Table of Contents

Volume 4, Issue 1

  January/February/March 2010

  Pages 1 - 52

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COMMENTARIES & VIEWS

Open Access Article

Exploring prion protein biology in flies: Genetics and beyond

Diego E. Rincon-Limas, Sergio Casas-Tinto and Pedro Fernandez-Funez
Pages 1 - 8
http://dx.doi.org/10.4161/pri.4.1.10504
Abstract | PDF

The fruit fly Drosophila melanogaster has been a favored tool for genetic studies for over 100 years and has become an excellent model system to study development, signal transduction, cell biology, immunity, and behavior. The relevance of Drosophila to humans is perhaps best illustrated by the fact that more than 75% of the genes identified in human diseases have counterparts in Drosophila. During the last decade, many fly models of neurodegenerative disorders have contributed to the identification of novel pathways mediating pathogenesis. However, the development of prion disease models in flies has been remarkably challenging. We recently reported a Drosophila model of sporadic prion pathology that shares relevant features with the typical disease in mammals. This new model provides the basis to explore relevant aspects of the biology of the prion protein, such as uncovering the genetic mechanisms regulating prion protein misfolding and prion-induced neurodegeneration, in a dynamic, genetically tractable in vivo system.

Open Access Article

New insights into the molecular mechanism of amyloid formation from cysteine scanning

Li Fei and Sarah Perrett
Pages 9 - 12
http://dx.doi.org/10.4161/pri.4.1.10670
Abstract | PDF

Our laboratory recently reported the identification of a peptide region, QVNI, within the prion domain of the yeast protein Ure2 that may act as an initiation point for fibril formation.1 This potential amyloid-forming region, which corresponds to residues 18-21 of Ure2, was initially identified by systematic cysteine scanning of the Ure2 prion domain. The point mutant R17C, and the corresponding octapeptide CQVNIGNR, were found to form fibrils rapidly under oxidative conditions due to the formation of a disulfide bond. Deletions within the QVNI sequence cause the fibril formation ability of R17C Ure2 to be lost. The aggregation propensity of this region is strongly modulated by its preceding residue: replacement of R17 with a hydrophobic residue promotes fibril formation in both full-length Ure2 and in the corresponding octapeptides. The wild-type octapeptide, RQVNIGNR, also forms fibrils, and is the shortest amyloid-forming peptide found for Ure2 to date. Interestingly, the wild-type octapeptide crystallizes readily and so provides a starting point towards obtaining high resolution structural information for the amyloid core of Ure2 fibrils.

Open Access Article

Insights into prion protein function from atomistic simulations

Miroslav Hodak and Jerzy Bernholc
Pages 13 - 19
http://dx.doi.org/10.4161/pri.4.1.10969
Abstract | PDF

Computer simulations are a powerful tool for studies of biological systems. They have often been used to study prion protein (PrP), a protein responsible for neurodegenerative diseases, which include “mad cow disease” in cattle and Creutzfeldt-Jacob disease in humans. An important aspect of the prion protein is its interaction with copper ion, which is thought to be relevant for PrP’s yet undetermined function and also potentially play a role in prion diseases. For studies of copper attachment to the prion protein, computer simulations have often been used to complement experimental data and to obtain binding structures of Cu-PrP complexes. This paper summarizes the results of recent ab initio calculations of copper-prion protein interactions focusing on the recently discovered concentration-dependent binding modes in the octarepeat region of this protein. In addition to determining the binding structures, computer simulations were also used to make predictions about PrP’s function and the role of copper in prion diseases. The results demonstrate the predictive power and applicability of ab initio simulations for studies of metal-biomolecular complexes.

Open Access Article

Fibril fragmentation in amyloid assembly and cytotoxicity: When size matters

Wei-Feng Xue, Andrew L. Hellewell, Eric W. Hewitt and Sheena E. Radford
Pages 20 - 25
http://dx.doi.org/10.4161/pri.4.1.11378
Abstract | PDF

Amyloid assemblies are associated with several debilitating human disorders. Understanding the intra- and extracellular assembly of normally soluble proteins and peptides into amyloid aggregates and how they disrupt normal cellular functions is therefore of paramount importance. In a recent report, we demonstrated a striking relationship between reduced fibril length caused by fibril fragmentation and enhanced ability of fibril samples to disrupt membranes and to reduce cell viability. These findings have important implications for our understanding of amyloid disease in that changes in the physical dimensions of fibrils, without parallel changes in their composition or molecular architecture, could be sufficient to alter the biological responses to their presence. These conclusions provide a new hypothesis that the physical dimensions and surface interactions of fibrils play key roles in amyloid disease. Controlling fibril length and stability toward fracturing, and thereby the biological availability of fibril material, may provide a new target for future therapeutic strategies towards combating amyloid disease.




