Verma Manish, Callio Jason, Otero P Anthony, Sekler Israel, Wills Zachary P, Chu Charleen T
Department of Pathology, Division of Neuropathology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213.
Department of Physiology and Cell Biology, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel, and.
J Neurosci. 2017 Nov 15;37(46):11151-11165. doi: 10.1523/JNEUROSCI.3791-16.2017. Epub 2017 Oct 16.
Mutations in leucine-rich repeat kinase 2 (LRRK2) contribute to development of late-onset familial Parkinson's disease (PD), with clinical features of motor and cognitive dysfunction indistinguishable from sporadic PD. Calcium dysregulation plays an important role in PD pathogenesis, but the mechanisms of neurodegeneration remain unclear. Recent reports indicate enhanced excitatory neurotransmission in cortical neurons expressing mutant LRRK2, which occurs before the well-characterized phenotype of dendritic shortening. As mitochondria play a major role in the rapid buffering of cytosolic calcium, we hypothesized that altered mitochondrial calcium handling contributes to dendritic retraction elicited by the LRRK2-G2019S and -R1441C mutations. In primary mouse cortical neurons, we observed increased depolarization-induced mitochondrial calcium uptake. We found that expression of mutant LRRK2 elicited transcriptional upregulation of the mitochondrial calcium uniporter (MCU) and the mitochondrial calcium uptake 1 protein (MICU1) with no change in levels of the mitochondrial calcium antiporter NCLX. Elevated MCU and MICU1 were also observed in LRRK2-mutated patient fibroblasts, along with increased mitochondrial calcium uptake, and in postmortem brains of sporadic PD/PDD patients of both sexes. Transcriptional upregulation of MCU and MICU1 was caused by activation of the ERK1/2 (MAPK3/1) pathway. Inhibiting ERK1/2 conferred protection against mutant LRRK2-induced neurite shortening. Pharmacological inhibitors or RNAi knockdown of MCU attenuated mitochondrial calcium uptake and dendritic/neuritic shortening elicited by mutant LRRK2, whereas expression of a constitutively active mutant of NCLX that enhances calcium export from mitochondria was neuroprotective. These data suggest that an increased susceptibility to mitochondrial calcium dysregulation contributes to dendritic injury in mutant LRRK2 pathogenesis. Cognitive dysfunction and dementia are common features of Parkinson's disease (PD), causing significant disability. Mutations in LRRK2 represent the most common known genetic cause of PD. We found that PD-linked LRRK2 mutations increased dendritic and mitochondrial calcium uptake in cortical neurons and familial PD patient fibroblasts, accompanied by increased expression of the mitochondrial calcium transporter MCU. Blocking the ERK1/2-dependent upregulation of MCU conferred protection against mutant LRRK2-elicited dendrite shortening, as did inhibiting MCU-mediated calcium import. Conversely, stimulating the export of calcium from mitochondria was also neuroprotective. These results implicate increased susceptibility to mitochondrial calcium overload in LRRK2-driven neurodegeneration, and suggest possible interventions that may slow the progression of cognitive dysfunction in PD.
富含亮氨酸重复激酶2（LRRK2）的突变会导致晚发性家族性帕金森病（PD）的发生，其运动和认知功能障碍的临床特征与散发性PD难以区分。钙调节异常在PD发病机制中起重要作用，但神经退行性变的机制仍不清楚。最近的报道表明，在表达突变型LRRK2的皮质神经元中兴奋性神经传递增强，这发生在特征明确的树突缩短表型之前。由于线粒体在细胞质钙的快速缓冲中起主要作用，我们推测线粒体钙处理的改变有助于LRRK2 - G2019S和 - R1441C突变引起的树突回缩。在原代小鼠皮质神经元中，我们观察到去极化诱导的线粒体钙摄取增加。我们发现突变型LRRK2的表达引起线粒体钙单向转运体（MCU）和线粒体钙摄取1蛋白（MICU1）的转录上调，而线粒体钙反向转运体NCLX的水平没有变化。在LRRK2突变的患者成纤维细胞中也观察到MCU和MICU1升高，同时线粒体钙摄取增加，在散发性PD/PDD患者的男女尸检大脑中也观察到这种情况。MCU和MICU1的转录上调是由ERK1/2（MAPK3/1）途径的激活引起的。抑制ERK1/2可保护细胞免受突变型LRRK2诱导的神经突缩短。MCU的药理抑制剂或RNA干扰敲低可减弱突变型LRRK2引起的线粒体钙摄取和树突/神经突缩短，而增强线粒体钙输出的组成型活性突变体NCLX的表达具有神经保护作用。这些数据表明，线粒体钙调节异常的易感性增加有助于突变型LRRK2发病机制中的树突损伤。认知功能障碍和痴呆是帕金森病（PD）的常见特征，会导致严重残疾。LRRK2突变是已知最常见的PD遗传病因。我们发现与PD相关的LRRK2突变增加了皮质神经元和家族性PD患者成纤维细胞中的树突和线粒体钙摄取，同时伴随着线粒体钙转运体MCU表达的增加。阻断ERK1/2依赖的MCU上调可保护细胞免受突变型LRRK2引起的树突缩短，抑制MCU介导的钙内流也有同样的效果。相反，刺激线粒体钙输出也具有神经保护作用。这些结果表明，LRRK2驱动的神经退行性变中线粒体钙超载的易感性增加，并提示可能减缓PD认知功能障碍进展的干预措施。
