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Promoter Methylation and Tissue-Specific Transcription of the α7 Nicotinic Receptor Gene, CHRNA7

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Abstract

The α7 nicotinic acetylcholine receptor is known to regulate a wide variety of developmental and secretory functions in neural and non-neural tissues. The mechanisms that regulate its transcription in these varied tissues are not well understood. Epigenetic processes may play a role in the tissue-specific regulation of mRNA expression from the α7 nicotinic receptor subunit gene, CHRNA7. Promoter methylation was correlated with CHRNA7 mRNA expression in various tissue types and the role of DNA methylation in regulating transcription from the gene was tested by using DNA methyltransferase (DNMT1) inhibitors and methyl donors. CHRNA7 mRNA expression was silenced in SH-EP1 cells and bisulfite sequencing PCR revealed the CHRNA7 proximal promoter was hypermethylated. The proximal promoter was hypomethylated in the cell lines HeLa, SH-SY5Y, and SK-N-BE which express varying levels of CHRNA7 mRNA. Expression of CHRNA7 mRNA was present in SH-EP1 cells after treatment with the methylation inhibitor, 5-aza-2-deoxycytidine (5-Aza-CdR), and increased in SH-EP1 and HeLa cells using another methylation inhibitor, zebularine (ZEB). Transcription from the CHRNA7 promoter in HeLa cells was increased when the methyl donor methionine (MET) was absent from the media. Using methylation-sensitive restriction enzyme analysis (MSRE), there was a strong inverse correlation between CHRNA7 mRNA levels and promoter DNA methylation across several human tissue types. The results support a role for DNA methylation of the proximal promoter in regulation of CHRNA7 transcription.

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Abbreviations

α7nACHR:

α7 neuronal nicotinic acetylcholine receptor

CHRNA7 :

Human α7 neuronal nicotinic acetylcholine receptor subunit gene

5-Aza-CdR:

5-Aza-2-deoxycytidine

ZEB:

Zebularine

BSP:

Sodium-bisulfite DNA modification coupled with PCR and sequencing

MSRE:

Methylation-sensitive restriction enzyme analysis

PNMT:

Phosphotidylethanolamine N-methyltransferase gene

RELN :

Human reelin gene

GAD1 :

Human glutamate decarboxylase 1 gene

DNMT1 :

DNA methyltransferase 1

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Acknowledgments

This study was supported by NIH grants DA09457, MH081177, the Veterans’ Affairs Medical Research Service to SL, and a T32-MH15442 fellowship and research funds from the Developmental Psychobiology Research Group at UCD to AC.

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Authors and Affiliations

  1. Department of Psychiatry, University of Colorado Denver, Mail Stop: 8344, P.O. Box 6511, Aurora, CO, 80045, USA

    Andrew Canastar, Judith Logel, Sharon Graw, Jessica Finlay-Schultz, Christina Osborne, Milda Palionyte, Carla Drebing, Molly Plehaty, Lisa Wilson, Rosemary Eyeson & Sherry Leonard

  2. Department of Biochemistry and Molecular Genetics, University of Colorado Denver, Aurora, CO, 80045, USA

    Jessica Finlay-Schultz

  3. The Veterans Affairs Medical Research Center, Denver, CO, 80220, USA

    Sherry Leonard

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  1. Andrew Canastar
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  2. Judith Logel
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Canastar, A., Logel, J., Graw, S. et al. Promoter Methylation and Tissue-Specific Transcription of the α7 Nicotinic Receptor Gene, CHRNA7 . J Mol Neurosci 47, 389–400 (2012). https://doi.org/10.1007/s12031-011-9663-7

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