^CtmPrP and ER stress

Abstract

The pathogenic agent is hypothesized to be PrP^Sc in prion diseases. However, little accumulation of PrPSc is repeatedly observed in some kinds of natural and experimental prion diseases, including some special genetic human prion diseases. One of the specific topology forms of PrP, ^CtmPrP, representing a key neurotoxic intermediate in prion disorders, has been testified in cell-free translation systems and transgenic mice models. Recently, some studies have showed that point-mutations within the hydrophobic transmembrane region increase the amount of ^CtmPrP in cells, such as human homologue A117V which is associated with GSS and G114V associated with gCJD, while the mutations outsides transmembrane region do not. The retention of the CtmPrP in ER subsequently is able to induce ER stress and apoptosis, which is supported by up-regulation of ER chaperone synthesis, such as Grp78, Grp58, Grp94, Bip and the transcription factor CHOP/GADD153. In conclusion, some kinds of intermediate forms of PrP^Sc, including ^CtmPrP, may work as the ultimate cause of neurodegeneration.

Introduction

Prion diseases or transmissible spongiform encephalopathies (TSE) are a group of neurodegenerative diseases, including Creutzfeldt-Jacob disease (CJD), Gerstmann-Sträussler-Scheinker syndrome (GSS), fatal familial insomnia (FFI) and Kuru in humans, scrapie in sheep and bovine spongiform encephalopathy (BSE) in cattle.1 The pathogenic agent is hypothesized to be PrP^Sc, an abnormal isoform that is infectious in the absence of nucleic acid and is converted from PrP^C, a normal cell-surface glycoprotein. The secondary structural difference between two PrP isoforms seems to be clear that PrP^Sc contains significantly more β-sheet structure.2 Although deposits of PrP^Sc in brains are common pathological features of prion diseases, significant pathology and clinical dysfunction with little accumulation of PrP^Sc are repeatedly observed in some kinds of natural and experimental prion diseases.3 Hence, the infectious form of PrP may not be the proximate cause of neuronal dysfunction and degeneration. Several alternative forms of PrP have therefore been hypothesized to be the primary neurotoxic species designated PrP^toxic, such as ^CtmPrP.4

Approximately 10–15% of human prion cases are related with the mutations of prion protein gene (PRNP) on chromosome 20, which form a special subtype of human prion diseases or CJD, including genetic or familial CJD (gCJD or fCJD), Gerstmann-Sträussler-Scheinker syndrome (GSS) and fatal familial insomnia (FFI).5 More than 55 mutations in PRNP have been confirmed to be associated with or directly linked to the development of genetic human prion diseases.6 The mutations consist of a series of insertions or deletion of octarepeats in the N-terminus of PrP and numerous point-mutations in the middle and C-terminal sequences of PrP. Interestingly, the pathological characteristics and clinical phenotypes vary largely among various gCJDs, which strongly highlights that different mutated PrP forms may have distinct pathologic pathways.

^CtmPrP, One of the Membrane Topology Forms of PrP

Most mature PrP molecules present on the surface of cells through their GPI anchors that serve as their sole means of attachment to the lipid bilayer after post-translational modification processes in endoplasmic reticulum (ER) lumen and Golgi apparatus.7 Additionally, PrP contains a conserved hydrophobic sequence that can span the lipid bilayer in either direction, resulting in two transmembrane forms, designated ^CtmPrP and ^NtmPrP. ^CtmPrP is believed to span the membrane once, with its highly conserved hydrophobic region in the center of the molecule (aa 111–134) as a transmembrane anchor and the C terminus in the ER lumen. ^NtmPrP spans the membrane with the same transmembrane segment, but with its N terminus in the ER lumen reversely.8

^CtmPrP, a Neurotoxic Element in Some Prion Neurodegeneration

Presence of several topological forms of PrP, which is firstly noticed in cell-free translation/translocation systems, has been observed for long time.9,10 In the past years, expressions of ^CtmPrP in experimental rodents have been confirmed to be associated with neurodegenerative diseases. In the transgenic mice expressing PrP molecules carrying mutations within or near the transmembrane domain, obvious ^CtmPrP can be identified in brain membranes.11 More interestingly, transgenic mice expressing such ^CtmPrP-favoring mutations are more likely to develop a spontaneous neurodegenerative illness that bears some similarities to scrapie and some kinds of human genetic prion diseases, without detectable PrP^Sc in brains.4,8 Unlike the natural PrP^Sc, ^CtmPrP inducing neurodegenerative diseases show extremely low infectivity.12 Although the exact pathogenic role of ^CtmPrP in prion diseases still remains unclear, numerous data have illustrated that this special molecule possesses significant neurotoxicity. Moreover, Hegde et al. have reported that accumulation of PrP^Sc induces an enhanced generation of ^CtmPrP, implying that ^CtmPrP represents a key neurotoxic intermediate in prion disorders.4

