Adrienne Ficq

BINDING SITES OF 14C-ACTINOMYCIN IN AMPHIBIAN OVOCYTES AND AN AUTORADIOGRAPHY TECHNIQUE FOR THE DETECTION OF CYTOPLASMIC DNA.

Abstract An autoradiography method based upon the affinity of actinomycin D for the intact DNA molecule has been developed. The results show the presence of DNA within the cytoplasm (yolk) of the large ovocytes.

Biochemical studies of nucleate and anucleate fragments isolated from sea-urchin eggs: A comparison between fertilization and parthenogenetic activation

Abstract Nucleate and anucleate fragments of sea-urchin eggs as well as whole eggs were given RNA and protein precursors in the presence of puromycin and actinomycin. A comparison was made between events occurring after fertilization and after parthenogenetic activation.

Incorporationin vitro de glycocolle-I-14C dans les oocytes d'Astéries

An autoradiographic method, based on the observation of individual particles, was used to determine the relative specific radioactivity of RNA and of the proteins in the different parts of the oocytes ofAsterias rubens. The nucleolus incorporates glycine in both fractions about 100 times more rapidly than the cytoplasm.

The uptake of [8-14C]adenine and [2-14C]phenylalanine by rat-liver nuclei in vitro

Abstract 1. 1. Normal rat liver nucleic have been shown to take up [8- 14 C]adenine and [2- 14 C]phenylalanine in vitro . 2. 2. Nuclei, prepared from regenerating rat liver 24, 48 and 72 hours after partial hepatectomy, take up labelled adenine and phenylalanine in vitro . The nucleic acid metabolism of the isolated nuclei of regenerating rat liver recovers much more rapidly from the effects of partial hepatectomy than does the protein metabolism. 3. 3. The cytoplasmic fractions of rat liver enhance the uptake of adenine and phenylalanine. The microsomes appear to influence the uptake of adenine by isolated nuclei, while the uptake of phenylalanine appears to be influenced by the mitochondria.

Métabolisme des acides nucléiques dans l'oeuf d'oursin en développement. Étude autoradiographique

Abstract Autoradiographic experiments undertaken with 3H-thymidine, 3H-uridine and 3H-cytidine in order to follow nucleic acid metabolism in developing sea urchin eggs have shown that, during the first steps (cleavage) of development, a mechanism of rapid DNA synthesis must exist in the egg. The metabolically active RNA, strictly restricted to the nucleus, that we have demonstrated in these young stages could be a “messenger RNA”.

The effect of X-rays and ultraviolet light on the uptake in vitro of [8-14C]adenine and [2-14C]-phenylalanine by isolated nuclei

Abstract 1. 1. The uptake of [8-14C]adenine and [2-14C]phenylalanine into isolated calfthymus nuclei have been shown to be inhibited by X-irradiation. 2. 2. X-irradiation of isolated calf-thymus nuclei inhibited the uptake of [14C]adenine into the nuclear RNA while the uptake into the DNA was unaffected. 3. 3. X-irradiation of isolated rat liver nuclei and/or microsomes inhibited the uptake of [14C]adenine by the nuclei. u.v. irradiation of the nuclei and/or the microsomes, in this system, gave rise to an inhibition of adenine uptake which was reversed by incubating in daylight. 4. 4. X-irradiation of isolated rat-liver nuclei and/or mitochondria inhibited the uptake of [14C]phenylalanine by the nuclei. u.v. irradiation of the nuclei and/or the microsomes, in this system, gave rise to an inhibition of phenylalanine uptake which was reversed by incubating in daylight.

