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Mxi1 inhibits the proliferation of U87 glioma cells through down-regulation of cyclin B1 gene expression.

Manni I, Tunici P, Cirenei N, Albarosa R, Colombo BM, Roz L, Sacchi A, Piaggio G, Finocchiaro G - Br. J. Cancer (2002)

Bottom Line: Interestingly, the expression of cyclin B1 was inhibited to about 60% in U87.Mxi1.14 cells.This inhibition occurs at the transcriptional level and depends, at least in part, on the E-box present on the cyclin B1 promoter.Consistent with this, the endogenous Mxi1 binds this E-box in vitro.

View Article: PubMed Central - PubMed

Affiliation: Istituto Regina Elena, Centro Ricerca Sperimentale, Laboratorio di Oncogenesi Molecolare, Via delle messi D'Oro 156, 00158 Rome, Italy.

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Max/Mxi1 heterodimers bind the E-box present on the cyclin B1 promoter. Gel mobility retardation assays were performed with a double-stranded, 32P-labelled oligonucleotide, spanning position −133/−110, containing the E-box of the cyclin B1 promoter (B1 E-box). Addition of nuclear extract from U87 cells resulted in a retarded band. A 200-fold molar excess of unlabelled probe (lane 6) but not an oligonucleotide containing a non-specific binding site (lane 7) specifically inhibits the formation of the complex. In lanes 3, 4, nuclear extracts were pre-incubated with anti-Mxi1 antibodies and, in lane 5, with anti-Max antibody.
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fig8: Max/Mxi1 heterodimers bind the E-box present on the cyclin B1 promoter. Gel mobility retardation assays were performed with a double-stranded, 32P-labelled oligonucleotide, spanning position −133/−110, containing the E-box of the cyclin B1 promoter (B1 E-box). Addition of nuclear extract from U87 cells resulted in a retarded band. A 200-fold molar excess of unlabelled probe (lane 6) but not an oligonucleotide containing a non-specific binding site (lane 7) specifically inhibits the formation of the complex. In lanes 3, 4, nuclear extracts were pre-incubated with anti-Mxi1 antibodies and, in lane 5, with anti-Max antibody.

Mentions: We previously demonstrated that the Max protein recognizes the E-box present on the cyclin B1 promoter (Farina et al, 1996). By EMSAs we verified the ability of Mxi1 to bind the cyclin B1 E-box. Electromobility shift assays, performed with U87 nuclear extracts, reveal one protein complex binding to the radiolabelled probe, spanning nt −133 to −110 in the cyclin B1 promoter (B1 E-box) (Figure 8Figure 8


Mxi1 inhibits the proliferation of U87 glioma cells through down-regulation of cyclin B1 gene expression.

Manni I, Tunici P, Cirenei N, Albarosa R, Colombo BM, Roz L, Sacchi A, Piaggio G, Finocchiaro G - Br. J. Cancer (2002)

Max/Mxi1 heterodimers bind the E-box present on the cyclin B1 promoter. Gel mobility retardation assays were performed with a double-stranded, 32P-labelled oligonucleotide, spanning position −133/−110, containing the E-box of the cyclin B1 promoter (B1 E-box). Addition of nuclear extract from U87 cells resulted in a retarded band. A 200-fold molar excess of unlabelled probe (lane 6) but not an oligonucleotide containing a non-specific binding site (lane 7) specifically inhibits the formation of the complex. In lanes 3, 4, nuclear extracts were pre-incubated with anti-Mxi1 antibodies and, in lane 5, with anti-Max antibody.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2375210&req=5

fig8: Max/Mxi1 heterodimers bind the E-box present on the cyclin B1 promoter. Gel mobility retardation assays were performed with a double-stranded, 32P-labelled oligonucleotide, spanning position −133/−110, containing the E-box of the cyclin B1 promoter (B1 E-box). Addition of nuclear extract from U87 cells resulted in a retarded band. A 200-fold molar excess of unlabelled probe (lane 6) but not an oligonucleotide containing a non-specific binding site (lane 7) specifically inhibits the formation of the complex. In lanes 3, 4, nuclear extracts were pre-incubated with anti-Mxi1 antibodies and, in lane 5, with anti-Max antibody.
Mentions: We previously demonstrated that the Max protein recognizes the E-box present on the cyclin B1 promoter (Farina et al, 1996). By EMSAs we verified the ability of Mxi1 to bind the cyclin B1 E-box. Electromobility shift assays, performed with U87 nuclear extracts, reveal one protein complex binding to the radiolabelled probe, spanning nt −133 to −110 in the cyclin B1 promoter (B1 E-box) (Figure 8Figure 8

Bottom Line: Interestingly, the expression of cyclin B1 was inhibited to about 60% in U87.Mxi1.14 cells.This inhibition occurs at the transcriptional level and depends, at least in part, on the E-box present on the cyclin B1 promoter.Consistent with this, the endogenous Mxi1 binds this E-box in vitro.

View Article: PubMed Central - PubMed

Affiliation: Istituto Regina Elena, Centro Ricerca Sperimentale, Laboratorio di Oncogenesi Molecolare, Via delle messi D'Oro 156, 00158 Rome, Italy.

Show MeSH
Related in: MedlinePlus