Regulating Factors of PrPres Glycosylation in Creutzfeldt-Jakob Disease - Implications for the Dissemination and the Diagnosis of Human Prion Strains
Etienne Levavasseur1, Isabelle Laffont-Proust1, Émilie Morain1, Baptiste A. Faucheux1,2, Nicolas Privat1, Katell Peoc'h4, Véronique Sazdovitch1,2, Jean-Philippe Brandel1, Jean-Jacques Hauw2,3, Stéphane Haïk1,2,3*
1 INSERM, Avenir Team - Human Prion Diseases, Paris, France2 APHP, R. Escourolle Neuropathology Laboratory, Paris, France3 InVS, French National Center of Reference for Unconventional Transmissible Agents, Paris, France4 Biochemistry and Molecular Biology Department, Lariboisière Hospital, Paris, France
Abstract Objective The glycoprofile of pathological prion protein (PrPres) is widely used as a diagnosis marker in Creutzfeldt-Jakob disease (CJD) and is thought to vary in a strain-specific manner. However, that the same glycoprofile of PrPres always accumulates in the whole brain of one individual has been questioned. We aimed to determine whether and how PrPres glycosylation is regulated in the brain of patients with sporadic and variant Creutzfeldt-Jakob disease.
Methods PrPres glycoprofiles in four brain regions from 134 patients with sporadic or variant CJD were analyzed as a function of the genotype at codon 129 of PRNP and the Western blot type of PrPres.
Results The regional distribution of PrPres glycoforms within one individual was heterogeneous in sporadic but not in variant CJD. PrPres glycoforms ratio significantly correlated with the genotype at codon 129 of the prion protein gene and the Western blot type of PrPres in a region-specific manner. In some cases of sCJD, the glycoprofile of thalamic PrPres was undistinguishable from that observed in variant CJD.
Interpretation Regulations leading to variations of PrPres pattern between brain regions in sCJD patients, involving host genotype and Western blot type of PrPres may contribute to the specific brain targeting of prion strains and have direct implications for the diagnosis of the different forms of CJD.
Discussion This study provides new insight into the analysis of PrPres heterogeneity within sCJD cases, based upon the biochemical analysis of at least 3 different brain regions from 123 sCJD cases. This large series of sCJD tissues allowed us to identify the diversity of PrPres glycoforms within sporadic CJD forms and to establish a correlation with PRNP genotype, brain region and Western blot type of PrPres. Our results demonstrate that the regional distribution of PrPres glycoforms within one individual is heterogeneous in the sporadic form of CJD and suggest the involvement of precise regulating mechanisms.
Different factors may contribute to this regulation. In the normal brain of human, hamster and mouse, PrPc glycosylation pattern is submitted to a regional diversity , , . In infected human brains, the diversity of PrPres glycoprofiles may result from such a regional variation of the substrate for prion replication. According to the prion hypothesis, this mechanism suggests that the glycoprofile of PrPc influences the glycoform ratio of PrPres in the conversion process. In acellular models of PrP conversion indeed, the use of PrPc with modified glycoforms ratio partially modifies the glycosylation pattern of newly formed PrPres . In the present study, we observed that the genotype at codon 129 of PRNP influenced the glycoform ratio of PrPres in a region dependent manner. This raises the possibility that the regional glycosylation state of PrPc may be affected by PRNP polymorphisms. It may be of interest to document such a regulation in the normal human brain.
Several studies have pointed out a relation between PrP amino acid sequence (i.e. PRNP genotype) and PrPres glycosylation in inherited diseases associated to mutation located close to (D178N, V180I, T183A, E200K) or remote (P102L) from the N-glycosylation sites , . Whether the presence of a mutation may modify the glycoform ratio of PrPc in the human brain is poorly documented. However we previously showed that in patients with the V180I mutation, no diglycosylated PrPres is produced, while PrPc glycosylation is normally processed . This clearly indicates that additional factors, different from the glycosylation state of PrPc, may influence the glycoform ratio of the accumulated PrPres in the pathological condition.
Type 1 or type 2 PrPres are thought to correspond to distinct accessibilities of proteinase K to cleavage sites resulting from different conformations of PrPres. A diversity of glycotypes has been described for type 2 PrPres, linked to the various forms of the diseases (sporadic, variant CJD, familial fatal insomnia). In sporadic CJD patients with the same genotype at codon 129, we found that PrPres type markedly influences the glycoform ratio of the accumulated PrPres in definite brain regions. This suggests that some conformers are more prone to convert distinct glycoforms of PrPc in sporadic diseases and that this phenomenon is regionally regulated. In addition, to explain the strain mutation phenomenon, it has been recently proposed that a single strain may consist of an ensemble of molecular species containing a dominant PrPres type . Such conformers may differ by the glycoform species they favor during replication. It can be speculated that regional factors influence the selection of PrPres subpopulations leading to the variation of the glycoform ratio that we finally detected.
