Octarepeat region flexibility impacts prion function, endoproteolysis and disease manifestation.

Agnes Lau,Beipei Shi,Jing Yang,Aru Balachandran,Jennifer Grams,Gerold Schmitt‐Ulms,Chae Kim,Robin Aglietti,Holger Wille,Serene Wohlgemuth,David Westaway,Hristina Gapeshina,Nathalie Daude,Jacques Van Der Merwe,Robert C.c Mercer ,Alex J Mcdonald ,Glenn L Millhauser ,Éric Walter ,Charles E Mays ,Jiri Safar

doi:10.15252/emmm.201404588

Abstract

The cellular prion protein (PrPC) comprises a natively unstructured N-terminal domain, including a metal-binding octarepeat region (OR) and a linker, followed by a C-terminal domain that misfolds to form PrPSc in Creutzfeldt-Jakob disease. PrPC β-endoproteolysis to the C2 fragment allows PrPSc formation, while α-endoproteolysis blocks production. To examine the OR, we used structure-directed design to make novel alleles, ‘S1’ and ‘S3’, locking this region in extended or compact conformations, respectively. S1 and S3 PrP resembled WT PrP in supporting peripheral nerve myelination. Prion-infected S1 and S3 transgenic mice both accumulated similar low levels of PrPSc and infectious prion particles, but differed in their clinical presentation. Unexpectedly, S3 PrP overproduced C2 fragment in the brain by a mechanism distinct from metal-catalysed hydrolysis reported previously. OR flexibility is concluded to impact diverse biological endpoints; it is a salient variable in infectious disease paradigms and modulates how the levels of PrPSc and infectivity can either uncouple or engage to drive the onset of clinical disease.

Highlights

Prion diseases such as Creutzfeldt-Jakob disease (CJD) and bovine spongiform encephalopathy (BSE) are transmissible neurodegenerative disorders
Using the C-terminal antibody Sha31, electrophoretic profiles of proteinase K (PK)-digested PrP species from brain homogenates of the transgenic mice were similar to the RML inoculum; this resemblance remained after incubation with the glycosidase PNGaseF, when a single band was present in all three genotypes (Fig 5A and B)
Since our studies define octarepeat region (OR) flexibility as impacting diverse biological endpoints, we suggest it will prove to be an important variable in infectious disease paradigms where levels of PrPSc and infectivity appear dissociated from the onset of clinical disease

Summary

Introduction

Prion diseases such as Creutzfeldt-Jakob disease (CJD) and bovine spongiform encephalopathy (BSE) are transmissible neurodegenerative disorders. A-secretase processing of APP by a disintegrin and metalloproteases (ADAM) generates a benign peptide called P3 and prevents amyloid formation. PrPC is subject to analogous endoproteolysis events (Fig 1A); a-cleavage of PrPC occurs adjacent to the linker region at residue 110 to generate a fragment called C1 and is attributed to the action of an ADAM protease (Chen et al, 1995; Jimenez-Huete et al, 1998; Vincent et al, 2000, 2001; LaffontProust et al, 2005; Walmsley et al, 2009). A-cleavage of PrPC serves a benign role insofar as formation of C1 precludes the formation of PrPSc upon exposure to infectious prion inocula as the protease-resistant core of PrPSc called PrP27-30 is longer, starting in

C Wildtype

Results

Materials and Methods