Mutant prion protein acquires resistance to protease in mouse neuroblastoma cells.

Conversion of the cellular isoform of the prion protein (PrP(C)) into the pathogenic isoform (PrP(Sc)) is thought to be the causative event in prion diseases. Biochemically, PrP(Sc) differs from PrP(C) in its partial resistance to proteinase K (PK). The amino acid sequence AGAAAAGA, comprising residues 112-119 of the murine PrP(C), has been shown to be amyloidogenic and evolutionarily conserved. To assess the effect of mutations at and around this hydrophobic sequence on protease resistance, the sequence was replaced either by alanines or by glycines and, in a third mutant, a large part surrounding this region was removed. The PrP mutant carrying substitutions of glycines for alanines showed PK resistance and aberrant proteolytic processing. Tetracycline-induced expression of this mutant indicated that resistance to protease is acquired concurrent with the synthesis of the protein. These findings indicate that mutations in the central hydrophobic region lead to immediate alterations in PrP structure and processing.

Ets1 is a common element in directing transcription of the alpha and beta genes of human protein kinase CK2.

Protein kinase CK2 is a conserved and vital Ser/Thr phosphotransferase with various links to malignant diseases, occurring as a tetramer composed of two catalytically active (CK2alpha and/or CK2alpha') and two regulatory subunits (CK2beta). There is balanced availability of CK2alpha and CK2beta transcripts in proliferating and differentiating cultured cells. Examination of the human CK2beta gene for transcriptionally active regions by systematic deletions and reporter gene assays indicates strong promoter activity at positions -42 to 14 and 12 to 72 containing transcription start sites 1 and 2 of the gene (positions +1 and 33), respectively, an upstream and a downstream enhancer activity at positions -241 to -168 and 123 to 677, respectively, and silencer activity at positions -241 to -261. Of the various transcription factor binding motifs present in those regions, Ets1 and CAAT-related motifs turned out to be of particular importance, Ets1 for promoter activation and CAAT-related motifs for enhancer activation. In addition, there are contributions by Sp1. Most strikingly, the Ets1 region representing two adjoining consensus motifs also occurs with complete identity in the recently characterized promoter of the CK2alpha gene [Krehan, A., Ansuini, H., Böcher, O., Grein, S., Wirkner, U. & Pyerin, W. (2001) J. Biol. Chem. 275, 18327-18336], and affects comparably, when assayed in parallel, the promoters of both CK2 genes, both by motif mutations and by Ets1 overexpression. The data strongly support the hypothesis that Ets1 acts as a common regulatory element of the CK2alpha and CK2beta genes involved in directing coordinate transcription and contributing to the balanced availability of transcripts.