SHORT COMMUNICATIONS

Open Access Article

Pyrroloquinoline quinone inhibits the fibrillation of amyloid proteins

Jihoon Kim, Masaki Kobayashi, Makoto Fukuda, Daisuke Ogasawara, Natsuki Kobayashi, Sung woong Han, Chikashi Nakamura, Masaki Inada, Chisato Miyaura, Kazunori Ikebukuro and Koji Sode
Pages 26 - 31
http://dx.doi.org/10.4161/pri.4.1.10889
Abstract | PDF

Several neurodegenerative diseases involve the selective damage of neuron cells resulting from the accumulation of amyloid fibril formation. Considering that the formation of amyloid fibrils as well as their precursor oligomers is cytotoxic, the agents that prevent the formation of oligomers and/or fibrils might allow the development of a novel therapeutic approach to neurodegenerative diseases. Here, we show pyrroloquinoline quinone (PQQ) inhibits the amyloid fibril formation of the amyloid proteins, amyloid β (1-42) and mouse prion protein. The fibril formation of mouse prion protein in the presence of PQQ was dramatically prevented. Similarly, the fibril formation of amyloid β (1-42) also decreased. With further advanced pharmacological approaches, PQQ may become a leading anti-neurodegenerative compound in the treatment of neurodegenerative diseases.

Open Access Article

Mouse neuronal cells expressing exogenous bovine PRNP and simultaneous downregulation of endogenous mouse PRNP using siRNAs

Sang-Gyun Kang, Yu Mi Roh, Mi Lan Kang, Yong-Sun Kim and Han Sang Yoo
Pages 32 - 37
http://dx.doi.org/10.4161/pri.4.1.11218
Abstract | PDF

Prion diseases, which are called transmissible spongiform encephalopathies (TSE), comprise a group of fatal infectious neurodegenerative disorders. Investigation of prion strains and generation of species dependent TSE model are necessary to understand pathogenesis of the disease. To establish a BSE-specific in vitro cell culture model, N2a and GT1 mouse neuronal cell lines were generated to express the bovine prion protein by transfection of the bovine prion gene (Prnp). In addition, the endogenous mouse prion protein was suppressed in N2a, NbP, GT1, and GbP cell lines using the siRNA duplexes, siRNA1 and siRNA2 that target the N- and C- termini of murine Prnp, respectively. Both siRNA1 and siRNA2 effectively decreased murine prion protein levels by more than 80% and the down-regulation efficacy was increased in siRNA dose-dependent manner. The greatest down-regulation was observed 48 h after siRNA delivery. The moPrnp knockdown NbP and GbP cell lines and the Prnp-targeting siRNA technique established in the present study would be useful tools for dissecting the basic mechanisms of prion infection, especially for BSE.




RESEARCH PAPERS

Open Access Article

Fragment molecular orbital calculations reveal that the E200K mutation markedly alters local structural stability in the human prion protein

Koji Hasegawa, Shirou Mohri and Takashi Yokoyama
Pages 38 - 44
http://dx.doi.org/10.4161/pri.4.1.10890
Abstract | PDF

The E200K mutation of the human prion protein (PrP) is known to cause familial Creutzfeldt–Jakob disease. In order to elucidate the effects of the mutation on the local structural stability of PrP, we performed ab initio fragment molecular orbital calculations for the wild-type human PrP and the E200K variant modeled under neutral and mild acidic conditions. The calculations revealed that this substitution markedly altered the intramolecular interactions in the PrP, suggesting that the local structural instabilities induced by the E200K mutation might cause initial denaturation of the PrP and its subsequent conversion to a pathogenic form. This work presents a new approach for quantitatively elucidating structural instabilities in proteins that cause misfolding diseases.

Open Access Article

Interdependence of amyloid formation in yeast: Implications for polyglutamine disorders and biological functions

Valery N. Urakov, Aleksandra B. Vishnevskaya, Ilya M. Alexandrov, Vitaly V. Kushnirov, Vladimir N. Smirnov and Michael D. Ter-Avanesyan
Pages 45 - 52
http://dx.doi.org/10.4161/pri.4.1.11074
Abstract | PDF

In eukaryotic cells amyloid aggregates may incorporate various functionally unrelated proteins. In mammalian diseases this may cause amyloid toxicity, while in yeast this could contribute to prion phenotypes. Insolubility of amyloids in the presence of strong ionic detergents, such as SDS or sarcosyl, allows discrimination between amorphous and amyloid aggregates. Here, we used this property of amyloids to study the interdependence of their formation in yeast. We observed that SDS-resistant polymers of proteins with extended polyglutamine domains caused the appearance of SDS or sarcosyl-insoluble polymers of three tested chromosomally-encoded Q/N-rich proteins, Sup35, Rnq1 and Pub1. These polymers were non-heritable, since they could not propagate in the absence of polyglutamine polymers. Sup35 prion polymers caused the appearance of non-heritable sarcosyl-resistant polymers of Pub1. Since eukaryotic genomes encode hundreds of proteins with long Q/N-rich regions, polymer interdependence suggests that conversion of a single protein into polymer form may significantly affect cell physiology by causing partial transfer of other Q/N-rich proteins into a non-functional polymer state.