Neural specific expression of the m4 muscarinic acetylcholine receptor gene is mediated by a RE1/NRSE-type silencing element.
Muscarinic receptor genes are members of the G-protein receptor superfamily that, with the inclusion of the odorant receptors, is believed to contain over a thousand members. Each member of this superfamily, which has been studied to date, appears to have a distinct pattern of expression, but little work has been done on the regulation of these complex expression patterns. We have recently isolated the rat m4 muscarinic receptor gene and identified a genomic 1520-nucleotide sequence that appeared capable of directing cell-specific expression (Wood, I. C., Roopra. A., Harrington, C., and Buckley, N. J. (1995) J. Biol. Chem. 270, 30933-30940). In the present study we have constructed a set of deletion promoter constructs to more closely define the DNA elements that are responsible for m4 gene expression. We have found that deletion of a RE1/NRSE silencer element between nucleotides -574 and -550, similar to that found in other neural specific genes, results in activation of reporter expression in non-m4-expressing cells. Gel mobility shift analysis has shown that a protein present in nonexpressing cells is capable of binding to this element and is probably the recently identified neural silencer, REST/NRSF. Of the constitutively active proximal promoter only a tandem Sp-1 site appears to recruit DNA binding proteins that are present in all cells tested. This represents the first report documenting the role of this silencer in regulating expression of a member of the G-protein receptor family.
Wood
I C
IC
Wellcome Laboratory for Molecular Pharmacology, University College London, London, WC1E 6BT, United Kingdom.
Roopra
A
A
Buckley
N J
NJ
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002645:Chickens; D006224:Cricetinae; D005786:Gene Expression Regulation; D005910:Glioma; D006801:Humans; D006822:Hybrid Cells; D051379:Mice; D008969:Molecular Sequence Data; D009447:Neuroblastoma; D009474:Neurons / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D051381:Rats; D043588:Receptor, Muscarinic M4; D011976:Receptors, Muscarinic / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D017384:Sequence Deletion; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D043588:Receptor, Muscarinic M4; D011976:Receptors, Muscarinic; D011994:Recombinant Proteins