Genomic organization and promoter analysis of the human nicotinic acetylcholine receptor alpha6 subunit (CHNRA6) gene: Alu and other elements direct transcriptional repression.
Nicotinic acetylcholine receptors (nAChRs) form ligand-gated ion channels involved in fast synaptic transmission. Recently mutations in nAChR genes have been reported in nocturnal frontal lobe epilepsy. We performed molecular analysis on the human neuronal nAChR alpha6 subunit (CHRNA6) gene, a member of nAChR gene family, to understand its role in disease. Genomic analysis revealed that the gene consisted of six exons with an estimated size of 16 kb. We mapped the CHRNA6 gene to chromosome 8p11.21-11.22 by fluorescence in situ hybridization, the same putative region responsible for adolescent-onset idiopathic generalized epilepsy. Examination of the 5'-regulatory region failed to identify either a TATA box or GC-rich sequences, but did highlight tandem Alu sequences, located between 910 and 370 bp upstream from a potential cap site. Analyses of transcriptional activity, performed using nested deletions of the 5'-upstream region, showed that the downstream Alu repeat and another element(s) in the promoter region, function as negative regulators. Further analyses of the tandem Alu repeats, examined by fusing them to a different ion channel gene promoter, confirmed their role as transcriptional repressors regardless of their orientation and copy number. These data may explain the limited expression of the CHRNA6 gene in the brain.
Ebihara
Mitsuru
M
Laboratory for Molecular Psychiatry, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Ohba
Hisako
H
Ohno
Shin-ich
SI
Yoshikawa
Takeo
T
D020087:Alu Elements / Q000235:genetics*; D001483:Base Sequence; D002874:Chromosome Mapping; D002898:Chromosomes, Human, Pair 8 / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000235:genetics; D005786:Gene Expression Regulation; D005796:Genes / Q000235:genetics; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics; D011978:Receptors, Nicotinic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
C470410:CHRNA6 protein, human; D011978:Receptors, Nicotinic; D011993:Recombinant Fusion Proteins; D004247:DNA; D008156:Luciferases