Is RE1/NRSE a Common cis-Regulatory Sequence for ChAT and VAChT Genes?

Stéphanie De Gois,Leı̈La Houhou,Yoshio Oda,Marilys Corbex,Fabrice Pajak,Etienne Thévenot,Guilan Vodjdani,Jacques Mallet,Sylvie Berrard

doi:10.1074/jbc.m006895200

Abstract

Choline acetyltransferase (ChAT), the biosynthetic enzyme of acetylcholine, and the vesicular acetylcholine transporter (VAChT) are both required for cholinergic neurotransmission. These proteins are encoded by two embedded genes, the VAChT gene lying within the first intron of the ChAT gene. In the nervous system, both ChAT and VAChT are synthesized only in cholinergic neurons, and it is therefore likely that the cell type-specific expression of their genes is coordinately regulated. It has been suggested that a 2336-base pair genomic region upstream from the ChAT and VAChT coding sequences drives ChAT gene expression in cholinergic structures. We investigated whether this region also regulates VAChT gene transcription. Transfection assays showed that this region strongly represses the activity of the native VAChT promoters in non-neuronal cells, but has no major effect in neuronal cells whether or not they express the endogenous ChAT and VAChT genes. The silencer activity of this region is mediated solely by a repressor element 1 or neuron-restrictive silencer element (RE1/NRSE). Moreover, several proteins, including RE1-silencing transcription factor or neuron-restrictive silencer factor, are recruited by this regulatory sequence. These data suggest that this upstream region and RE1/NRSE co-regulate the expression of the ChAT and VAChT genes.

Highlights

In cholinergic neurons, the neurotransmitter acetylcholine is synthesized by choline acetyltransferase (ChAT)1 and is translocated into synaptic vesicles by the vesicular acetylcholine transporter (VAChT)
Various data suggest that a 2336-bp region just upstream and designated region 1 (Fig. 2) regulates the cholinergic neuronspecific expression of the Choline acetyltransferase (ChAT) gene. (i) In non-neuronal cells, in vitro, this region is able to repress the activity of the ChAT/ VAChT promoter region 2, as well as that of a heterologous promoter [8, 24]; (ii) in transgenic mice, region 1 drove the expression of a downstream heterologous promoter in various cholinergic structures [25]; (iii) the pattern of transgene expression in the brain of these mice paralleled both qualitatively and quantitatively that of endogenous ChAT mRNAs; and (iv) in spinal cord, transgene expression was developmentally regulated, to the endogenous ChAT gene
Region 1 Is Involved in VAChT Gene Cell Type-specific Expression—Our previous studies have shown that the rat VAChT gene can be transcribed from two distinct domains of the cholinergic gene locus: (i) one upstream from exon R, which promotes the transcription of the ChAT gene [3, 11]; (ii) one downstream from exon R, which contains two distinct promoters for the VAChT gene

Summary

Introduction

The neurotransmitter acetylcholine is synthesized by choline acetyltransferase (ChAT)1 and is translocated into synaptic vesicles by the vesicular acetylcholine transporter (VAChT). The RE1/NRSE of region 1, in vitro, represses the expression of a heterologous downstream promoter in non-neuronal cells by binding REST/NRSF [24]. We used transient transfections to examine the potential involvement of a 5Ј upstream domain, region 1, in the regulation of the cell type-specific expression of the VAChT gene.

Results

Conclusion