Cloned endogenous mouse mammary tumor virus DNA is biologically active in transfected mouse cells and its expression is stimulated by glucocorticoid hormones

Diggelmann, Heidi; Vessaz, Annelyse; Buetti, Elena

doi:10.1016/0042-6822(82)90233-1

Cited by 23 publications

(22 citation statements)

References 36 publications

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“…Deletions to -204 retained the ability to respond to the hormonal treatment ( Figure 4A, lanes 5 and 6; lanes 13-20; Figure 5A, lanes 5-8). The observed stimulation factor was 10to 20-fold, therefore still in the range of other experimental observations (Varmus et al, 1979;Buetti and Diggelmann, 1981;Hynes et al, 1981;Diggelmann et al, 1982). Although sequences between -204 and -600 may affect its amplitude, the glucocorticoid response itself was still present in all transfectants with deletions to position -204.…”

Section: Construction Of Deleted Plasmidssupporting

confidence: 77%

“…Since the stimulation is largely of the rate of transcription (Young et al, 1977;Ringold et al, 1977b;Ucker et al, 1981), MMTV has become a model system for studying steroid hormone action. Hormonal regulation was also observed in cells transfected with molecularly cloned forms of MMTV DNA, both of exogenous (Buetti and Diggelmann, 1981;Owen and Diggelmann, 1983) and endogenous origin Diggelmann et al, 1982). Transfections with subgenomic fragments (Fasel et al, 1982;Payvar et al, 1982) or with recombinant DNAs between MMTV fragments and a marker gene (Lee et al, 1981;Huang et al, 1981;Groner et al, 1981) indicated that hormone-responsive sequences are located within the viral long terminal repeat (LTR).…”

Section: Introductionmentioning

confidence: 99%

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Glucocorticoid regulation of mouse mammary tumor virus: identification of a short essential DNA region.

Buetti¹,

Diggelmann²

1983

The EMBO Journal

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119

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Transcription of mouse mammary tumor virus (MMTV) DNA is stimulated by steroid hormones. To determine the DNA sequences involved in this regulation, we constructed a plasmid containing the MMTV long terminal repeat (LTR) in front of the coding region of the herpes simplex thymidine kinase gene, from which the promoter had been removed. Portions of the LTR were removed by the nuclease Ba/31, and the deleted molecules were recloned and tested for transcriptional activity in transfections of Ltk‐aprt‐ cells. Stably transfected cell clones were selected and hormone‐dependent transcription from the MMTV promoter was studied by the S1 nuclease mapping method. The results show that DNA sequences between ‐105 and ‐204 base pairs upstream from the initiation site of viral transcription are required for glucocorticoid stimulation.

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Section: Construction Of Deleted Plasmidssupporting

confidence: 77%

Section: Introductionmentioning

confidence: 99%

Glucocorticoid regulation of mouse mammary tumor virus: identification of a short essential DNA region.

Buetti¹,

Diggelmann²

1983

The EMBO Journal

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119

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“…The newly acquired MMTV proviruses are transcriptionally active and regulated by glucocorticoid hormones (6). The nuclei of this cell line were purified as described above, and the preexisting RNA was removed by exposing the nuclei to RNase A. RNA fragments of 60 nucleotides in length are protected from the RNase A digestion by RNA polymerase II molecules bound to their DNA template (33).…”

Section: Resultsmentioning

confidence: 99%

“…By using this assay, three different cell lines (16-3, 15-1, L-cells) were investigated for expression of MMTV-specific sequences. Cell clone 16-3 is derived from Ltk-cells which had been stably transfected with cloned endogenous integrated proviral DNA together with plasmid containing the herpes tk gene (6). This clone contains approximately six copies of foreign MMTV DNA in addition to the L-cell endogenous MMTV proviruses.…”

Section: Resultsmentioning

confidence: 99%

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Dexamethasone increases the number of RNA polymerase II molecules transcribing integrated mouse mammary tumor virus DNA and flanking mouse sequences.

Firzlaff¹,

Diggelmann²

1984

Mol. Cell. Biol.

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In mouse Ltk-cells that were transfected with recombinant bacteriophage DNA containing a complete proviral copy of an integrated endogenous mouse mammary tumor virus (MMTV) with its flanking cellular sequences, the newly acquired MMTV proviruses were transcribed in a glucocorticoid-responsive fashion. After hormone treatment of selected cell clones in culture we isolated the nuclei, elongated the nascent RNA chains in vitro, and determined the number of RNA polymerase II molecules on the transcribed MMTV DNA as well as on the flanking mouse DNA sequences. We found that the specific increase in the polymerase loading after hormone treatment is proportional to the increase in the amount of stable MMTV mRNA. When the DNA sequences which are responsible for hormone-receptor binding and for the increased MMTV mRNA levels were deleted, no increase in RNA polymerase II loading on MMTV DNA was observed. Nuclear RNA chains which were transcribed in response to hormone treatment were detected not only from the transfected MMTV DNA but also from the mouse DNA sequences adjacent to the 3' end of the provirus.The expression of mouse mammary tumor virus (MMTV) in mice as well as in tissue culture cells is regulated by steroid hormones (18,23,25,28,38). Studies utilizing homologous and heterologous cell lines revealed that the strong stimulation of viral RNA synthesis could be attributed to the action of glucocorticoids alone. The regulation is mediated by a hormone-receptor complex; it is very rapid and independent of simultaneous protein synthesis (30, 34). The hormone-receptor complex binds specifically to sequences in the long terminal repeat (LTR) of MMTV DNA (9,11,26,27,32). In transfection experiments it was shown that the same DNA region is involved in glucocorticoidregulated transcription (7,13,16,17,19). Upon deletion of some of these sequences, the hormonal effect on MMTV transcription is lost (2,7,21). It was demonstrated that the sequences required for the hormone response reside between -100 and -200 nucleotides upstream from the initiation site for viral RNA synthesis (2, 21). Larger DNA fragments containing this sequence but lacking the MMTV promoter were shown to confer hormone inducibility to other, normally non-hormone-responsive promoters (3, 21). As a result of hormonal treatment, the rate of viral RNA synthesis increases, as measured by pulse labeling (12,29,36,37,40), and higher levels of stable mRNA accumulate. From these studies it was deduced that the rate of initiation of viral RNA synthesis is increased, but they could not exclude the possibility of rapid metabolic effects of glucocorticoids, which may alter for example the stability of the MMTV mRNA. To further study the molecular events which are caused by hormone-receptor binding to DNA, it was therefore necessary to apply an approach which looks more directly at initiation of transcription. In the experiments presented here we measured the number of RNA polymerase II molecules transcribing MMTV DNA with and without hormone treatment. With the sa...

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Hormonal Control of Mouse Mammary Tumor Virus Transcription

Hynes

Groner

Cato

et al. 1987

Cellular and Molecular Biology of Mammary Cancer

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Cloned endogenous mouse mammary tumor virus DNA is biologically active in transfected mouse cells and its expression is stimulated by glucocorticoid hormones

Cited by 23 publications

References 36 publications

Glucocorticoid regulation of mouse mammary tumor virus: identification of a short essential DNA region.

Glucocorticoid regulation of mouse mammary tumor virus: identification of a short essential DNA region.

Dexamethasone increases the number of RNA polymerase II molecules transcribing integrated mouse mammary tumor virus DNA and flanking mouse sequences.

Hormonal Control of Mouse Mammary Tumor Virus Transcription

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