Dual function of a nuclear factor I binding site in MMTV transcription regulation

Buetti, Elena; Kühnel, Blanka; Diggelmann, Heidi

doi:10.1093/nar/17.8.3065

Cited by 54 publications

(35 citation statements)

References 61 publications

(87 reference statements)

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“…3b). Cell-specific differences in the sensitivity to mutations in the proximal element have also been observed by others (9,11 (38) and does not bind NF-I in a DNase I in vitro footprinting analysis with nuclear extracts, whereas 90/78 does (6). T47D cells also contain an NF-I binding activity (Fig.…”

Section: Discussionsupporting

confidence: 79%

“…That physical interactions between receptors can occur was shown in an electron microscopy study (50) in which progesterone receptors appeared associated in pairs on the MMTV HRE. Using the same constructs, we observed a similar requirement for spacing of the elements in the glucocorticoid response in Ltk-cells (6).…”

Section: Discussionsupporting

confidence: 57%

“…4), corresponding to the NF-I binding site (38). No DNase I protection in this area was observed on DNA of the 90/70 mutant (6). In the region closer to the promoter, the mutant 37/11 (which removes the TATA homology) gave extremely low levels of correctly initiated transcripts, even in the presence of progestin.…”

Section: Methodsmentioning

confidence: 98%

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Mutations in the hormone regulatory element of mouse mammary tumor virus differentially affect the response to progestins, androgens, and glucocorticoids.

Gowland¹,

Buetti²

1989

Mol. Cell. Biol.

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Transcription of the mouse mammary tumor virus DNA is known to be induced by several steroid hormones. Using chimeric MMTV plasmids containing mutations within the hormone regulatory element, we have previously studied the regions required for the glucocorticoid response in mouse fibroblasts. Here we report the characterization of elements essential for the stimulation by progestins and androgens as compared with glucocorticoids. The same set of mutant plasmids was transfected into the human mammary tumor cell line T47D, and the specific transcripts were analyzed by an Si nuclease protection assay. Androgen-mediated stimulation, although weak, showed an extended sensitivity to mutations, with a slight preference for the proximal region. The results with progestin suggest that sequences within all the described sites protected by the receptor in vitro are required and that the promoter-proximal region (-128 to -78 from the RNA start site) is more important than the distal one (-190 to -160). Moreover, a binding site for nuclear factor I was not required for the progestin response, whereas it was required for glucocorticoids. Thus, the various steroid receptors play a role in the differential regulation of mouse mammary tumor virus transcription by recognizing distinct sequence differences in the hormone regulatory element and interacting with different factors bound to the promoter.The mouse mammary tumor virus (MMTV) has been successfully used as a model for studying the control of gene expression by steroid hormones (reviewed in references 41 and 54). In particular, glucocorticoids have been shown to stimulate MMTV transcription in infected cells (42,55) through the action of the glucocorticoid receptor. Mainly because of the widespread occurrence of this receptor in cultured cell lines, the DNA sequences required for glucocorticoid regulation have been investigated in more detail by using cells transfected with chimeric DNA molecules (25,28) and have subsequently been mapped to the 200 base pairs (bp) upstream of the MMTV RNA start site in the viral long terminal repeat (LTR) (4,26,31). Using a series of mutant plasmids containing clustered point mutations, small deletions, or additions, we demonstrated that maximal stimulation by glucocorticoids requires the cooperative action of multiple, distinct sequence elements (5, 27). The strongest, distal element maps at -181 to -172, inside an area that binds the purified glucocorticoid receptor (39, 44). Further elements map around -120, in one of the three proximal receptor-binding sites (43), and in the symmetric sequence around -70 recognized by nuclear factor I (NF-I [35,38] described previously (5, 27). The results provide evidence that in the natural context of the MMTV promoter, the proximal upstream region is more important than the distal one for the progestin and possibly the androgen responses and that the NF-I binding site is not required for the progestin response, in contrast to the situation observed with glucocorticoids. It is suggested that subtle ...

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Section: Discussionsupporting

confidence: 79%

Section: Discussionsupporting

confidence: 57%

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Mutations in the hormone regulatory element of mouse mammary tumor virus differentially affect the response to progestins, androgens, and glucocorticoids.

Gowland¹,

Buetti²

1989

Mol. Cell. Biol.

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“…In addition, basal activity can be detected both when MMTV is in a stably integrated state and transfected transiently ISmith et al 1993). One of the immediate proximal promoter factors, NF-I, has been implicated in the basal regulation of MMTV [Buetti et al 1989; Kalff et al 1990; Toohey et al 19901, but NF-I does not bind its cognate site in nucleosomal Nuc-B reconstitutes [Pina et al 1990b;Archer et al 1991). It is conceivable that NF-I binds a specific configuration of Nuc-B nucleosomes that was not reproduced previously in vitro.…”

Section: Discussionmentioning

confidence: 99%

Nucleosome positioning on the MMTV LTR results from the frequency-biased occupancy of multiple frames.

Fragoso

John²,

Roberts³

et al. 1995

Genes Dev.

