Peng Xiangmin, Tehranian Roya, Dietrich Paula, Stefanis Leonidas, Perez Ruth G
Department of Neurology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213, USA.
J Cell Sci. 2005 Aug 1;118(Pt 15):3523-30. doi: 10.1242/jcs.02481. Epub 2005 Jul 19.
alpha-Synuclein is an abundant presynaptic protein implicated in neuronal plasticity and neurodegenerative diseases. Although the function of alpha-synuclein is not thoroughly elucidated, we found that alpha-synuclein regulates dopamine synthesis by binding to and inhibiting tyrosine hydroxylase, the rate limiting enzyme in dopamine synthesis. Understanding alpha-synuclein function in dopaminergic cells should add to our knowledge of this key protein, which is implicated in Parkinson's disease and other disorders. Herein, we report a mechanism by which alpha-synuclein diminishes tyrosine hydroxylase phosphorylation and activity in stably transfected dopaminergic cells. Short-term regulation of tyrosine hydroxylase depends on the phosphorylation of key seryl residues in the amino-terminal regulatory domain of the protein. Of these, Ser40 contributes significantly to tyrosine hydroxylase activation and dopamine synthesis. We observed that alpha-synuclein overexpression caused reduced Ser40 phosphorylation in MN9D cells and inducible PC12 cells. Ser40 is phosphorylated chiefly by the cyclic AMP-dependent protein kinase PKA and dephosphorylated almost exclusively by the protein phosphatase, PP2A. Therefore, we measured the impact of alpha-synuclein overexpression on levels and activity of PKA and PP2A in our cells. PKA was unaffected by alpha-synuclein. PP2A protein levels also were unchanged, however, the activity of PP2A increased in parallel with alpha-synuclein expression. Inhibition of PP2A dramatically increased Ser40 phosphorylation only in alpha-synuclein overexpressors in which alpha-synuclein was also found to co-immunoprecipitate with PP2A. Together the data reveal a functional interaction between alpha-synuclein and PP2A that leads to PP2A activation and underscores a key role for alpha-synuclein in protein phosphorylation.
α-突触核蛋白是一种丰富的突触前蛋白，与神经元可塑性和神经退行性疾病有关。尽管α-突触核蛋白的功能尚未完全阐明，但我们发现α-突触核蛋白通过与多巴胺合成的限速酶酪氨酸羟化酶结合并抑制它来调节多巴胺合成。了解α-突触核蛋白在多巴胺能细胞中的功能，应有助于我们对这种与帕金森病和其他疾病有关的关键蛋白的认识。在此，我们报告了一种机制，通过该机制α-突触核蛋白可降低稳定转染的多巴胺能细胞中酪氨酸羟化酶的磷酸化和活性。酪氨酸羟化酶的短期调节取决于该蛋白氨基末端调节域中关键丝氨酸残基的磷酸化。其中，Ser40对酪氨酸羟化酶的激活和多巴胺合成有显著贡献。我们观察到，α-突触核蛋白过表达导致MN9D细胞和诱导型PC12细胞中Ser40磷酸化减少。Ser40主要由环磷酸腺苷依赖性蛋白激酶PKA磷酸化，几乎完全由蛋白磷酸酶PP2A去磷酸化。因此，我们测量了α-突触核蛋白过表达对我们细胞中PKA和PP2A水平及活性的影响。PKA不受α-突触核蛋白的影响。PP2A蛋白水平也未改变，然而，PP2A的活性与α-突触核蛋白的表达平行增加。仅在α-突触核蛋白过表达细胞中抑制PP2A可显著增加Ser40磷酸化，在这些细胞中还发现α-突触核蛋白与PP2A共免疫沉淀。这些数据共同揭示了α-突触核蛋白与PP2A之间的功能相互作用，这种相互作用导致PP2A激活，并强调了α-突触核蛋白在蛋白质磷酸化中的关键作用。
