P4-122: Abnormal accumulation of citrullinated proteins in scrapie-infected mouse brain

Abstract

Prion diseases are neurodegenerative disorders by a pathogenic isoform of the prion protein (PrPSc). Although the key event in the pathology of prion diseases is thought to be the conversion of cellular prion protein (PrPC) to the protease–resistant pathogenic form (PrPSc), the factors that contribute to neurodegeneration in scrapie–infected animals are poorly understood. It can be suggested that there are possibilities of structural differences or differential post–translational modification between PrPC and PrPSc. Citrullination is a post–translational modification process that arginine residue in protein converts to citrulline in a Ca2+–dependent condition by peptidylarginine deiminases (PADs). To elucidate the involvement of protein citrullination in prion diseases, we examined whether citrullinated proteins are produced in scrapie–infected mouse brains. To determine the expression of PAD2 and the detection of citrullinated proteins, we carried out Western blot and immunohistochemistry analyses using the brains of control and scrapie–infected mice. To identify the new substrate for PAD2, we carried out two–dimensional gel electrophoresis and proteome analysis. We have found that PAD2 immunoreactivity and its expression levels of both mRNA and protein were significantly increased in the brains of scrapie–infected mice and that PAD2 was distributed mainly in reactive astrocytes. In addition, we have found that citrullinated proteins of varied molecular weights were detected in both control and scrapie–infected brains and that the level of citrullinated proteins increased in scrapie–infected mouse brain by Western blot analysis with anti–modified citrulline antibody. Interestingly, one of the citrullinated proteins was identified as dynamin using MALDI–TOF mass spectrometry. This study suggests that PAD2 has an important role in the pathogenesis of prion diseases and that the citrullinated dynamin may be involved in PrP trafficking and become a useful marker for prion diseases. This study was supported by grants A020007 from Ministry of Health & Welfare.