Isolation and functional characterization of the 5 '-upstream region of mouse P/Q-type Ca2+ channel alpha(1A) subunit gene

The omega-agatoxin-IVA-sensitive P/Q;type Ca2+ channel is predominantly exp ressed in the nervous system. To dissect the molecular mechanisms underlyin g the neuron-specific expression of the P/Q-type channel, we have isolated and characterized the 5'-upstream region of the mouse cu,, subunit gene, A transcription start site appeared to exist at -269 bp upstream from the sta rt codon as found by 5' RACE analysis. The proximal promoter of the alpha(1 A) subunit gene lacks a typical TATA box, but contains several transcriptio n factor binding sequences, including two Spl sites. When linked to a place ntal alkaline phosphatase (PLAP) reporter gene to examine the promoter acti vity, the 6.3-kb (-6,273 to +269) 5'-upstream region, but not a smaller 3.0 -kb construct (-3,021 to +269), was able to drive the reporter gene in neur on-like PC12 cells. In contrast, neither of these constructs enhanced the F LAP expression in fibroblast NIH3T3 cells. The sequence between 6.3 and 3.9 kb of the 5'-upstream region did not show promoter activity in either of t he cell lines, but enhanced TK promoter activity in PC12 cells, though not in NIH3T3 cells. These results suggest that neuron-specific elements of the alpha(1A), subunit gene are likely to be located in the dietal upstream re gions (-6,273 to -3,021) of the 5'-upstream sequence. (C) 1999 Academic Pre ss.