GTPase activity regulates kinase activity and cellular phenotypes of Parkinson's disease-associated LRRK2
GTPase activity regulates kinase activity and cellular phenotypes of Parkinson's disease-associated LRRK2
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DOI：
10.1093/hmg/dds522
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发表时间：
2013-03-15
影响因子：
3.5
通讯作者：
Moore, Darren J.
中科院分区：
文献类型：
--
作者：
Biosa, Alice;Trancikova, Alzbeta;Moore, Darren J.
LRRK2基因突变导致常染色体显性帕金森病。LRRK2编码一个多结构域蛋白，包含一个Ras-of-complex (Roc) GTPase结构域、一个Roc结构域的c端和一个蛋白激酶结构域。LRRK2可以作为GTPase和蛋白激酶，尽管这两个酶结构域之间的相互作用尚不清楚。尽管鸟嘌呤核苷酸结合对LRRK2的激酶活性至关重要，但GTP水解的作用尚不清楚。总的来说，调控GTPase活性的分子决定因素以及GTPase结构域如何影响LRRK2的特性仍有待阐明。在这里，我们在GTPase结构域中发现了一些合成错义突变，这些突变在功能上调节GTP结合和GTP水解，我们利用这些突变体全面探索GTPase活性对LRRK2激酶活性和细胞表型的贡献。我们的数据表明，鸟嘌呤核苷酸结合和GTP水解（在较小程度上）是维持正常激酶活性所必需的，这两种活性都有助于GTP依赖性的LRRK2激酶活性激活。鸟嘌呤核苷酸结合而非GTP水解调节LRRK2的二聚化、结构和稳定性。此外，GTP水解调节初级皮质神经元中lrrk2依赖性的神经突生长抑制，但无法强有力地调节家族性G2019S突变的影响。我们的研究阐明了GTPase活性在调节LRRK2的激酶活性和细胞表型中的作用，并对GTPase结构域作为减轻LRRK2介导的神经退行性变的分子靶点的验证具有重要意义。
Mutations in the LRRK2 gene cause autosomal dominant Parkinson's disease. LRRK2 encodes a multidomain protein containing a Ras-of-complex (Roc) GTPase domain, a C-terminal of Roc domain and a protein kinase domain. LRRK2 can function as a GTPase and protein kinase, although the interplay between these two enzymatic domains is poorly understood. Although guanine nucleotide binding is critically required for the kinase activity of LRRK2, the contribution of GTP hydrolysis is not known. In general, the molecular determinants regulating GTPase activity and how the GTPase domain contributes to the properties of LRRK2 remain to be clarified. Here, we identify a number of synthetic missense mutations in the GTPase domain that functionally modulate GTP binding and GTP hydrolysis and we employ these mutants to comprehensively explore the contribution of GTPase activity to the kinase activity and cellular phenotypes of LRRK2. Our data demonstrate that guanine nucleotide binding and, to a lesser extent, GTP hydrolysis are required for maintaining normal kinase activity and both activities contribute to the GTP-dependent activation of LRRK2 kinase activity. Guanine nucleotide binding but not GTP hydrolysis regulates the dimerization, structure and stability of LRRK2. Furthermore, GTP hydrolysis regulates the LRRK2-dependent inhibition of neurite outgrowth in primary cortical neurons but is unable to robustly modulate the effects of the familial G2019S mutation. Our study elucidates the role of GTPase activity in regulating kinase activity and cellular phenotypes of LRRK2 and has important implications for the validation of the GTPase domain as a molecular target for attenuating LRRK2-mediated neurodegeneration.