Genetic Mapping of Activity Determinants within Cellular Prion Proteins: N-terminal modules in PrP super(C) offset pro-apoptotic activity of the Doppel helix B/B' region

• Published in: The Journal of biological chemistry 2004-12, Vol.279 (53), p.55443-55454
• Main Authors: Drisaldi, Bettina, Coomaraswamy, Janaky, Mastrangelo, Peter, Strome, Bob, Yang, Jing, Watts, Joel C, Chishti, MAzhar, Marvi, Melissa, Windl, Otto, Ahrens, Rosemary, Major, Francois, Sy, Man-Sun, Kretzschmar, Hans, Fraser, Paul E, Mount, Howard TJ, Westaway, David
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X

The PrP-like Doppel (Dpl) protein causes apoptotic death of cerebellar neurons in transgenic mice, a process prevented by expression of the wild type (wt) cellular prion protein, PrP super(C). Internally deleted forms of PrP super(C) resembling Dpl such as PrPdelta32-121 produce a similar PrP super(C)-sensitive pro-apoptotic phenotype in transgenic mice. Here we demonstrate that these phenotypic attributes of wt Dpl, wt PrP super(C), and PrPdelta132-121 can be accurately recapitulated by transfected mouse cerebellar granule cell cultures. This system was then explored by mutagenesis of the co-expressed prion proteins to reveal functional determinants. By this means, neuroprotective activity of wt PrP super(C) was shown to be nullified by a deletion of the N-terminal charged region implicated in endocytosis and retrograde axonal transport (PrPdelta23-28), by deletion of all five octarepeats (PrPdelta51-90), or by glycine replacement of four octarepeat histidine residues required for selective binding of copper ions (Prnp"H/G"). In the case of Dpl, overlapping deletions defined a requirement for the gene interval encoding helices B and B' (Dpldelta101-125). These data suggest contributions of copper binding and neuronal trafficking to wt PrP super(C) function in vivo and place constraints upon current hypotheses to explain Dpl/PrP super(C) antagonism by competitive ligand binding. Further implementation of this assay should provide a fuller understanding of the attributes and subcellular localizations required for activity of these enigmatic proteins.