Characterization of the cystatin B gene promoter harboring the dodecamer repeat expanded in progressive myoclonus epilepsy, EPM1.

Mutations in the gene encoding cystatin B (CSTB) are responsible for the primary defect in progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1). A novel and unique type of disease-causing mutation, an unstable dodecamer repeat expansion, accounts for the majority of EPM1 patients world-wide. This minisatellite repeat expansion, located in the putative promoter of CSTB 175 bp upstream from the translation initiation codon, appears to downregulate CSTB gene expression in vivo. We report here the characterization of the CSTB promoter using different promoter-luciferase gene constructs. Transient transfections of cultured mammalian cells suggest that the region from -670 to -1 bp from the translation initiation codon functions as the CSTB promoter. Active binding to five Sp1 and four AP1 sites as well as weak binding to an androgen response element (ARE) half site was demonstrated by electrophoretic mobility shift assays. The effect of the minisatellite expansion on the promoter activity was evaluated by comparing the activity of constructs containing wild-type and expanded alleles. An increase in the number of dodecamer units from three to 19 repeats lowered transcription in vitro by 10-fold. Northern analysis of lymphoblastoid RNA from individuals with 'premutation' length dodecamer repeat (12-17 copies) expansions showed decreased levels of CSTB mRNA expression. These data indicate that expansion of the dodecamer repeat located in the proximal promoter of CSTB severely disrupts the function of the promoter and thereby reduces transcription of CSTB.