Transcriptional Repression by a Conserved Intronic Sequence in the Nicotinic Receptor α3 Subunit Gene

Yuly F Fuentes Medel,Paul D Gardner

doi:10.1074/jbc.m702354200

Abstract

The genes encoding the nicotinic acetylcholine receptor alpha3, alpha5, and beta4 subunits are genomically clustered. These genes are co-expressed in a variety of cells in the peripheral and central nervous systems. Their gene products assemble in a number of stoichiometries to generate several nicotinic receptor subtypes that have distinct pharmacological and physiological properties. Signaling through these receptors is critical for a variety of fundamental biological processes. Despite their importance, the transcriptional mechanisms underlying their coordinated expression remain to be completely elucidated. By using a bioinformatics approach, we identified a highly conserved intronic sequence within the fifth intron of the alpha3 subunit gene. Reporter gene analysis demonstrated that this sequence, termed "alpha3 intron 5," inhibits the transcriptional activities of the alpha3 and beta4 subunit gene promoters. This repressive activity is position- and orientation-independent. Importantly, repression occurs in a cell type-specific manner, being present in cells that do not express the receptor genes or expresses them at very low levels. Electrophoretic mobility shift assays demonstrate that nuclear proteins specifically interact with alpha3 intron 5 at two distinct sites. We propose that this intronic repressor element is important for the restricted expression patterns of the nicotinic receptor alpha3 and beta4 subunit genes.

Highlights

We show that this intronic sequence is capable of repressing the transcriptional activities of the ␣3 and ␤4 subunit gene promoters and does so in a cell type-specific manner
Again as expected, there was relatively little conservation (Ͻ50%) between intronic sequences (Fig. 1b) with one notable exception: there was a striking degree of conservation (Ͼ50% identity) between an ϳ1-kb region of the fifth intron of the human and mouse ␣3 subunit genes (Fig. 1b)
As at least 12 genes encode nicotinic receptor subunits expressed in the nervous system, there is the potential for a substantial number of functionally distinct receptor subtypes

Summary

Introduction

The genes exhibit overlapping expression patterns, these patterns are not identical either temporally or spatially suggesting that in addition to a possible common regulatory mechanism, these genes are most likely subject to independent regulation as well. In this regard, it is important to note that a growing body of evidence suggests that the ␣3 and ␤4 subunit genes may be coordinately expressed, whereas the ␣5 gene may be independently regulated (47, 49 –51). We show that the intronic sequence directly interacts with nuclear proteins

Methods

Results

Conclusion