The gene encoding the β subunit of the inhibitory glycine receptor (GlyR) is widely expressed throughout the mammalian central nervous system. To unravel the elements regulating its transcription, we isolated its 5′ non-coding and upstream flanking regions from mouse. Sequence analysis revealed significant differences between the 5′ region of the β subunit gene and the corresponding regions of the homologous GlyR α subunit genes; it also identified a novel exon (exon 0) that encodes most of the 5′-untranslated portion of the GlyR β mRNA. Primer extension experiments disclosed multiple transcriptional start sites. Transfection experiments with luciferase reporter gene constructs showed that sequences encompassing 1.58kb of upstream flanking region and 180bp of exon 0 displayed high promoter activity in two neuroblastoma cell lines but not in non-neural cells. Analysis of various deletion constructs showed that the 5′ flanking region preceding the transcriptional start sites silences expression in non-neural cells but is not essential for general promoter activity. In contrast, the deletion of sequences within exon 0 drastically decreased or abolished transcription; the removal of sequences harbouring Sp1 consensus sequences within exon 0 decreased expression specifically in a neuroblastoma cell line. Band-shift assays confirmed the binding of Sp1 to sites within the deleted sequence. Our results indicate that neural-specific expression of the GlyR β subunit gene might depend on a direct interaction of Sp1 transcription factors with cis elements located downstream from transcription initiation sites.

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