Lac
 Operator Analogues: Bromodeoxyuridine Substitution in the
 lac
 Operator Affects the Rate of Dissociation of the
 lac
 Repressor

Lin, Syr-Yaung; Riggs, Arthur D.

doi:10.1073/pnas.69.9.2574

Cited by 141 publications

(56 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The recent report of Lin and Riggs provides a possible explanation for the (CH3)2SO-induction of differentiation as wvell as for the BrdU-inhibition of this process (23). These investigators, studying the lac operon, demonstrated that the rate of dissociation of repressor from BrdU-substituted operator DNA is 10-times slower than from normal operator.…”

Section: Resultsmentioning

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.

View full text Add to dashboard Cite

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:

show abstract

Section: Resultsmentioning

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.

View full text Add to dashboard Cite

show abstract

“…Another possibility is that they exist in many copies in the cell (7). Extending the former hypothesis LIN and RIGGS have proposed that cellular functions which are subject to regulation are selectively inhibited by BUdR because BUdR-substituted DNA binds regulatory proteins tighter than does normal DNA (35). This may explain not only the high sensitivity to BUdR substitution of differentiating systems, but also the appearance of virus-like particles after growth of guinea pig cells (11), polyoma transformed cells (20), spleen cells (44), rat embryo cells (48) and mouse melanoma cells (49) in BUdR containing media.…”

Section: Effects On Enzyme Activities Of Budr Substitution In Dnamentioning

confidence: 99%

“…This may explain not only the high sensitivity to BUdR substitution of differentiating systems, but also the appearance of virus-like particles after growth of guinea pig cells (11), polyoma transformed cells (20), spleen cells (44), rat embryo cells (48) and mouse melanoma cells (49) in BUdR containing media. In all cases altered binding to DNA of regulatory proteins may result in disruption of finely adjusted regulatory systems which normally check viral growth (35). However, induction of virus-like particles could also reflect increased probability of strand breaks by which virus particles are released.…”

Section: Effects On Enzyme Activities Of Budr Substitution In Dnamentioning

confidence: 99%

“…Change in the dissociation constant of chromatin containing BUdR-substituted DNA has been found (21). In E. coli the binding of lac-repressor to lac-operator is increased when the latter is substituted with BUdR (35). Since structural changes in the DNA may cause serious changes of normal cell functions precautions must be taken before BUdR is used as a tracer compound in studies of normal cellular processes.…”

Section: Effects Of Budr Incorporation Into the Dna Moleculementioning

confidence: 99%

See 1 more Smart Citation

Mechanisms of action of 5-bromodeoxyuridine based on studies with tetrahymena pyriformis

Lykkesfeldt

1977

Carlsberg Res. Commun.

View full text Add to dashboard Cite

The effect of BUdR on cell multiplication, DNA-, RNA-and protein synthesis in Tetrahymena pyriformis has been studied. Incorporation of BUdR up to 50% of the thymine sites in the DNA has no effect on the DNA synthetic rate at least for two generation times. RNA synthesis is inhibited shortly after addition of BUdR to exponentially growing populations, and experiments with synchronously growing populations have revealed that inhibition of RNA synthesis follows the incorporation of BUdR into the ribosomal RNA genes. A preferential inhibition of the synthesis of 25S and 17S ribosomal RNA has been found to take place at all degrees of BUdR substitution in DNA. Local changes in the structure of the DNA molecule leading to alterations in the pattern of transcription have been proposed as a model for the mode of action of BUdR. CONTENTS

show abstract

“…This phenomenon has been most extensively investigated by the filter binding technique with the lac repressor (5), CAP protein (6), and animal cell histones (7). In order to further investigate the possibility that BrdU acts on gewg expression through an altered binding of regulatory proteins to DNA it is important to establish whether or not this tighter binding phenomenon extends to other chromosomal proteins as well.…”

Section: Introductionmentioning

confidence: 99%

Interaction of chromosomal proteins with BrdU substituted DNA as determined by chromatin-DNA competition

Bick

Devine

1977

Nucl Acids Res

View full text Add to dashboard Cite

Chromatin-DNA competition has been utilized to examine the general nature of chromosomal proteins interacting more strongly with BrdU substituted DNA. When chromatin is incubated with an excess of purified DNA, a portion of the chromosomal proteins will exchange to the purified DNA. These two complexes can then be separated on Metrizamide gradients due to their differing protein/DNA ratios. Using this technique we observe that most nonhistone chromosomal proteins will exchange to a competitor DNA, the extent of exchange being directly dependent upon the competitor DNA being present in excess. While essentially the same proteins will migrate to either unsubstituted or BrdU substituted DNA, the substituted DNA is found to be a quantitatively better competitor and its effectiveness as a competitor is directly related to the level of BrdU substitution. INTRODUCTIONThe observation that the thymidine analog, 5-bromodeoxyuridine (BrdU), will alter the expression of specific genetic loci in animal cells (1-4) is frequently considered to be related to the demonstration of Lin and Riggs that both structural and regulatory proteins will bind more tightly to substituted DNA. This phenomenon has been most extensively investigated by the filter

show abstract

Lac Operator Analogues: Bromodeoxyuridine Substitution in the lac Operator Affects the Rate of Dissociation of the lac Repressor

Cited by 141 publications

References 25 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

Mechanisms of action of 5-bromodeoxyuridine based on studies with tetrahymena pyriformis

Interaction of chromosomal proteins with BrdU substituted DNA as determined by chromatin-DNA competition

Contact Info

Product

Resources

About