Functional proteome analysis of age associated PrPC knockout mice liver along with regulatory response of cytoskeleton associated tau protein and fatty liver disease.
by Amandeep Singh Arora
Date of Examination:2015-04-14
Date of issue:2015-12-03
Advisor:Prof. Dr. Mathias Bähr
Referee:Prof. Dr. Mikael Simons
Referee:Prof. Dr. Uwe-Karsten Hanisch
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EnglishPrPC is a membrane associated cellular glycoprotein. Its mis-folded form, PrPSc causes a set of neurodegenerative diseases called transmissible spongiform encephalopathies (TSEs). PrPC is abundantly expressed in the brain tissue and a relatively lower expression has been reported in the peripheral tissues. It has contradictorily been described to be involved in multiple functions beside the consensually accepted function as an anti-oxidant. PrPC has evolutionary conserved sequence and it shows that it should have some fundamental functions. The functional studies of PrPC in the peripheral tissues could be useful to elucidate its fundamental function and to better understand its role in the disease mechanism. Among the peripheral tissues, the liver has low PrPC expression, but interestingly its expression increases in oxidative stress during the activation of hepatic stellate cells (HSCs), which have role in liver diseases. In addition, our first set of results have shown a significant up-regulation of PrPC expression in the liver of aging mice (14 months) as compared to the 3 and 9 months old mice with a differential expression in the male and female groups. To understand the age and gender dependent altered expression of PrPC and to find out that if there was any manifestation of liver diseases, 2D gel electrophoresis based proteomics analysis was performed in the liver of PrPC knockout mice (Zürich I). Proteins differentially regulated in the liver of PrPC Knockout mice were identified by the tandem mass spectrometry (MS/MS). Further, the functional analysis of proteomics dataset was performed by using bioinformatics tool called Ingenuity pathway analysis (IPA). IPA predicted the manifestations of the liver diseases including the fatty liver disease along with the regulation of insulin receptor (INSR) and PPARα transcriptional factor. The biochemical measurement of triglycerides and the staining of liver tissue by Sudan III confirmed the fatty liver disease in the liver of aging PrPC knockout mice. Further, the regulation of apoptotic markers by the Western blot indicated the progression of fatty liver disease into non-alcoholic steatohepatitis (NASH). As majority of the research work in the PrPC knockout mouse models has been performed in the brain tissue. Thus, we further performed an additional IPA analysis from the same proteomics dataset by setting brain as a reference tissue. This analysis included the broader PrPC relevant literature information stored in the ingenuity Pathways Knowledge Base (IPKB). It predicted the role of PrPC in cellular development, cell signaling, cellular growth and proliferation including the role of microtubule associated tau protein (MAPT). Further, by using Western blot analysis we observed an age dependent decrease in tau expression and an increase of ptau expression in the liver of old age mice. Furthermore, we showed the PrPC dependent regulation of ptau by gsk3beta, which is an already known mechanism during prion infection. Fatty liver disease has already been described to be associated with the imbalance of liver cytoskeleton by the regulation of tau protein expression. Hence, our study showed that the absence of PrPC causes the fatty liver disease along with associated change in cytoskeleton function in the aging mice liver. In conclusion, from this current study, by using PrPC knockout mice, we showed a critical role of PrPC in the liver of aging mice by regulating the glucose/lipid metabolism, which further seems to be associated with its cytoskeleton function.
Keywords: Prions+PrPC knockout mice+Liver