Yablonska Svitlana, Strohlein Colleen E, Baranov Sergei V, Yeh Stacy M, Patel Aashka, Singh Tanisha, Jauhari Abhishek, Kim JinHo, Khattar Nicolas K, Li Fang, Wang Xiaomin, Chang Yue-Fang, Lee C Y Daniel, Yang X William, Carlisle Diane L, Friedlander Robert M
Neuroapoptosis Laboratory, Department of Neurological Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213.
Center for Neurobehavioral Genetics, Semel Institute for Neuroscience & Human Behavior at UCLA, Los Angeles, California 90095.
J Neurosci. 2025 Feb 19;45(8):e1254242024. doi: 10.1523/JNEUROSCI.1254-24.2024.
Huntington's disease (HD), a neurodegenerative disease, affects approximately 30,000 people in the United States, with 200,000 more at risk. Mitochondrial dysfunction caused by mutant huntingtin (mHTT) drives early HD pathophysiology. mHTT binds the translocase of the mitochondrial inner membrane (TIM23) complex, inhibiting mitochondrial protein import and altering the mitochondrial proteome. The 17 aa HTT N-terminal sequence (N17) acts as a regulatory domain in HD pathogenesis; phosphomimetic modification of serines 13 and 16 of the N17 domain impacts subcellular localization and degradation and ameliorates toxicity in mouse and cell models of HD. Using cellular and mouse (either sex) HD models, we investigated the mechanisms by which HTT phosphorylation affects intracellular localization. We demonstrate that introducing phosphomimetic mutations within the mHTT fragment N17 domain decreased TIM23 binding affinity and reduced inhibition of mHTT-mediated mitochondrial protein import. BACHD-SD mice expressing full-length mHTT harboring the same two N17 phosphomimetic mutations have an ameliorated HD-like phenotype as compared with mice expressing mHTT. Consistent with reduced toxicity in vivo, we found that the amount of full-length mHTT in the brain mitochondria of BACHD-SD transgenic mice is less when the mHTT has two phosphomimetic mutations. To complement the relevance of the phosphomimetic HTT findings, endogenous N17 phospho-mHTT is less likely to translocate to the mitochondria compared with nonphosphorylated mHTT. We demonstrate that phosphorylation of mHTT at serines 13 and 16 is critical for negatively regulating mHTT mitochondrial targeting and that reducing mHTT mitochondrial localization and binding to TIM23 results in amelioration of mHTT-induced mitochondrial and neuronal toxicity.
亨廷顿舞蹈症（HD）是一种神经退行性疾病，在美国约有3万人受其影响，另外还有20万人面临患病风险。突变型亨廷顿蛋白（mHTT）导致的线粒体功能障碍驱动了HD早期的病理生理过程。mHTT与线粒体内膜转位酶（TIM23）复合体结合，抑制线粒体蛋白导入并改变线粒体蛋白质组。17个氨基酸的HTT N端序列（N17）在HD发病机制中作为一个调节结构域；N17结构域丝氨酸13和16的磷酸模拟修饰影响亚细胞定位和降解，并改善HD小鼠和细胞模型中的毒性。利用细胞和小鼠（雌雄均可）HD模型，我们研究了HTT磷酸化影响细胞内定位的机制。我们证明，在mHTT片段N17结构域内引入磷酸模拟突变会降低TIM23结合亲和力，并减少mHTT介导的线粒体蛋白导入的抑制作用。与表达mHTT的小鼠相比，表达具有相同两个N17磷酸模拟突变的全长mHTT的BACHD-SD小鼠具有改善的HD样表型。与体内毒性降低一致，我们发现当mHTT具有两个磷酸模拟突变时，BACHD-SD转基因小鼠脑线粒体中全长mHTT的量较少。为补充磷酸模拟HTT研究结果的相关性，与未磷酸化的mHTT相比，内源性N17磷酸化mHTT向线粒体转位的可能性较小。我们证明，mHTT丝氨酸13和16的磷酸化对于负向调节mHTT的线粒体靶向至关重要，并且减少mHTT的线粒体定位和与TIM23的结合会改善mHTT诱导的线粒体和神经元毒性。
