scholarly journals NUCLEOLI OF DIPLOID CELL STRAINS

1971 ◽  
Vol 49 (3) ◽  
pp. 785-802 ◽  
Author(s):  
Stephanie G. Phillips ◽  
David M. Phillips
Keyword(s):  
Ribosomal Rna ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Chinese Hamster ◽  
Diploid Cell ◽  
Particulate Material ◽  
Gradual Disappearance ◽  
Mouse Cells ◽  
Cell Strains ◽  
Nucleolar Components

Nucleoli of cultured Chinese hamster or mouse cells in early passages had a loosely reticular substructure. Within the reticulum small, irregularly shaped, light fibrillar zones occurred which were contiguous with denser fibrillar zones. These denser zones appeared to be connected in some places to the particulate material which composed the mass of the nucleolus. Generally, electron-transparent spaces separated the particulate zones from the fibrillar areas. Treatment with toyocamycin, an agent which is reported to cause a blockage in the processing of ribosomal RNA, greatly inhibited the accumulation of newly synthesized RNA in the cytoplasm, as monitored by radioautography. Toyocamycin treatment caused the gradual disappearance of the granules from the particulate region of the nucleoli, and resulted ultimately in the nucleoli appearing homogeneously fibrillar. Actinomycin D treatment, which inhibited virtually all RNA synthesis, caused a segregation, and finally a disaggregation, of nucleolar components.

scholarly journals SITES OF NUCLEOLUS PRODUCTION IN CULTURED CHINESE HAMSTER CELLS

1969 ◽  
Vol 40 (1) ◽  
pp. 248-268 ◽  
Author(s):  
Stephanie Gordon Phillips ◽  
David M. Phillips
Keyword(s):  
Fine Structure ◽  
Ribosomal Rna ◽  
Rna Synthesis ◽  
Chinese Hamster ◽  
Cell Types ◽  
Chinese Hamster Cell ◽  
Chinese Hamster Cells ◽  
Cell Strains ◽  
Almost All ◽  
Nucleolar Components

Chinese hamster cell strains in the early passages in culture display wide variation in number of nucleolus-like bodies per cell, though such strains are characteristically euploid. A variety of criteria indicate that the nucleolus-like bodies are true nucleoli. Their Azure B- and fast green-staining properties indicate the presence of RNA and protein; they have typical nucleolar fine structure, including both fibrous and granular components; radioautography reveals that their patterns of uptake of uridine-3H into RNA are similar to those reported for nucleoli of other cell types; actinomycin D, at a level which selectively inhibits ribosomal RNA synthesis, greatly reduces their RNA synthesis and also causes segregation of fibrous and granular nucleolar components. Colchicine was used to experimentally fragment the nuclei of these cells into a number of separate karyomeres, each presumably containing some, or only one, of the chromosomes of the complement. Almost all the karyomeres contain nucleolus-like bodies which, by the same criteria applied to the multiple nucleolus-like bodies of uninuclear cells, appear to be true nucleoli. The nucleoli of individual karyomeres of the same cell often differ from each other in fine structure while the multiple nucleoli of a uninuclear cell generally resemble each other. The evidence presented in this study indicates that Chinese hamster cells contain many nucleolus-producing sites scattered through the genome.

scholarly journals DISTINCTIVE CHARACTERISTICS OF NUCLEOLI OF TWO ESTABLISHED CELL LINES

1971 ◽  
Vol 49 (3) ◽  
pp. 803-815 ◽  
Author(s):  
David M. Phillips ◽  
Stephanie G. Phillips
Keyword(s):  
Cell Lines ◽  
Cultured Cells ◽  
Chinese Hamster ◽  
Mouse Cell ◽  
Particulate Material ◽  
Gradual Disappearance ◽  
Established Cell ◽  
Nucleolar Volume ◽  
Light Area ◽  
Nucleolar Components

Nucleoli of cultured cells of the established lines KB and L were found to possess a distinctive fine structural organization. The major portion of the nucleolar volume was composed of compact, particulate material. Spheroidal fibrillar zones about 0.4 µ in diameter occurred within the particulate mass. These fibrillar zones had a central light area and a denser rim. Toyocamycin treatment, which sharply inhibited the appearance of newly synthesized RNA in the cytoplasm, caused the gradual disappearance of the fibrillar material from nucleoli. Actinomycin D treatment, which inhibited virtually all RNA synthesis, caused varying types of segregation of nucleolar components. The morphology of nucleoli of KB and L cells and the reorganization of these nucleoli in response to drugs appear to be different from those of nucleoli of freshly initiated Chinese hamster and mouse cell lines.

scholarly journals REPOPULATION OF THE POSTMITOTIC NUCLEOLUS BY PREFORMED RNA

1972 ◽  
Vol 53 (3) ◽  
pp. 611-623 ◽  
Author(s):  
Stephanie Gordon Phillips
Keyword(s):  
Actinomycin D ◽  
Chinese Hamster ◽  
Mitotic Cell ◽  
Fast Green ◽  
Azure B ◽  
Mouse Cells ◽  
Nucleolar Rna ◽  
L929 Mouse ◽  
Nucleolar Components ◽  
Mitotic Cells

