The mouse calretinin gene promoter region: structural and functional components

Abstract

The 5′ flanking region of the mouse calretinin gene was cloned and a 1.8 kbp region adjacent to exon 1 was sequenced. Putative upstream promoter and enhancer elements were identified, including appropriately positioned TATA and CAAT boxes (positions −50 and −68, respectively). There was considerable sequence and structural homology between mouse and human upstream elements. Neuron-restrictive activity was demonstrated via transfection of calretinin promoter-reporter constructs into primary embryonic mouse brain cultures expressing calretinin. In promoterless reporter constructs, the proximal upstream 1.5 kbp of the mouse calretinin gene boosted luciferase activity (up to 100-fold) exclusively in the neuronal population. Deletion analysis revealed the minimal promoter to be within the 95-bp proximal to the transcription start site. Transfections with SV40 promoter constructs in these cultures resulted in reporter gene expression predominantly in non-neuronal cells. Inserting the proximal 1.5 kbp of mouse calretinin upstream in SV40 promoter-reporter constructs reduced luciferase activity. Thus, calretinin upstream sequences increased reporter expression in cultured neurons and decreased expression from the SV40 promoter in non-neuronal cultured brain cells. The calretinin promoter contained relevant regulatory element consensus motifs and demonstrated in vitro neuron-restrictive bioactivity.