The regional variation of PrPres glycoform ratio demonstrated in this study may be of interest in the understanding of “lesion profile” and brain targeting by different prion strains. The mechanisms that favor the accumulation of a given strain in a given brain region are not known, but the regulation of PrPc/ PrPres glycosylation is likely involved in this process , , . As a matter of fact, we found in sCJD cases that the thalamus favored the production of type 2 PrPres presenting a high content in diglycosylated forms. It is noteworthy that besides sCJD, vCJD and FFI are associated with a high content of diglycosylated forms and are characterized by a “targeting” of the thalamus. In all studied brain areas from French vCJD patients, as already reported in the U.K. cases, we observed that the glycoform ratio was remarkably constant. In this infectious form of prion disorders, the pathogenic events are initiated by an exogenous PrPres from contaminated bovine tissues with its own defined characteristics, such as a high glysosylated content that is maintained through interspecies transmission. By contrast, the causative event in sCJD is thought to be a rare, spontaneous and endogenous conversion of the host-encoded PrPc into PrPres. The diversity of glycoform ratios that we observed in sCJD may reflect the regulation of an endogenous phenomenon by various host factors (genotype, brain region) that may be overwhelmed in the case of an infection by a virulent prion strain such as the bovine agent in vCJD patients.
Our observation of PrPres glycoprofile diversity in sCJD raises the question of the strain diagnosis based on biochemical PrPres analysis. The type 2B PrPres profile is widely used as a diagnosis marker of definite vCJD that until now has been observed in MM patients only. However, in some VV2 CJD patients, the high proportion of diglycosylated PrPres we observed in the thalamus was undistinguishable from the PrPres profile detected in vCJD cases. Studies using transgenic mice expressing human PrP suggest that clinicopathological and biochemical presentations of BSE infection may vary in humans with the genotype at codon 129 . One possible explanation of our findings in these particular VV2 cases could be BSE infection (i.e. vCJD) in valine homozygote patients. However, the mean age at death (>65 years, except one case who was 27) and the neuropathological pattern (absence of florid plaques) do not support such hypothesis. While the first MM vCJD patients occurred simultaneously in UK and France, the peak of French vCJD epidemics occurred in 2002 more than 5 years later than in the UK. Comparison of the 1980–1995 pattern of BSE exposition in the UK and France indicated that it peaked also later in France . This probably explains the different temporal pattern of vCJD incidence. We analyzed retrospectively whether some French VV2 CJD patients who died between 1993 and 1999 exhibited also a thalamic PrPres profile with high glycosylation site occupancy similar to what is observed in vCJD. The same profile of PrPres ressembling type 2B could also be detected in patients who died during the 1993–1999 period (Fig 6). This suggests that these VV patients had indeed a genuine sporadic disease rather than an atypical vCJD, which is very unlikely to have occurred in an unrelated genotype and before 1996 in France.
Finally, the present work evidences regulations that lead to highly significant variations of PrPres pattern between brain regions in sCJD patients, involving host genotype and Western blot type of PrPres as recapitulated in figure 7. These may contribute to the specific brain targeting of prion strains and have some diagnosis implications.
Figure 6. Similar PrPres glycoform ratios with high glycosylation site occupancy in thalamus from VV2 sCJD patients and vCJD patients.
Each dot represents one brain area from a patient and is defined on the ternary plot by three coordinates corresponding to the relative intensity of di-, mono- and nonglycosylated bands. We compared the VV2 patients from our sCJD series and vCJD patients with VV2 sCJD patients who died before or just after the first variant breakout in 1996. No difference was observed between VV2 patients from the 2000–2007 series and VV2 patients from the period 1993–1999.
Figure 7. Schematic representation of regulation of PrPres glycoforms by regional and molecular factors in sporadic Creutzfeldt-Jakob disease.