This study is concerned with the fate of the nucleolar contents, particularly nucleolar RNA, during mitosis Mitotic cells harvested from monolayer cultures of Chinese hamster embryonal cells, KB6 (human) cells, or L929 (mouse) cells were allowed to proceed into interphase in the presence or absence (control) of 0.04–0 08 µg/ml of actinomycin D, a concentration which preferentially inhibits nucleolar (ribosomal) RNA synthesis 3 hr after mitosis, control cells had large, irregularly shaped nucleoli which stained intensely for RNA with azure B and for protein with fast green. In cells which had returned to interphase in the presence of actinomycin D, nucleoli were segregated into two components easily resolvable in the light microscope, and one of these components stained intensely for RNA with azure B. Both nucleolar components stained for protein with fast green In parallel experiments, cultures were incubated with 0.04–0 08 µg/ml actinomycin D for 3 hr before harvesting of mitotic cells, then mitotic cells were washed and allowed to return to interphase in the absence of actinomycin D. 3 hr after mitosis, nuclei of such cells were devoid of large RNA-containing structures, though small, refractile nucleolus-like bodies were observed by phase-contrast microscopy or in material stained for total protein. These experiments indicate that nucleolar RNA made several hours before mitosis persists in the mitotic cell and repopulates nucleoli when they reform after mitosis

scholarly journals RNA SYNTHESIS IN CHINESE HAMSTER CELLS

1968 ◽  
Vol 36 (3) ◽  
pp. 583-593 ◽  
Author(s):  
M. D. Enger ◽  
R. A. Tobey ◽  
A. G. Saponara
Keyword(s):  
Ribosomal Rna ◽  
Rna Synthesis ◽  
Specific Activity ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Sedimentation Analysis ◽  
Ovary Cells ◽  
Chinese Hamster Cells ◽  
Mechanical Selection ◽  
Selection Of

The incorporation of methionine-methyl-14C into 18S ribosomal RNA of cultured Chinese hamster ovary cells in early and late interphase has been determined by zone-sedimentation analysis of phenol-extracted RNA preparations. Synchronized cell cultures were prepared for these studies by thymidine treatment and by mechanical selection of mitotic cells. The specific activity of 18S RNA labeled in late interphase was found to be 1.1–1.2 times that of 18S RNA labeled in early interphase. Upon correction for increase in RNA mass, the rate of methylation of 18S RNA in late interphase is about 1.9 times that in early interphase.

scholarly journals A comparative study of the effect of aflatoxin B1 and actinomycin D on HeLa cells

1969 ◽  
Vol 114 (2) ◽  
pp. 289-298 ◽  
Author(s):  
E. H. Harley ◽  
K. R. Rees ◽  
A. Cohen
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Rna ◽  
Mechanism Of Action ◽  
Aflatoxin B1 ◽  
Hela Cells ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Inhibitory Effect ◽  
Cytotoxic Effects ◽  
Rna Precursor

1. The cytotoxic effects of aflatoxin B1 on HeLa cells were examined and effects of short exposures of the cells to the toxin were found to be reversible. 2. Aflatoxin B1 inhibited the synthesis of both ribosomal and heterodisperse RNA. It is proposed that the toxin's mechanism of action on ribosomal RNA synthesis is related to its inhibitory effect on the maturation of the 45s-ribosomal-RNA precursor. 3. Protein synthesis is inhibited to a greater extent by aflatoxin B1 than by actinomycin D. In contrast with actinomycin D, aflatoxin B1 was shown to disaggregate polyribosomes directly.

Correlated inhibition of ribosomal RNA synthesis and silver staining by actinomycin D

1979 ◽  
Vol 47 (3) ◽  
pp. 329-333 ◽  
Author(s):  
F. J. Hofg�rtner ◽  
W. Krone ◽  
Kamlesh Jain
Keyword(s):  
Ribosomal Rna ◽  
Silver Staining ◽  
Rna Synthesis ◽  
Actinomycin D

scholarly journals SOME BIOCHEMICAL PROPERTIES OF CHINESE HAMSTER CELLS SENSITIVE AND RESISTANT TO ACTINOMYCIN D

1974 ◽  
Vol 63 (3) ◽  
pp. 773-779 ◽  
Author(s):  
Robert H. F. Peterson ◽  
Judith A. O'Neil ◽  
June L. Biedler
Keyword(s):  
Rna Synthesis ◽  
Actinomycin D ◽  
Chinese Hamster ◽  
Biochemical Properties ◽  
Metabolic Inhibitors ◽  
Drug Resistant ◽  
Temperature Dependent ◽  
Chinese Hamster Cells ◽  
Uptake And Release ◽  
Resistant Cells