166

124

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The translational positions of nucleosomes in the promoter region of the mouse mammary tumor virus (MMTV) were defined at high resolution. Nucleosome boundaries were determined in primer extension assays using full-length single-stranded mononucleosomal DNA prepared from cells treated with formaldehyde, a reversible protein-DNA cross-linking agent. Multiple boundaries were observed in both the nucleosome A (Nuc-A) and Nuc-B region of the promoter, indicating multiple nucleosome translational frames. The different nucleosome frames in both the Nuc-A and Nuc-B regions were occupied unequally. The most frequently occupied frames were found clustered within 50-60 bases of each other, resulting in a distribution centered in the positions defined previously at low resolution for Nuc-A and Nuc-B. The most abundant 5' ends of the frames in the B region were found between -235 and -187, and the 3' ends between -86 and -36, whereas in the A region the most abundant 5' ends were between -22 and +42, and the 3' ends between + 121 and +186. Although frames in the Nuc-B region of the LTR extend at a low frequency in the 5' direction toward the Nuc-C region, there is a sharp discontinuity in the 3' direction toward Nuc-A, suggesting the presence of a boundary constraint in the A-B linker. The positions and relative occupancies of nucleosome frames, in either the B or the A region, did not change when the promoter was activated with dexamethasone.

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“…A direct physical interaction of glucocorticoid receptor (GR) with transcription factors such as the p65 subunit of NF-B (8), Ets related factor (9), octamer transcription factor 1 (10), Spi-1 (11), and SWI1 proteins (12) have been shown to modulate glucocorticoid receptor function. Transcription factors such as NF-1 (13) and Oct-1 (14,15) which bind close to proximal GRE sites seem to modulate the MMTV promoter transcription. The glucocorticoid receptor bound to the distal GRE site (Ϫ186 to Ϫ170) of the MMTV LTR was found to cooperate with other binding sites to which transcription factors such as NF-1, octamer transcription factor (Oct-1), SP1, and CACCC box binding factor (15)(16)(17) were shown to bind.…”

mentioning

confidence: 99%

Cellular Factors Binding to a Novel cis-Acting Element Mediate Steroid Hormone Responsiveness of Mouse Mammary Tumor Virus Promoter

Lee

Reddy

1995

Journal of Biological Chemistry

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Steroid hormone receptors regulate mouse mammary tumor virus (MMTV) gene expression by binding to hormone response DNA elements present in the long terminal repeat. Tissue-specific expression of MMTV is unlikely to be regulated by steroid hormone-receptor complex alone, and mammary cell-specific factors might play a role in the hormone-induced transcriptional activation. In this report we have investigated the function of a novel cis-acting element designated Kil (؊204 to ؊188) which is located adjacent to the distal glucocorticoid response element, in steroid hormone-induced transcription of MMTV. Electrophoretic mobility shift assays indicate that cellular factors bind to the Kil element, and dexamethasone stimulation results in alterations in the binding pattern of proteins in this region. By transient transfection assays using wild type and deletion mutants of the Kil element, we show that this novel cis-acting element is necessary for hormone-induced transcription of MMTV and functions in mammary tumor cells but not in NIH/3T3 cells. Mutagenesis of the Kil sequence suggests that the entire Kil element functioning as one unit is necessary for hormone-induced transcription of MMTV. When placed in the context of heterologous promoters, neither Kil element nor glucocorticoid response element is able to induce significant hormone-induced transcription of MMTV. The presence of both the DNA elements in tandem results in optimal induction of transcription in the presence of steroid hormones. Our results also indicate that the Kil element functions in human breast carcinoma cell lines such as T47D and MCF-7. These results suggest that Kil element in combination with distal glucocorticoid response element functions as a mammary cell-specific enhancer to regulate MMTV transcription.Steroid hormone receptors are cytoplasmic proteins, which, upon binding to their ligands, translocate into the nucleus and bind to DNA elements in a sequence-specific manner and modulate the transcription of cellular genes (1). Transcription from the long terminal repeat (LTR) 1 of mouse mammary tumor virus (MMTV) has been used extensively to understand the regulation of gene expression by steroid hormones. Glucocorticoids, progesterones, and androgens induce the transcription of MMTV (2-4) upon binding of their respective receptors to the hormone response elements. These elements are located between Ϫ202 and Ϫ59 bases upstream of the transcription initiation site in the MMTV promoter, which can be divided into the proximal and distal glucocorticoid response elements (GREs). The GRE consists of four repeats of the hexanucleotide 5Ј-TGTTCT-3Ј to which the steroid hormone receptors were shown to bind and exert their effects on transcription. The proximal glucocorticoid responsive element (GRE) contains three repeats of the hexanucleotide, while the distal GRE (Ϫ188 to Ϫ170) contains only one of the hexanucleotide. Apart from steroid hormone-receptor complex, which is a major determinant of MMTV transcription, cellular factors have been implic...

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Dual function of a nuclear factor I binding site in MMTV transcription regulation

Cited by 54 publications

References 61 publications

Mutations in the hormone regulatory element of mouse mammary tumor virus differentially affect the response to progestins, androgens, and glucocorticoids.

Mutations in the hormone regulatory element of mouse mammary tumor virus differentially affect the response to progestins, androgens, and glucocorticoids.

Nucleosome positioning on the MMTV LTR results from the frequency-biased occupancy of multiple frames.

Cellular Factors Binding to a Novel cis-Acting Element Mediate Steroid Hormone Responsiveness of Mouse Mammary Tumor Virus Promoter

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