Retinoic acid stimulates α-CAMKII gene expression in PC12 cells at a distinct transcription initiation site

Abstract

The promoter region of the α-subunit of the calcium/calmodulin- dependent protein kinase II (α-CaMKII) gene was inserted into a β- galactosidase (β-gal) reporter plasmid, and β-gal activities were examined in neuroblastoma (NB2a) and pheochromocytoma (PC12) cells after transient or stable transfections. The α-CaMKII promoter was 12- to 45-fold more active in NB2a compared with PC12 cells after transient or stable transfections. All-trans retinoic acid (RA) stimulated reporter gene expression at both protein and mRNA levels in transfected PC12 cells. RA increased the level of endogenous α-CaMKII mRNA in untransfected PC12 cells by 4.4-fold. The transcription initiation site(s) (TIS) of the α-CaMKII gene in PC12 cells and rat brain was examined by RNase protection assays (RPA) and reverse transcriptase PCRs. The TIS for the α-CaMKll/β-gal reporter gene in transfected PC12 cells was indistinguishable from the TIS. 1 in rat hippocampus. In contrast, the only detectable TIS for the α-CaMKII gene in untransfected PC12 cells was located near the ATG translation start codon, 147 nucleotides 3' to TIS. 1 in hippocampus. This unusual TIS was also the predominant TIS in rat cerebellum. These results suggest that the αCaMKII promoter may contain sequences that respond directly or indirectly to RA. In addition, the unusual TIS of the α-CaMKII gene in PC12 cells and rat cerebellum may contribute to the very low expression of this gene compared with that in hippocampus.