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The von Hippel-Lindau (VHL) Tumor-suppressor Gene is Down-regulated by Selenium Deficiency in Caco-2 Cells and Rat Colon Mucosa

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Abstract

To test the hypothesis that selenium affects DNA methylation and hence gene regulation, we employed a methylation array (Panomics) in the human colonic epithelial Caco-2 cell model. The array profiles DNA methylation from promoter regions of 82 human genes. After conditioning cells to repeatedly reduced concentrations of fetal bovine serum, a serum-free culture was established. Se-methylselenocysteine (SeMSC) was added at 0 (deficient Se) or 250 (control Se) nM to cells maintained in DMEM. After 7 days, cells were collected and stored at −80°C until analysis; experiments were replicated three times. Glutathione peroxidase activity was significantly decreased in cells grown in low SeMSC. Cells grown in 250 nM SeMSC had maximal GPx activity. Genomic DNA from cells grown in the low-SeMSC media and media containing 250 nM SeMSC was incubated with methylation-binding protein followed by isolation of methylated DNA. The methylated DNA was labeled with biotin and hybridized to the methylation array. Thus, genes with promoter methylation will produce a higher chemiluminescence signal than those genes with no promoter methylation. Of the genes profiled, the von Hippel-Lindau (VHL) gene was most different as indicated by quantification following chemiluminescence detection demonstrating that the promoter region of VHL was hypermethylated in cells from the low-SeMSC media. To determine whether promoter methylation affected transcription, we isolated RNA from replicate samples and performed real-time RT PCR. VHL (mRNA) was down-regulated (fold change significantly <1) in cells grown in low SeMSC compared to cells grown in 250 nM SeMSC (control; fold change = 1). We also show that (mRNA) Vhl expression is significantly reduced in mucosa from rats fed a diet deficient in Se. Our results suggest that low Se status affects DNA promoter region methylation and that this can result in down-regulation of the tumor suppressor gene VHL.

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Abbreviations

VHL:

von Hippel-Lindau gene (human)

Vhl:

von Hippel-Lindau gene (rat)

SeMSC:

Se-methylselenocysteine

GPx:

glutathione preoxidase

DNMT:

DNA methyltransferase

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Disclosure

E. O. Uthus, A. Begaye, S. A. Ross, and H. Zeng have no conflicts of interest.

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Authors and Affiliations

  1. USDA ARS Human Nutrition Res Center, PO Box 9034, Grand Forks, ND, 58202-9034, USA

    Eric Uthus & Huawei Zeng

  2. University of Arizona, Tucson, USA

    Adrienne Begaye

  3. NIH/NCI, Nutritional Sciences Research Group, 6130 Executive Blvd, MSC 7328, Rockville, MD, 20892-7328, USA

    Sharon Ross

Authors
  1. Eric Uthus
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  2. Adrienne Begaye
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  3. Sharon Ross
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  4. Huawei Zeng
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Correspondence to Eric Uthus.

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The US Department of Agriculture, Agricultural Research Service, Northern Plains Area, is an equal opportunity/affirmative action employer and all agency services are available without discrimination. Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the US Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable. This work was supported by the US Department of Agriculture and National Cancer Institute.

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Uthus, E., Begaye, A., Ross, S. et al. The von Hippel-Lindau (VHL) Tumor-suppressor Gene is Down-regulated by Selenium Deficiency in Caco-2 Cells and Rat Colon Mucosa. Biol Trace Elem Res 142, 223–231 (2011). https://doi.org/10.1007/s12011-010-8764-4

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