A Minimal Promoter for the GABAA Receptor α6-Subunit Gene Controls Tissue Specificity

Authors

  • Pamela J. McLean,

    1. Laboratory of Molecular Neurobiology, Department of Pharmacology, Boston University School of Medicine, Boston, Massachusetts, U.S.A.
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  • Diana Shpektor,

    1. Laboratory of Molecular Neurobiology, Department of Pharmacology, Boston University School of Medicine, Boston, Massachusetts, U.S.A.
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  • Sabita Bandyopadhyay,

    1. Laboratory of Molecular Neurobiology, Department of Pharmacology, Boston University School of Medicine, Boston, Massachusetts, U.S.A.
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  • Shelley J. Russek,

    1. Laboratory of Molecular Neurobiology, Department of Pharmacology, Boston University School of Medicine, Boston, Massachusetts, U.S.A.
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  • David H. Farb

    1. Laboratory of Molecular Neurobiology, Department of Pharmacology, Boston University School of Medicine, Boston, Massachusetts, U.S.A.
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  • Lippincott Williams & Wilkins, Inc., Philadelphia

  • The present address of Dr. P. J. McLean is Alzheimer's Disease Research Unit, Department of Neurology, Massachusetts General Hospital—East, 149 13th Street, Charlestown, MA 02129, U.S.A.

  • Abbreviations used: GABA, γ-aminobutyric acid; GABAA, type A γ-aminobutyric acid; Inr element, initiator element.

Address correspondence and reprint requests to Dr. D. H. Farb at Laboratory of Molecular Neurobiology, Department of Pharmacology, Boston University School of Medicine, 80 East Concord Street, Boston, MA 02118, U.S.A. E-mail: dfarb@bu.edu

Abstract

Abstract: The ability of nerve cells to regulate the expression of specific neurotransmitter receptors is of central importance to nervous system function, but little is known about the DNA elements that mediate neuron specific gene expression. The type A γ-aminobutyric acid (GABAA) receptor α6-subunit gene, which is expressed exclusively in cerebellar granule cells, presents a unique opportunity to study the cis elements involved in restricting gene expression to a distinct neuronal population. In an effort to identify the regulatory elements that govern cerebellar granule cell-specific gene expression, the proximal 5′ flanking regions for the human, rat, and mouse α6 genes were cloned and sequenced, and a major transcriptional initiation site was identified in the rodent genes. Functional analysis of rat α6 gene-reporter constructs in primary neuronal cultures reveals that a 155-bp TATA-less promoter region (-130 to +25 bp) constitutes a minimal promoter that can drive cerebellar granule cell-specific expression. Internal deletion and decoy competition studies demonstrate that the minimal promoter contains a 60-bp region (-130 to -70 bp) that is critical for enhanced promoter activity in cerebellar granule cells. Activity of the compromised promoter containing the deletion cannot be rescued by placing the 60-bp region downstream of the reporter gene, demonstrating that it is not a classical enhancer but rather a positionally dependent regulator. An additional cerebellar-specific activating sequence is located between -324 and -130 bp, and a downstream negative regulatory region (+158 to +294) has been shown to be active in fibroblasts but inactive in cerebellar granule cells. Taken together, the results suggest a possible mechanism for the control of cerebellar granule cell-specific expression of the GABAA receptor α6 subunit gene.

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