional gel electrophoresis - 2,4-D 2,4-dichlorophenoxyacetic acid