5-Bromodeoxyuridine-Induced Differentiation of a Neuroblastoma

Schubert, David; Jacob, François

doi:10.1073/pnas.67.1.247

Cited by 202 publications

(38 citation statements)

References 8 publications

(1 reference statement)

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“…If globin mRNA synthesized by BrU-treatect cells contained frequent errors in transcription so that one base in 50 was incorrectly transcribed, a difference in the stability of duplexes formed between cDNA and globin mRNA from cells treated with (CH3)2S0 and with BrdU and (CH3)2S0 would have been detected (22). No An unusual observation has been the report that BrdU induces the differentiation of neuroblastoma cells in vitro, an effect independent of the incorporation of DNA (5). This latter observation is contrary to other studies which indicated that BrdU incorporation into DNA was essential for its biological effects (1,2,6).…”

Section: Resultsmentioning

confidence: 99%

“…5-Bromo-2 '-deoxyuridine (BrdU) provides a tool for investigating some of the molecular events involved in ontogenesis as well as oncogenesis, for it affects tissues as dissimilar as muscle cells of chick embryo (1), mammary tissue of pregnant mice (2), and malignant cells (3)(4)(5)(6)(7). The effects on each of these systems are considered to be specific and nonrandom.…”

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confidence: 99%

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Effects of 5-Bromo-2′-Deoxyuridine on Production of Globin Messenger RNA in Dimethyl Sulfoxide-Stimulated Friend Leukemia Cells

Preisler¹,

Housman

Scher³

et al. 1973

Proc. Natl. Acad. Sci. U.S.A.

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From these studies, where low doses which did not significantly interfere with cell replication had been used, synthesis of proteins characteristic of the final differentiated state was inhibited. For example, hemoglobin synthesis by chick embryo was inhibited during culture in the presence of BrdU (8, 9).We previously reported that dimethyl sulfoxide [(CH3)2S0 I enhances heme synthesis and erythroid differentiation of Friend leukemia cells in vitro (10) and that inhibition of these effects by BrdU was dependent on its incorporation into DNA (7). Similarly, other investigators have observed inhibition by BrdU of (CH3)2S0-induced stimulation of globin synthesis (11). Since appreciable amounts of globin mRNA were detected in (CH3)2S0-stimulated, but not in control, Friend leukemia cells cultures, it appeared that (CH3)2S0 might act at a transcriptional level (12, 21.)The present studies were designed to determine whether BrdU might also affect transcription of globin mRNA by Friend leukemia cells. We found that a decrease in the amount of globin mRNA accompanied inhibition of (CH3) BrdU-and (CH3)2S0-treated-cells cultured in the presence of both 2% (CH3)2S0 and 3 ,g/ml of BrdU. The cultures in each experiment were seeded simultaneously and maintained in the dark.Preparation of RNA RNA was prepared by a modification of the method of Natta (29). All glassware was treated with 0.1% ethyl oxidiformate (Eastman Kodak Co.) and then autoclaved; all plastic was treated with 2.0% Na dodecyl sulfate. 1 to 2 X 109 cells were suspended in 10 ml of isotonic saline and added by drops to an extraction mixture consisting of 40 ml of water-saturated distilled phenol, 20 ml of buffer R [46 mM NaCl-20 mM sodium acetate-2 mM NaEDTA (pH 5.2) made 2% with Na dodecyl sulfate], and 50 mg of bentonite dry weight, prepared by the method of Fraenkel-Conrat et al. (13). After the mixture was shaken for 30 min at 40, the aqueous and phenol phases were separated by centrifugation at 17,300 X g. 10 ml of phenol was added to the aqueous phase, which was kept on ice while the phenol phase and the interphase were again extracted with 20 ml of buffer R by shaking for 15 min at 4°. The aqueous and phenol phases were again separated by centrifugation. The aqueous phases were combined and extracted with 30 ml of additional phenol by shaking for 20 min at 4°. After centrifugation for 5 min, the phenol phase was discarded. The aqueous phase was centrifuged for 20 min at 30,900 X g and the pellet was discarded. 0.1 Volume of 3 M NaCl and 2 volumes of absolute ethanol were added to the aqueous phase, and the mixture was allowed to stand over-2956 Abbreviations: BrdU, 5-bromo-2'-deoxyuridine; (CH3)2SO, dimethyl sulfoxide; cDNA, complementary DNA. t Present address:

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Effects of 5-Bromo-2′-Deoxyuridine on Production of Globin Messenger RNA in Dimethyl Sulfoxide-Stimulated Friend Leukemia Cells

Preisler¹,

Housman

Scher³

et al. 1973

Proc. Natl. Acad. Sci. U.S.A.

