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Prion protein-induced proteome alterations in sporadic Creutzfeldt-Jakob disease and in SH-SY5Y cell culture model

dc.contributor.advisorBähr, Mathias Prof.
dc.contributor.authorGawinecka, Joannade
dc.titlePrion protein-induced proteome alterations in sporadic Creutzfeldt-Jakob disease and in SH-SY5Y cell culture modelde
dc.contributor.refereeZerr, Inga Prof.
dc.subject.dnb500 Naturwissenschaftende
dc.description.abstractengCellular prion protein (PrPC) is a glycosylated membrane glycoprotein mainly expressed in the central nervous system. Some still undefined molecular events can lead to the conversion of PrPC into an abnormal conformer called scrapie prion protein (PrPSc). PrPSc is characterized by increased â-sheet content, detergent insolubility, partial resistance to protease digestion and tendency to aggregate in the brain tissue. The conversion and subsequent aggregation of PrPSc in the brain tissue underlay pathogenesis of transmissible spongiform encephalopathies. In turn, sporadic Creutzfeldt-Jakob disease is most common form of human transmissible spongiform encephalopathie. The heterogeneous disease phenotype is mainly influenced by the methionine/valine (M/V) polymorphism at codon 129 in the human prion protein gene (PRNP gene) and by the presence of two major types of pathological, protease-resistant forms of the prion protein (PrPSc) leading to 2 different profiles in Western blot (type 1 and type 2). This thesis includes 2 original publications that investigate the physiology and the pathology of the human prion protein. In our first study, CSF proteome alterations in living, symptomatic sCJD patients with two different codon 129 genotypes (MM and VV) were analyzed using a proteomic approach consisted of 2D Fluorescence Difference Gel Electrophoresis (2D-DIGE) and mass spectrometry analysis. CSF proteome profiling revealed up-regulation of 27 and down-regulation of 3 proteins in the MM-sCJD as well as the up-regulation of 24 proteins in the VV-sCJD when compared to control. Beside proteins showing common regulation for both codon 129 genotypes in sCJD, some proteins seem to be specifically regulated in certain genotype. Almost 40% of sCJD specifically regulated proteins in CSF are involved in glucose metabolism, regardless of codon 129 polymorphism. The validation phase of selected glycolytic enzymes using Western blot technique confirmed up-regulation of ALDOA, LDH and G6PI when compared to three different control groups (NDC, AD and VD). Furthermore, the immunolabeling showed that G6PI is present in reactive astrocytes in sCJD affected brain while it is predominantly localized in neurons in age-matched control brain. Additionally, decreased level of G6PI was found in the brain of MM1-sCJD subtype. With these data, for the first time the implication of G6PI in prion-induced pathology was demonstrated. The identification of sCJD-regulated proteins in CSF proteome alterations in living, symptomatic sCJD- patients will broaden our knowledge about pathological processes occurring in sCJD, as they are still not fully understood. Moreover, they could serve as protein source to identify novel biomarkers for differential sCJD diagnosis. In our second study, a proteomic approach was applied in order to reveal proteins, and thereby biological processes, affected by stable overexpression of human PrPC in human neuroblastoma SH-SY5Y cell line. Densitometric analysis of silver stained 2D gels showed 18 differentially regulated proteins in SH-SY5Y cells overexpressing human PrPC. Between them, 13 proteins were up- and 5 down-regulated. The PrPC overexpression in SH-SY5Y cells affected mostly few biological processes such as signal transduction, cytoskeleton organization and protein folding. Proteomics gives a unique opportunity to analyze both physiological and pathological processes at the protein level on a global scale. Proteome analysis of SH-SY5Y human neuroblastoma cells stably overexpressing PrPC revealed proteins whose expression is directly modulated through PrPC and consequently physiological processes which are influenced by PrPC level in cells. On the other hand side, identification of CSF proteome alterations in sCJD provides more information about pathological processes occurring in the brain and caused by presence of PrPSc. Both these studies broaden our knowledge about still not fully understood pathobiology of
dc.contributor.coRefereeBayer, Thomas A. Prof.
dc.contributor.thirdRefereeSimons, Mikael Prof.
dc.subject.topicBiology (incl. Psychology)de
dc.subject.gerCreutzfeldt-Jakob Krankheitde
dc.subject.engCreutzfeldt-Jakob diseasede
dc.affiliation.instituteBiologische Fakultät inkl. Psychologiede

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