Specificity and regulation of casein kinase-mediated phosphorylation of α-synuclein

Abstract

α-Synuclein (α-syn) is the major component of pathologic inclusions that characterize neurodegenerative disorders such as Parkinson disease, dementia with Lewy body disease, and multiple system atrophy. The present study uses novel phospho-specific antibodies to assess the presence and regulation of phosphorylated Ser87 and Ser129 in α-syn in human brain samples and in a transgenic mouse model of α-synucleinopathies. By immunohistochemistry, α-syn phosphorylated at Ser129, but not at Ser87, was abundant in α-syn inclusions. Under normal conditions, Ser129 phosphorylation, but not Ser87 phosphorylation, was detected at low levels in the soluble biochemical fractions in human α-syn transgenic mice and stably transfected cultured cells. Therefore, a role for Ser87 phosphorylation in α-synucleinopathies is unlikely, and in vitro assays showed that phosphorylation at this site would inhibit polymerization. In vitro studies also indicated that hyperphosphorylation of Ser129 α-syn in pathologic inclusions may be due in part to the intrinsic properties of aggregated α-syn to act as substrates for kinases but not phosphatases. Further studies in transgenic mice and cultured cells suggest that cellular toxicity, including proteasomal dysfunction, increases casein kinase 2 activity, which results in elevated Ser129 α-syn phosphorylation. These data provide novel explanations for the presence of hyperphosphorylated Ser129 α-syn in pathologic inclusions. © 2008 American Association of Neuropathologists, Inc.