Neurotoxic Mutants of the Prion Protein Induce Spontaneous Ionic Currents in Cultured Cells

Abstract

The mechanisms by which prions kill neurons and the role of the cellular prion protein in this process are enigmatic. Insight into these questions is provided by the neurodegenerative phenotypes of transgenic mice expressing prion protein (PrP) molecules with deletions of conserved amino acids in the central region. We report here that expression in transfected cells of the most toxic of these PrP deletion mutants (⌬105-125) induces large, spontaneous ionic currents that can be detected by patchclamping techniques. These currents are produced by relatively non-selective, cation-permeable channels or pores in the cell membrane and can be silenced by overexpression of wild-type PrP, as well as by treatment with a sulfated glycosaminoglycan. Similar currents are induced by PrP molecules carrying several different point mutations in the central region that cause familial prion diseases in humans. The ionic currents described here are distinct from those produced in artificial lipid membranes by synthetic peptides derived from the PrP sequence because they are induced by membrane-anchored forms of PrP that are synthesized by cells and that are found in vivo. Our results indicate that the neurotoxicity of some mutant forms of PrP is attributable to enhanced ion channel activity and that wild-type PrP possesses a channel-silencing activity. Drugs that block PrP-associated channels or pores may therefore represent novel therapeutic agents for treatment of patients with prion diseases.

A C-terminal Membrane Anchor Affects the Interactions of Prion Proteins with Lipid Membranes

“…Interestingly, ⌬CRPrP was found to contain the full set of posttranslational modifications (N-glycosylations and GPI attachment) typical of wild-type PrP C (49). Expression of ⌬CRPrP in transfected cell lines was found to induce spontaneous ionic currents, and the same was true for PrPs harboring several point mutations in the central hydrophobic region that are linked to familial prion diseases in humans (37,38). These ionic currents can be silenced SCHEME 1.…”

“…9B). In previous experiments based on electrophysiological measurements with neuronal cells expressing ⌬CR_PrP, Solomon et al (37,38) already surmised that ⌬CR_PrP might form ion channel-like assemblies.…”

Knockdown of prion protein (PrP) by RNA interference weakens the protective activity of wild-type PrP against copper ion and antagonizes the cytotoxicity of fCJD-associated PrP mutants in cultured cells

“…transgenic mice expressing insert mutants of PrP develop prion disease with accumulation of detergent-insoluble, protease-resistant PrP in the brain. When expressed in cell lines, PrP with additional repeats displays detergent insolubility, resistance to proteinase K digestion similar to PrP Sc , and alters cell surface expression and hinders export to the cell surface, leading to obvious cytotoxicity (24,36). The significant antagonizing activity of PrP-specific sirnas on the octarepeats insertion-induced cytotoxicity illustrates not only the molecular mechanism of the mutants, but also the potential to use those sirnas for eliminating the relevant neurotoxicity.…”

Cytoprotective and toxic functions of the prion protein family

“…Even though the expression of these mutants causes severe toxicity in PrP null transgenic mice (33)(34)(35) and in CGN cultures (64,97), they show reduced effect (43,98), or no effect (42,96) when expressed in immortalized cell lines. The most obvious phenotype associated with the Shmerling or CR mutations in PrP-s in cell models is the drug hypersensitivity and the occurrence of spontaneous inward cationic currents (37,39). Strong evidence supports that the same functional changes of PrPCR is behind these two phenotypes detected in different models (40).…”

“…PrPCR and several other HD mutants have also been found to cause spontaneous inward ionic currents through the plasma membranes of various cell types (39,40), which activities were observed also by PrP-s harbouring familiar TSE-related point mutations. These effects, although not being directly toxic, were also abrogated by the co-expression of WT PrP.…”