NR AQQM
AU Yadavalli,R.; Guttmann,R.P.; Seward,T.; Centers,A.P.; Williamson,R.A.; Telling,G.C.
TI Calpain-dependent endoproteolytic cleavage of PrPsc modulates scrapie prion propagation
QU The Journal of Biological Chemistry 2004 May 21; 279(21): 21948-56
PT journal article
AB Previous studies using post-mortem human brain extracts demonstrated that PrP in Creutzfeldt-Jakob disease (CJD) brains is cleaved by a cellular protease to generate a C-terminal fragment, referred to as C2, which has the same molecular weight as PrP-(27-30), the protease-resistant core of PrPsc (1). The role of this endoproteolytic cleavage of PrP in prion pathogenesis and the identity of the cellular protease responsible for production of the C2 cleavage product has not been explored. To address these issues we have taken a combination of pharmacological and genetic approaches using persistently infected scrapie mouse brain (SMB) cells. We confirm that production of C2 is the predominant cleavage event of PrPsc in the brains of scrapie-infected mice and that SMB cells faithfully recapitulate the diverse intracellular proteolytic processing events of PrPsc and PrPc observed in vivo. While increases in intracellular calcium (Ca(2+)) levels in prion-infected cell cultures stimulate the production of the PrPsc cleavage product, pharmacological inhibitors of calpains and overexpression of the endogenous calpain inhibitor, calpastatin, prevent the production of C2. In contrast, inhibitors of lysosomal proteases, caspases, and the proteasome have no effect on C2 production in SMB cells. Calpain inhibition also prevents the accumulation of PrPsc in SMB and persistently infected ScN2A cells, whereas bioassay of inhibitor-treated cell cultures demonstrates that calpain inhibition results in reduced prion titers compared with control-treated cultures assessed in parallel. Our observations suggest that calpain-mediated endoproteolytic cleavage of PrPsc may be an important event in prion propagation.
MH Animals; Biological Assay; Brain/metabolism; Calcium/chemistry; Calcium-Binding Proteins/pharmacology; Calpain/chemistry/*metabolism; Cell Division; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Ionophores/pharmacology; Kinetics; Mice; PrPc Proteins/metabolism; PrPsc Proteins/*metabolism; Prions/*chemistry; Protein Structure, Secondary; Protein Structure, Tertiary; Scrapie/*metabolism; Support, U.S. Gov't, P.H.S.; Time Factors
AD Rajgopal Yadavalli, Rodney P. Guttmann, Tanya Seward, Adrian P. Centers, Glenn C. Telling (gtell2@uky.edu), Sanders Brown Center on Aging, University of Kentucky, Lexington, Kentucky 40536; Rodney P. Guttmann§, Departmens of Physiology, University of Kentucky, Lexington, Kentucky 40536; R. Anthony Williamson, Department of Immunology, The Scripps Research Institute, La Jolla, California 92037; Glenn C. Telling, Departments of Microbiology, Immunology and Molecular Genetics, Neurology, University of Kentucky, Lexington, Kentucky 40536
SP englisch
PO USA