Synthesis and detection of DNA in early oogenesis

Tm aim of the present work was, on one hand, to identify, by 3H-thymidine labelling, the different types of ovarian cells in which DNA synthesis takes place in very young Xenopus laevis females and, on the other hand, to localize the DNA in these early stage oocytes, by applying the method of “staining” with radioactive actinomycin we have described elsewhere [3, 41. The animals analized, numbering 6, were aged from 2 to 3 months, that is just at the beginning of metamorphosis or 3, 4, 5, 6 and 7 weeks later. Methyl 3H-thymidine (The Radiochemical Centre, Amersham, England) of specific activity 500 mC/mM was administered intraperitoneally, using 30 ,uC per female. The animals were sacrificed 24 h after injection of the radioisotope. Ovaries were fixed by freeze-substitution and were sectioned 8 p thick. Some preparations were submitted to prior digestion with DNAase (Worthington, 0.1 mg/ml +MgCl, M/300, at 37°C during 1 h). The “staining” of unlabelled sections of ovaries with 3H-actinomycin (Schwarz, Bioresearch, specific radioactivity 2.6 C/m&‘) was carried out in the dark, during 1 h, in a solution having a radioactivity of 5 &/ml; this was followed by washing with cold actinomycin (10 pm/ml) for 30 min in the dark, then by washing with running water for 12 h. The classical autoradiographical procedure [8] was applied to both types of preparations, both being developed after 15 days’ exposure. The autoradiographs show that: (1) Thymidine incorporation into nuclear DNA takes place in the ovaries of the female which underwent metamorphosis 4 to 7 weeks before. The labelling is found, essentially in the prophase figures of premeiotic cells and in the nuclei of the smallest oocytes (these having a diameter of 30 to 50 ,u) (Fig. 1). The germinal vesicles of larger oocytes remain unlabelled. Preparations treated with DNAase show no trace of radioactivity. (2) The binding of actinomycin occurs in the follicular cells nuclei. Actinomycin is also strongly fixed in the nuclei of the very young oocytes, having a cell diameter of 30 to 150 EC, in contrast with what is found in the germinal vesicles of mature oocytes [4]. Radioactivity diminishes regularly during the increase of volume of the germinal vesicle. The activity is condensed around the nucleoli and it probably originates from the chromatin associated with the nucleolus (Fig. 2). In the nuclear sap, it seems closely related to structures which probably correspond to the lampbrush chromosomes at the beginning of their expansion (Fig. 3). At this stage, no significant labelling by actinomycin can be found in the cytoplasm.

Analyse de l'induction neurale par autoradiographie

Neural induction is accompanied by a migration of granules from the organizer to the reacting ectoblast (Brachet). The radioautographs of embryos or explants of amphibians seem to show that such a migration to the induced organs takes place when a marked organizer is grafted in a non-radioactive host. Substances of high molecular weight (proteins or nucleic acids) diffuse from an organizer tagged either with radioactive glycine or with radioactive orotic acid.

Cytochemical studies on the oogenesis of Dytiscus marginalis L

Abstract In a brief description of the oogenesis of Dytiscus marginalis, attention is focused on the role and the significance of the Giardina body and the nurse cells, in this organism. A staining technique, with 3H-actinomycin followed by autoradiography, very sensitive for detecting acid-labile DNA has confirmed, with more details, the observations made after Feulgen staining: 1. 1. During the early stages, the Giardina body and the nuclei of the nurse and the follicle cells, bind 3H-actinomycin intensely. 2. 2. At more advanced stages, the nuclei of the nurse and of the follicle cells remain heavily labelled by 3H-actinomycin. Although these structures are the only ones to be Feulgen positive at that stage, after staining with 3H-actinomycin, the germinal vesicle of the oocyte is also labelled, and more weakly, the ooplasma. 3. 3. Feulgen positive granules are visible in the cytoplasm, surrounding the nuclear membrane. They are radioactive after staining with 3H-actinomycin and electron microscopy suggests a passage of nuclear material through the nuclear membrane. These results are discussed in the light of our present knowledge on the role of extra chromosomal DNA and of cytoplasmic DNA.

Metabolic processes in cell nuclei

Abstract The incorporation, as observed in radioautographic studies, of precursors of proteins and nucleic acids into the cell nucleus has been shown to occur both in whole organisms (embryonic cells, normal and regenerating liver); and in isolated thymus and liver nuclei. The significance of these phenomena and their dependency on nuclear or cytoplasmic energy sources has been discussed.