Cell-specific expression of the rat secretogranin II promoter.
Secretogranin II (SgII) is a member of the granin family of secretory proteins, which are selectively expressed in neuroendocrine cells. As a first step in understanding the molecular basis for cell type-specific expression of SgII, we isolated a 12-kb clone from a rat genomic library that contained the entire rat SgII coding region, the transcription initiation site, and approximately 3 kb of 5'-flanking region. Within 75 bp of the transcription start site (+1) we located a TATA box and a consensus cAMP responsive element. Within the 5'-flanking region, a number of potential cis-acting elements were identified, including 2 Pit-1 binding sites, 15 E box motifs, and near-perfect matches for AP-1 and AP-2 sites. To demonstrate cell type-specific expression the rat SgII gene, a plasmid containing 2.6 kb of the 5'-flanking region of the SgII gene fused to the luciferase reporter gene (p2774Luc) was transfected into rat pheochromocytoma PC-12 cells, rat pituitary GH4C1 (GH) cells, human BE(2)-M17 (M17) neuroblastoma cells, and mouse fibroblast NIH/3T3 cells. The promoter activity was 6- to 36-fold higher in neuroendocrine cells than in NIH/ 3T3 cells. Progressive deletions in the 5'-flanking region to 61 bp upstream of the start site (p223Luc) had no effect on promoter activity in PC-12 cells. On the other hand, a 5'-deletion in the SgII promoter to -1032 increased promoter activity 3.8-fold in GH cells. This level of expression was maintained when the SgII promoter was further truncated to -189, whereas truncation to -61 resulted in a 2.6-fold reduction in promoter activity. These results suggest that the sequence between -61 and +162 bp is sufficient for SgII promoter activity in PC-12 cells. However, other elements in the 5'-flanking region contribute to both positive and negative regulation of the rat SgII gene in GH cells.
Jones
L C
LC
Department of Pharmacology, University of South Alabama, Mobile 36688, USA.
Day
R N
RN
Pittler
S J
SJ
Valentine
D L
DL
Scammell
J G
JG
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002864:Chromogranins; D003001:Cloning, Molecular; D017353:Gene Deletion; D015870:Gene Expression; D018014:Gene Transfer Techniques; D016678:Genome; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000235:genetics; D016716:PC12 Cells / Q000378:metabolism; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011506:Proteins / Q000235:genetics*; D051381:Rats; D014158:Transcription, Genetic
D002864:Chromogranins; D015345:Oligonucleotide Probes; D011506:Proteins