Arrows indicate which glycoform of PrPres is favored by individual factors. As an example, the accumulation of diglycosylated PrPres is favored in the thalamus, in VV patients and in case of type 2 PrPres.
see full text ;
MAD COW DISEASE terminology UK c-BSE (typical), atypical BSE H or L, and or Italian L-BASEhttp://bse-atypical.blogspot.com/2008/03/mad-cow-disease-terminology-uk-c-bse.htmlSaturday, June 21, 2008 HUMAN and ANIMAL TSE Classifications i.e. mad cow disease and the UKBSEnvCJD only theory JUNE 2008
Manuscript Draft Manuscript Number:
Title: HUMAN and ANIMAL TSE Classifications i.e. mad cow disease and the UKBSEnvCJD only theory
Article Type: Personal View
Corresponding Author: Mr. Terry S. Singeltary,
Corresponding Author's Institution: na
First Author: Terry S Singeltary, none
Order of Authors: Terry S Singeltary, none;
Terry S. Singeltary
TSEs have been rampant in the USA for decades in many species, and they all have been rendered and fed back to animals for human/animal consumption. I propose that the current diagnostic criteria for human TSEs only enhances and helps the spreading of human TSE from the continued belief of the UKBSEnvCJD only theory in 2007. HUMAN and ANIMAL TSE Classifications i.e. mad cow disease and the UKBSEnvCJD only theory August 2007 August 2007 HUMAN and ANIMAL TSE Classifications i.e. mad cow disease and the UKBSEnvCJD only theory TSEs have been rampant in the USA for decades in many species, and they all have been rendered and fed back to animals for human/animal consumption. I propose that the current diagnostic criteria for human TSEs only enhances and helps the spreading of human TSE from the continued belief of the UKBSEnvCJD only theory in 2007. With all the science to date refuting it, to continue to validate this myth, will only spread this TSE agent through a multitude of potential routes and sources i.e. consumption, surgical, blood, medical, cosmetics etc. I propose as with Aguzzi, Asante, Collinge, Caughey, Deslys, Dormont, Gibbs, Ironside, Manuelidis, Marsh, et al and many more, that the world of TSE Transmissible Spongiform Encephalopathy is far from an exact science, but there is enough proven science to date that this myth should be put to rest once and for all, and that we move forward with a new classification for human and animal TSE that would properly identify the infected species, the source species, and then the route. This would further have to be broken down to strain of species and then the route of transmission would further have to be broken down. Accumulation and Transmission are key to the threshold from sub-clinical to clinical disease, and key to all this, is to stop the amplification and transmission of this agent, the spreading of, no matter what strain. In my opinion, to continue with this myth that the U.K. strain of BSE (one strain TSE in cows), and the nv/v CJD (one strain TSE humans) and that all the rest of human TSE are just one single strain i.e. sporadic CJD (when to date there are 6 different phenotypes of sCJD, and growing per Gambetti et al), and that no other animal TSE transmits to humans, to continue with this masquerade will only continue to spread, expose, and kill, who knows how many more in the years and decades to come. ONE was enough for me, My Mom, hvCJD i.e. Heidenhain Variant CJD, DOD 12/14/97 confirmed, which is nothing more than another mans name added to CJD, like CJD itself, Jakob and Creutzfeldt, or Gerstmann-Straussler-Scheinker syndrome, just another CJD or human TSE, named after another human. WE are only kidding ourselves with the current diagnostic criteria for human and animal TSE, especially differentiating between the nvCJD vs the sporadic CJD strains and then the GSS strains and also the FFI fatal familial insomnia strains or the ones that mimics one or the other of those TSE? Tissue infectivity and strain typing of the many variants Manuscript of the human and animal TSEs are paramount in all variants of all TSE. There must be a proper classification that will differentiate between all these human TSE in order to do this. With the CDI and other more sensitive testing coming about, I only hope that my proposal will some day be taken seriously. ...
Terry S. Singeltary Sr. P.O. Box 42 Bacliff, Texas USA 77518 firstname.lastname@example.org
Tuesday, July 29, 2008
Heidenhain Variant Creutzfeldt Jakob Disease Case Report
10 people killed by new CJD-like disease
Public release date: 9-Jul-2008
Since Gambetti's team wrote a paper describing an initial 11 cases referred to his centre between 2002 and 2006 (Annals of Neurology, vol 63, p 697), another five have come to light. "So it is possible that it could be just the tip of the iceberg," Gambetti says.
sporadic CJD, the big lie
Thursday, July 10, 2008 A Novel Human Disease with Abnormal Prion Protein Sensitive to Protease update July 10, 2008
Thursday, July 10, 2008 A New Prionopathy update July 10, 2008