Plasminogen stimulates propagation of protease-resistant prion protein in vitro

Abstract

To clarify the role of plasminogen as a cofactor for prion propagation, we conducted functional assays using a cell-free prion protein (PrP) conversion assay termed protein misfolding cyclic amplification (PMCA) and prion-infected cell lines. Here, we report that plasminogen stimulates propagation of the protease-resistant scrapie PrP (PrPSc). Compared to control PMCA conducted without plasminogen, addition of plasminogen in PMCA using wild-type brain material significantly increased PrP conversion, with an EC50 = similar to 56 nM. PrP conversion in PMCA was substantially less efficient with plasminogen-deficient brain material than with wild-type material. The activity stimulating PrP conversion was specific for plasminogen and conserved in its kringle domains. Such activity was abrogated by modification of plasminogen structure and interference of PrP-plasminogen interaction. Kinetic analysis of PrPSc generation demonstrated that the presence of plasminogen in PMCA enhanced the PrPSc production rate to similar to 0.97U/mu l/h and reduced turnover time to similar to 1 h compared to those (similar to 0.4 U/mu l/h and similar to 2.5 h) obtained without supplementation. Furthermore, as observed in PMCA, plasminogen and kringles promoted PrPSc propagation in ScN2a and Elk 21(+) cells. Our results demonstrate that plasminogen functions in stimulating conversion processes and represents the first cellular protein cofactor that enhances the hypothetical mechanism of prion propagation.-Mays, C. E., Ryou, C. Plasminogen stimulates propagation of protease-resistant prion protein in vitro. FASEB J. 24, 5102-5112 (2010). www.fasebj.org