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“…This analogue can promote neurite extension in the absence of DNA synthesis, however (Schubert and Jacob, 1970).…”

Section: B Neuroblasto Ma Cellsmentioning

confidence: 99%

Culture of Neural Tissue and Cells in Research Methods in Neurochemistry. Vol. Ii.

Herschman¹

1972

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“…In contrast, exposure of BrdU-resistant, thymidine kinase-NOTES negative (TK-) Friend cells to BrdU results in induction or potentiation of DMSO-induced differentiation (6). BrdU has been shown to have a selective effect upon a number of differentiating systems (1, 4, 13, 17, 22-24, 29, 30, 33) under conditions in which mutagenicity can be excluded as a mechanism (for review, see reference 25), and occasionally in which these effects are not related to the incorporation of the analog into DNA (5,6,24,26). Similarly, purine analogs have been shown to induce differentiation in purine analog-resistant Friend cells, whereas hypoxanthine is an effective inducer of wild-type and both purine and pyrimidine analog-resistant Friend erythroleukemia cells (12; unpublished data).…”

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confidence: 99%

Induction and inhibition of Friend erythroleukemia cell differentiation by pyrimidine analogs: analysis of the requirement for intracellular accumulation and incorporation into DNA.

Bilello¹,

Gauri²,

Kühne³

et al. 1982

Mol. Cell. Biol.

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Alkyldeoxyuridines which differ from thymidine by a C5 substitution of straight or branched alkyl chains of two to six carbon atoms have been tested for their ability to be taken up, phosphorylated, and incorporated into DNA. Analysis of the uptake of 5-ethyl-2'-deoxyuridine and 5-propyl-2'-deoxyuridine (n-PrdU)-similar to both thymidine and 5-bromo-2'-deoxyuridine-indicates that transport is dependent upon a functional cellular thymidine kinase. All of the aforementioned pyrimidines with the exception of n-PrdU are phosphorylated to the triphosphate and incorporated into DNA. The homologs 5-iso-propyl-2'-deoxyuridine (iso-PrdU) and 5-hexyl-2'-deoxyuridine are neither transported into the cell, phosphorylated, nor incorporated into DNA. These analogs were tested (i) for their ability to induce in the absence of dimethyl sulfoxide and (ii) to determine whether they enhance or inhibit dimethyl sulfoxide-induced differentiation of Friend erythroleukemia cells. Inhibition of erythroid differentiation appears to require the incorporation of thymidine analogs into DNA, and thus only 5-ethyl-2'-deoxyuridine and 5-bromo-2'-deoxyuridine were effective in inhibiting dimethyl sulfoxide-induced differentiation. The observation that iso-PrdU, and to a lesser extent n-PrdU and 5-propyldeoxyuridine monophosphate, induce differentiation under conditions in which they are not detectable intracellularly is strong evidence that this class of inducer acts at the cell membrane.Friend erythroleukemia cells are a suitable model system to study control mechanisms involved in alterations in cellular gene expression which lead to phenotypical and biochemical changes characteristic of erythroid differentiation in vivo. This system is particularly advantageous because induced erythroid differentiation in Friend cells is a synchronous process which gives rise to a homogeneous, differentiated cell population (for review, see reference 21). Erythroid differentiation, which is induced in this system by a number of compounds of diverse chemical structure, is readily assessed by measurement of hemoglobin, which comprises 20 to 50% of the total cytoplasmic protein in induced cells. Changes in cell architecture and the synthesis of globin mRNA and other differentiationspecific proteins are observed after exposure to inducers. Our laboratory has been concerned with changes in gene expression in response to alterations in the cellular environment and with the characterization of early events in the induction of Friend cells (3, 15). We have been particularly interested in whether inducers must enter the cell to affect differentiation and whether induction is a membrane-mediated process. Studies using labeled dimethyl sulfoxide (DMSO) (18), butyric acid (1), or actinomycin D (31) indicated that inducers accumulated intracellularly after a certain lag period. However, studies on membrane transport indicated that significant alterations in membrane function occur before the internalization of significant levels of inducer (3, 11, 15, 20; unpublished d...

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5-Bromodeoxyuridine-Induced Differentiation of a Neuroblastoma

Cited by 202 publications

References 8 publications

Effects of 5-Bromo-2′-Deoxyuridine on Production of Globin Messenger RNA in Dimethyl Sulfoxide-Stimulated Friend Leukemia Cells

Effects of 5-Bromo-2′-Deoxyuridine on Production of Globin Messenger RNA in Dimethyl Sulfoxide-Stimulated Friend Leukemia Cells

Culture of Neural Tissue and Cells in Research Methods in Neurochemistry. Vol. Ii.

Induction and inhibition of Friend erythroleukemia cell differentiation by pyrimidine analogs: analysis of the requirement for intracellular accumulation and incorporation into DNA.

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