The overall goal of this project is to investigate the pathogenesis of prion diseases using transgenic mouse models. Our objective is to understand the molecular and cellular mechanisms by which prions kill neurons and damage the central nervous system. During the last funding period, we investigated how three, distinct genetic mechanisms (loss, gain, and subversion of the normal function of PrPC) contribute to neurodegeneration in two different transgenic models: Tg(PG14) mice, which express an aggregation-prone PrP harboring a nine-octapeptide insertional mutation linked to familial Creutzfeldt-Jakob disease;and Tg(CR) mice, which express a highly neurotoxic form of PrP with a 21-amino acid deletion (residues 105-125) in the conserved, central region of the protein. In the course of our previous work, we made three key discoveries that form the basis for this renewal application. First, we found that mutations in the central region of PrP, including deletions such as CR, as well as point mutations associated with familial prion diseases of humans, induce a powerful ion channel activity that can be observed by patch-clamping techniques. Second, we have identified a group of PrP ligands that inhibit the ion channel activity of mutant PrP. Interestingly, most of these ligands also inhibit the formation of PrPSc, in cell-free systems or mice, suggesting that common structural domains control both the functional activity of PrP and its conversion to PrPSc. Third, we observed that a positively-charged, nine amino acid domain at the N-terminus plays an essential role in the channel-forming and neuroprotective properties of PrP molecules. Taken together, these new findings suggest the hypothesis that the pathogenesis of some inherited prion diseases, and possibly of infectiously acquired cases as well, may be due to PrP-related channel activity. Moreover, these pathogenic effects may be determined by a surprisingly short domain of the PrP primary sequence that controls both ion channel activity and conformational misfolding of the protein. To pursue these observations, we plan to (1) Analyze transgenic models of familial prion diseases associated with a "channel-inducing" PrP mutations;(2) Determine whether PrPSc induces PrPC-dependent, ion channel activation;(3) Test whether inhibitors of PrP-related ion channel activity have therapeutic effect in mouse models of familial and infectious prion diseases;and (4) Investigate the function of the N-terminal polybasic domain of PrP a critical neurotoxicity determinant. This project addresses a major gap in knowledge concerning the mechanisms by which prions are neurotoxic. It explores specific molecular and cellular pathways that may be activated by prions, and that may a role in their pathogenic effects. The proposal ties prion diseases to other neurodegenerative conditions due to abnormal activity of ion channels, and it sets the stage for treating prion diseases by blocking specific neurotoxic pathways in addition to prion propagation.

Public Health Relevance

Prion diseases are fatal neurodegenerative disorders of humans and animals that pose a grave threat to public health, and endanger the safety of the food, blood and organ supplies. This grant application utilizes genetically engineered mice to explore the mechanisms by which prions kill nerve cells and damage the brain. The project sets the stage for development of novel therapeutic approaches based on blocking specific neurotoxic pathways.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS040975-14
Application #
8492171
Study Section
Cellular and Molecular Biology of Neurodegeneration Study Section (CMND)
Program Officer
Wong, May
Project Start
2001-01-15
Project End
2016-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
14
Fiscal Year
2013
Total Cost
$449,317
Indirect Cost
$174,841
Name
Boston University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
Imberdis, Thibaut; Harris, David A (2014) Prion permissive pathways: extracellular matrix genes control susceptibility to prion infection. EMBO J 33:1506-8
Fluharty, Brian R; Biasini, Emiliano; Stravalaci, Matteo et al. (2013) An N-terminal fragment of the prion protein binds to amyloid-ýý oligomers and inhibits their neurotoxicity in vivo. J Biol Chem 288:7857-66
Biasini, Emiliano; Unterberger, Ursula; Solomon, Isaac H et al. (2013) A mutant prion protein sensitizes neurons to glutamate-induced excitotoxicity. J Neurosci 33:2408-18
Biasini, Emiliano; Turnbaugh, Jessie A; Massignan, Tania et al. (2012) The toxicity of a mutant prion protein is cell-autonomous, and can be suppressed by wild-type prion protein on adjacent cells. PLoS One 7:e33472
Solomon, Isaac H; Biasini, Emiliano; Harris, David A (2012) Ion channels induced by the prion protein: mediators of neurotoxicity. Prion 6:40-5
Biasini, Emiliano; Turnbaugh, Jessie A; Unterberger, Ursula et al. (2012) Prion protein at the crossroads of physiology and disease. Trends Neurosci 35:92-103
Westergard, Laura; Turnbaugh, Jessie A; Harris, David A (2011) A nine amino acid domain is essential for mutant prion protein toxicity. J Neurosci 31:14005-17
Solomon, Isaac H; Khatri, Natasha; Biasini, Emiliano et al. (2011) An N-terminal polybasic domain and cell surface localization are required for mutant prion protein toxicity. J Biol Chem 286:14724-36
Westergard, Laura; Turnbaugh, Jessie A; Harris, David A (2011) A naturally occurring C-terminal fragment of the prion protein (PrP) delays disease and acts as a dominant-negative inhibitor of PrPSc formation. J Biol Chem 286:44234-42
Massignan, Tania; Biasini, Emiliano; Harris, David A (2011) A Drug-Based Cellular Assay (DBCA) for studying cytotoxic and cytoprotective activities of the prion protein: A practical guide. Methods 53:214-9

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