A graded series of drug-resistant Chinese hamster sublines has been examined for biochemical changes accompanying resistance to actinomycin D. The most highly resistant subline, DC-3F/AD X, is maintained at 10 µg/ml of the antibiotic. It was shown that over 250 times more actinomycin D is required to inhibit RNA synthesis in this subline than in the parental DC-3F line. The DC-3F/AD X subline was also shown to have a somewhat reduced capacity to transport uridine as compared to parental cells. Sensitive cells took up over 50 times more tritiated antibiotic than the most resistant cells, as determined in a 1-h assay. Uptake of actinomycin D was shown to be temperature-dependent in both resistant and sensitive cells and was not influenced by various metabolic inhibitors. Resistance could not be explained by a rapid uptake and release of the antibiotic, as demonstrated in efflux experiments, or by its metabolism. In addition, highly resistant cells which are cross-resistant to puromycin were shown to have a reduced capacity to take up labeled puromycin. These studies provide further evidence indicating that the mechanism of resistance to actinomycin D is reduced permeability to drug and suggesting that cell membrane alteration accounts for resistance to both actinomycin D and puromycin.

Nature of the So-Called Fibrillar Component in the Segregated Nucleolus

1972 ◽  
Vol 30 ◽  
pp. 244-245
Author(s):  
T. Unuma ◽  
R. Senda ◽  
M. Muramatsu
Keyword(s):  
Rna Synthesis ◽  
Actinomycin D ◽  
Morphological Changes ◽  
Enzymatic Digestion ◽  
The Other ◽  
Granular Component ◽  
Fibrillar Component ◽  
Nucleolar Segregation ◽  
Nucleolar Components

Actinomycin D, selectively blocks DNA-dependent RNA synthesis, causes marked morphological changes of the nucleolus. One of the most prominent changes is the segregation of the nucleolar components. The granular component is redistributed and separated from the other components of the nucleolus. The term nucleolar segregation has been used to designate the separation of the granular and fibrillar components. In the present study, the nucleolus was segregated into two distinct zones, namely the granular and the so-called fibrillar component (Fig. 1). The purpose of this study is to present the evidences of protelnous nature of the so-called fibrillar component in the segregated nucleolus by morphological, biochemical and enzymatic digestion studies.

CHANGES IN LABELLING OF RIBONUCLEIC ACID SPECIES FRACTIONATED FROM THE RAT VENTRAL PROSTATE IN RESPONSE TO TESTOSTERONE

1975 ◽  
Vol 78 (2) ◽  
pp. 401-416 ◽  
Author(s):  
Anja Isotalo ◽  
R. S. Santti
Keyword(s):  
Body Weight ◽  
Glucose Metabolism ◽  
Ribosomal Rna ◽  
Orotic Acid ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Ventral Prostate ◽  
Total Rna ◽  
Testosterone Administration

ABSTRACT The total RNA isolated at various times up to 24 h after testosterone administration from the ventral prostate of castrated rats, was labelled either by injecting 3H-orotic acid directly into the ventral prostate 6 h before the animals were killed, or by incubating prostatic tissue in vitro with 3H-uridine for 20 to 60 min. The isolated RNA was separated into tRNA, ribosomal RNA (Q1 RNA) and two DNA-like RNA fractions (Q2) and TD RNA) by chromatography on methylated albumin kieselguhr (MAK) columns, and the fractions were further analysed by sucrose gradient centrifugation. Testosterone given into castrated animals for 12 h, stimulated the labelling of all main fractions. The radioactivity of TD RNA after a 60-min incubation period in vitro with 3H-uridine was approximately twice that seen in the castrated rat, while there was a 3.1- and 2.2-fold increase in the radioactivity of the Q1 and Q2 RNA fractions respectively. Kinetics of incorporation of 3H-uridine into different RNA fractions revealed that the hormone facilitated the labelling of the TD RNA fraction relatively more than that of the Q2 fraction. The injection of 3H-orotic acid into the ventral prostate labelled the Q1 RNA preferentially. More than 60 % of the recovered radioactivity was found in Q1 RNA (as 18 and 28 S). Testosterone increased markedly (9.4-fold) the labelling of this fraction. It was concluded that testosterone has an activatory effect on the production of ribosomal RNA, and the bulk of the testosterone effect on the total RNA labelling is to be found in this fraction. Furthermore, it seems likely that testosterone also stimulates both the synthesis and processing of DNA-like RNA. When actinomycin D was given 2 h before the hormone administration in a dose of 25 μg per 100 g of body weight, there was no noticeable increase in the labelling of any fraction above the level seen in the untreated castrated rat. There is evidence that testosterone exerts some effects on the labelling of proteins with radioactive amino acids and 14C-glucose metabolism in the absence of that fraction of the total RNA synthesis which is sensitive to a low dose (25 μg per 100 g of body weight) of actinomycin D (Isotalo & Santti 1972). In this way it may be concluded that the major changes of the RNA synthesis after testosterone administration are likely to be secondary to the protein synthesis and glucose metabolism, or the hormone exerts its anabolic effect on prostatic cells at different sites and by different modes of action, each of which can be operated independently.

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