Genomic organization and neuronal cell type specific promoter activity of β isoform of Ca 2+/calmodulin dependent protein kinase II of rat brain

Abstract

The gene encoding the β isoform of rat Ca 2+/calmodulin-dependent protein kinase II was cloned, and its exon–intron organization was analyzed. The gene consisted of 21 exons spanning more than 80 kilobase pairs and the coding sequence was made up of 20 exons. Each discrete functional unit, such as the ATP-binding site, the autophosphorylation site responsible for Ca 2+-independent activity, the calmodulin binding site, and the link structure, was encoded by a single exon. All splice junction sequences flanking the introns conformed to the consensus splice junction sequence and the GT–AG splice rule. The site of transcription initiation was −78 bases from the initiation codon as determined by 5′ RACE analysis. The promoter activity of the gene was analyzed using neuroblastomas, as well as non-neuronal cell lines. Neuronal cell type-specific promoter activity was found in the 5′-upstream region −66 to −35 bp from the transcription initiation site. Silence elements were found further upstream at −222 to −123 bp and −576 to −323 bp. A protein bound to the −66 to −35 region was found in the nuclear extract of rat brain, including the cerebellum, forebrain, and brainstem, by gel mobility shift assay.