The Comprehensive Native Interactome of a Fully Functional Tagged Prion Protein

Abstract

The enumeration of the interaction partners of the cellular prion protein, PrP^C, may help clarifying its elusive molecular function. Here we added a carboxy proximal myc epitope tag to PrP^C. When expressed in transgenic mice, PrP_myc carried a GPI anchor, was targeted to lipid rafts, and was glycosylated similarly to PrP^C. PrP_myc antagonized the toxicity of truncated PrP, restored prion infectibility of PrP^C-deficient mice, and was physically incorporated into PrP^Sc aggregates, indicating that it possessed all functional characteristics of genuine PrP^C. We then immunopurified myc epitope-containing protein complexes from PrP_myc transgenic mouse brains. Gentle differential elution with epitope-mimetic decapeptides, or a scrambled version thereof, yielded 96 specifically released proteins. Quantitative mass spectrometry with isotope-coded tags identified seven proteins which co-eluted equimolarly with PrP^C and may represent component of a multiprotein complex. Selected PrP^C interactors were validated using independent methods. Several of these proteins appear to exert functions in axomyelinic maintenance.