Mithramycin inhibits SP1 binding and selectively inhibits transcriptional activity of the dihydrofolate reductase gene in vitro and in vivo.

Blume, Scott W.; Snyder, Richard C.; Ray, Ratna B.; Thomas, Shelia D.; Koller, Charles; Miller, Donald M.

doi:10.1172/jci115474

Cited by 306 publications

(253 citation statements)

References 48 publications

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“…5B, lanes 3 and 4, and 7 and 8). These observations corroborate with the fact that mithramycin A modifies specific G/C-rich DNA target sequences, which interferes with Sp1 protein binding [25][26][27]. …”

Section: Mithramycin a Inhibits While Monokines Increase Sp1 Binding supporting

confidence: 81%

Transcriptional control of human T‐BET expression: The role of Sp1

Min

Boyd

et al. 2007

Eur J Immunol

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Murine T-bet (T-box expressed in T cells) is a master regulator of IFN-c gene expressionin NK and T cells. T-bet also plays a critical role in autoimmunity, asthma and other diseases. However, cis elements or trans factors responsible for regulating T-bet expression remain largely unknown. Here, we report on our discovery of six Sp1-binding sites within the proximal human T-BET promoter that are highly conserved among mammalian species. Electrophoretic mobility shift assays demonstrate a physical association between Sp1 and the proximal T-BET promoter with a direct dose response between Sp1 expression and T-BET promoter activity. Ectopic overexpression of Sp1 also enhanced T-BET expression and cytokine-induced IFN-c secretion in NK cells and T cells. Mithramycin A, which blocks the binding of Sp1 to the T-BET promoter, diminished both T-BET expression and IFN-c protein production in monokine-stimulated primary human NK cells. Collectively, our results suggest that Sp1 is a positive transcriptional regulator of T-BET. As T-BET and IFN-c are critically important in inflammation, infection, and cancer, targeting Sp1, possibly with mithramycin A, may be useful for preventing and/ or treating diseases associated with aberrant T-BET or IFN-c expression.

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Section: Mithramycin a Inhibits While Monokines Increase Sp1 Binding supporting

confidence: 81%

Transcriptional control of human T‐BET expression: The role of Sp1

Min

Boyd

et al. 2007

Eur J Immunol

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“…Pretreatment of WT BMDM with mithramycin A, a specific Sp1 inhibitor, 40,41 prevented BCM-induced expression of caveolin-1 mRNA and protein (Figures 5a and b), as well as BCM reduced Mϕ surface expression of TLR4, a reflection of TLR4 internalization ( Figure 5c). …”

Section: Figure 1 Continuedmentioning

confidence: 95%

Tissue damage negatively regulates LPS-induced macrophage necroptosis

Scott

Fan

et al. 2016

Cell Death Differ

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Infection is a common clinical complication following tissue damage resulting from surgery and severe trauma. Studies have suggested that cell pre-activation by antecedent trauma/tissue damage profoundly impacts the response of innate immune cells to a secondary infectious stimulus. Cell necroptosis, a form of regulated inflammatory cell death, is one of the mechanisms that control cell release of inflammatory mediators from important innate immune executive cells such as macrophages (Mϕ), which critically regulate the progress of inflammation. In this study, we investigated the mechanism and role of trauma/tissue damage in the regulation of LPS-induced Mϕ necroptosis using a mouse model simulating long-bone fracture. We demonstrate that LPS acting through Toll-like receptor (TLR) 4 promotes Mϕ necroptosis. However, necroptosis is ameliorated by high-mobility group box 1 (HMGB1) release from damaged tissue. We show that HMGB1 acting through cell surface receptor for advanced glycation end products (RAGE) upregulates caveolin-1 expression, which in turn induces caveolae-mediated TLR4 internalization and desensitization to decrease Mϕ necroptosis. We further show that RAGE-MyD88 activation of Cdc42 and subsequent activation of transcription factor Sp1 serves as a mechanism underlying caveolin-1 transcriptional upregulation. These results reveal a previous unidentified protective role of damage-associated molecular pattern (DAMP) molecules in restricting inflammation in response to exogenous pathogen-associated molecular pattern molecules.

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“…RNA isolation, Northern blots and ELISA 3T3-L1 adipocytes differentiated for 10 days were maintained with low glucose (5.5 mmol/l) or high glucose (25 mmol/l) for 48 h. Mithramycin A (100 nmol/l) (Sigma), an inhibitor of Sp1 binding to DNAs [29], was added 24 h before harvesting cells. Total RNAs were extracted using Trizol (Invitrogen) according to the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

Regulation of human resistin gene expression in cell systems: an important role of stimulatory protein 1 interaction with a common promoter polymorphic site

et al. 2005

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Aims/hypothesis: Resistin is an adipokine that might link obesity and insulin resistance. A common polymorphism of the human resistin gene, −420C>G, is a major determinant of plasma resistin concentrations as well as resistin mRNA expression in human adipose tissue. In this study, we investigated the regulatory mechanism by which this polymorphism affects resistin expression. Methods: Electrophoretic mobility shift assay was performed to identify the transcription factors binding to the −420G region. Transient transfection and reporter assay were used to measure promoter activities of the resistin gene. The binding ability of stimulatory protein 1 (Sp1) in response to adipocyte differentiation or high glucose concentrations was also measured. Results: Sp1 and stimulatory protein 3 (Sp3) specifically bound to the region around −420G of the human resistin gene. Overexpression of Sp1 increased the promoter activity regardless of −420 genotypes, while the promoter activity of the −420G construct was two-fold higher than that of the −420C construct. In contrast, overexpression of Sp3 scarcely increased the promoter activity. The binding ability of Sp1 to the −420G region was increased in response to adipocyte differentiation. Mithramycin A, an inhibitor of DNA binding of Sp1, reduced the effect of high glucose on transcription induction of the resistin gene in adipocytes. Conclusions/interpretation: These results suggest that Sp1 is an important factor regulating transcription of human resistin gene. A common polymorphism of the human resistin promoter, −420C>G, is critical for the binding of Sp1 and modulates the transcriptional activity of the resistin gene by changing the binding ability of Sp1. In addition, Sp1 may be involved in the increase of resistin expression by hyperglycaemia.

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Mithramycin inhibits SP1 binding and selectively inhibits transcriptional activity of the dihydrofolate reductase gene in vitro and in vivo.

Abstract: The promoter of the human dihydrofolate reductase (DHFR)

Cited by 306 publications

References 48 publications

Transcriptional control of human T‐BET expression: The role of Sp1

Transcriptional control of human T‐BET expression: The role of Sp1

Tissue damage negatively regulates LPS-induced macrophage necroptosis

Regulation of human resistin gene expression in cell systems: an important role of stimulatory protein 1 interaction with a common promoter polymorphic site

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