Dominant-negative effects of LRRK2 heterodimers: A possible mechanism of neurodegeneration in Parkinson's disease caused by LRRK2 I2020T mutation
Dominant-negative effects of LRRK2 heterodimers: A possible mechanism of neurodegeneration in Parkinson's disease caused by LRRK2 I2020T mutation
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LRRK2 异二聚体的显性负效应：LRRK2 I2020T 突变引起帕金森病神经退行性变的可能机制
DOI：
10.1016/j.bbrc.2012.11.113
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发表时间：
2013
期刊：
影响因子：
--
通讯作者：
Obata F
中科院分区：
文献类型：
--
作者：
Ohta E;Kawakami F;Kubo M;Obata F
富含亮氨酸重复序列激酶2（LRRK 2）是常染色体显性帕金森病（PD）PARK 8的致病分子，但其突变的病因学影响仍不清楚。在本研究中，我们研究了I2020 T突变体LRRK 2诱导的神经退行性变的新机制。使用天然凝胶电泳和免疫沉淀，我们发现，野生型（WT）LRRK 2形成异源二聚体与I2020 T LRRK 2在转染细胞，异源二聚体表现出显着较低的细胞内蛋白质水平比WT/WT-同源二聚体。I2020 T LRRK 2的量增加降低了共转染的WT LRRK 2的蛋白水平。脉冲追踪实验显示，通过与I2020 T LRRK 2共转染，WT LRRK 2的细胞内蛋白寿命缩短。这些结果表明，I2020 T LRRK 2通过WT/I2020 T-异源二聚体形成增强WT LRRK 2的细胞内降解。WT LRRK 2在HEK 293细胞中的过表达增加了Akt 1（S473）的磷酸化水平，Akt 1（S473）是LRRK 2的可能的生理底物，并且使细胞对过氧化氢诱导的凋亡具有抗性。然而，与WT/WT表达细胞相比，WT/I2020 T表达细胞中的Akt 1磷酸化和凋亡抗性均降低。当用I2020 T LRRK 2转染过表达WT LRRK 2的神经母细胞瘤SH-SY 5 Y克隆时，Akt 1磷酸化的减少和细胞凋亡抵抗也很明显。总而言之，这些结果表明I2020 T突变通过WT/I2020 T异二聚体形成增强了LRRK 2的细胞内降解，导致Akt 1磷酸化减少并降低了对细胞凋亡的保护作用。我们的研究结果表明，在PD的神经退行性变的可能性，由I2020 T LRRK 2突变引起的显性负性机制。
Leucine-rich repeat kinase 2 (LRRK2) is the molecule responsible for autosomal-dominant Parkinson’s disease (PD), PARK8, but the etiologic effects of its mutation remain unknown. In the present study, we investigated a novel mechanism for the neurodegeneration induced by I2020T mutant LRRK2. Using native gel electrophoresis and immunoprecipitation, we found that wild-type (WT) LRRK2 formed a heterodimer with I2020T LRRK2 in transfected cells, and that the heterodimer exhibited a markedly lower intracellular protein level than the WT/WT-homodimer. An increased amount of I2020T LRRK2 decreased the protein level of co-transfected WT LRRK2. A pulse-chase experiment revealed that the intracellular protein lifetime of WT LRRK2 was shortened by co-transfection with I2020T LRRK2. These results suggest that I2020T LRRK2 enhances the intracellular degradation of WT LRRK2 through WT/I2020T-heterodimer formation. Overexpression of WT LRRK2 in HEK293 cells increased the phosphorylation level of Akt1 (S473), a possible physiological substrate of LRRK2, and made cells resistant to hydrogen peroxide-induced apoptosis. However, both Akt1 phosphorylation and apoptosis resistance were reduced in WT/I2020T-expressing cells in comparison with WT/WT-expressing cells. Reduction of Akt1 phosphorylation and apoptosis resistance were also evident when a neuroblastoma SH-SY5Y clone overexpressing WT LRRK2 was transfected with the I2020T LRRK2. Altogether, these results suggest that the I2020T mutation enhances the intracellular degradation of LRRK2 through WT/I2020T-heterodimer formation, leading to reduced Akt1 phosphorylation and diminished protectivity against apoptosis. Our findings suggest the possibility of a dominant-negative mechanism of neurodegeneration in PD caused by I2020T LRRK2 mutation.