^CtmPrP, a Trigger for ER Stress and Apoptosis

Many cell biological assays highlight that ^CtmPrP contains an uncleaved signal peptide as well as a GPI anchor, which retains intracellularly in either the ER or the Golgi depending on the cell type.13 An ER stress and subsequent apoptosis has been repeatedly observed in the cells of retention of the abnormal ^CtmPrP in ER, which is believed to be the normal physiological reaction for cleaning out the misfolding proteins.14 Supporting this claim, many protective molecular chaperons, such as GRP78, GRP94, GRP29, ORP150, PDI, SERCA, HO-1, SERP1 and Herp, are obviously upregulated due to the ER stress.15 Normally, the unfolded protein response (UPR) could results in upregulation of ER chaperone synthesis, such as Grp78, Grp58, Grp94 and Bip, which are adaptive in nature, but the induction of the transcription factor CHOP/GADD153 and phosphorylation of the translation initiation factor eIF-2 can damage cells by triggering apoptosis.16,17 More recently, Wang et al. have proposed the evidences that enhanced levels of ER stress up-stream proteins Grp78, Grp58, PERK and Bip are specifically observed in the cells, after formation of ^CtmPrP by expressions of either the genetic engineering recombinant PrPs retained in ER (PrP-KDEL and PrP-3AV) or the genetic prion disease associated PrP mutants within the transmembrane region (PrP-G114V and PrP-A117V).18 Subsequently, the presences of the ER stress chaperones trigger the increase of the following apoptosis chaperones such as CHOP and pro-caspase-12.

^CtmPrP, a Cytolic PrP Special in Some PrP Mutants

Up to now, dozens of PrP mutants are definitely confirmed to be related with the prion diseases in humans, either familiarly or spontaneously. Additionally, numerous artificially modified PrP molecules possess the abilities to cause neurodegenerative disorders in transgenic mice. Isaac et al. have classified the PrP mutants into three categories,19 in which the second category of mutations are those in the N-terminal signal sequence and hydrophobic domain that influence the membrane topology of PrP. Wang et al. have illustrated that disease-related point-mutations within the hydrophobic transmembrane region increase the amount of ^CtmPrP in cells, such as human homologue A117V which is associated with GSS20 and G114V associated with gCJD.21 Meanwhile, the formation of ^CtmPrP in cells seems to be special to the mutations in this region, since expressions of other fCJD-related PrP mutants outside this region (P102L and E200K) in cells do not produce detectable ^CtmPrP, though those two mutants cause clearly cytotoxicity.18

Besides the transmembrane region, nonconservative substitutions in the core of the N-terminal signal sequence are also able to increase the proportion of ^CtmPrP, combining these mutations with ones in the central domain results in almost exclusive synthesis of the ^CtmPrP.22 The inserted mutation of fourteen octarepeats (PrP-PG14) seems to be localized in ER, which induce TSE-like disease after introducing into transgenic mice.23 More recently, Xu et al. propose the data that expressions of the insertions with nine-(PrP-PG9) and twelve-(PrP-PG12) octarepeats partially retain in ER, sensitize the transfected cells to ER stress stimuli and trigger the apoptosis via ER stress, whereas PrP mutant with octarepeats deletion (PrP-PG0) induce apoptosis probably through mitochondrial disruption pathway.24 It lays out a broad diversity of PrP mutations in damaging cells and in resulting in various clinical phenotypes of genetic human TSEs.

Speculation

PrP^Sc is found in the brains in most cases of infectious, familial and sporadic prion disease. Although close correlation between the accumulation of PrP^Sc and the appearance of neuropathological changes have been addressed in most human and animal TSEs, there are still a number of exceptions. Either in some transmission bioassays25 or in some human genetic prion diseases,26 remarkable pathological changes and clinical manifestations are accompanied little deposit of PrP^Sc in central nerve system. It raises the question whether PrP^Sc is really the proximate cause for neuron damage in all prion diseases. Partially in line with the phenomenon, the famous peptide PrP106-126 shows rapid toxicity, but almost loses its cytotoxicity after aggregation on the cultured cells,27 possibly indicating a similar manner that the aggregated PrP^Sc might be nontoxic. Instead of that, some kinds of intermediates, including ^CtmPrP, may work as the ultimate cause of neurodegeneration.

Acknowledgments

This work was supported by Chinese National Natural Science Foundation Grants 30800975, National Basic Research Program of China (973 Program) (2007CB310505), China Mega-Project for Infectious Disease (2009ZX10004-101 and 2008ZX10004-008) and the SKLID Development Grant (2008SKLID102, 2011SKLID204 and 2011SKLID211).