Molecular Neuropathobiology Laboratory, Department of Molecular Life Sciences, Tokai University School of Medicine, Isehara, Kanagawa, 259-1193, Japan.
Molecular Neuropathobiology Laboratory, Department of Molecular Life Sciences, Tokai University School of Medicine, Isehara, Kanagawa, 259-1193, Japan; The Institute of Medical Sciences, Tokai University, Isehara, Kanagawa, 259-1193, Japan; Micro/Nano Technology Center, Tokai University, Hiratsuka, Kanagawa, 259-1292, Japan.
Neurochem Int. 2022 Sep;158:105364. doi: 10.1016/j.neuint.2022.105364. Epub 2022 May 28.
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by selective loss of motor neurons in the brain and spinal cord. Recent studies have shown that mutations in SQSTM1 are linked to ALS. It has also been demonstrated that a systemic loss of SQSTM1 exacerbates disease phenotypes in an ALS mouse model. However, it is still unclear whether and how SQSTM1 in the central nervous system (CNS) specifically regulates ALS-associated disease phenotypes. To address this issue, we generated CNS-specific Sqstm1 deficient SOD1 transgenic mice, and conducted gross phenotype analyses as well as the immunohistochemical and biochemical examinations of spinal cord tissues using these mice. CNS-specific SQSTM1 deficiency accelerated the disease onset and shortened the lifespan of SOD1 mice. The CNS-specific SQSTM1 ablation also resulted in increased number of ubiquitin-positive aggregates, while their size rather became much smaller. Remarkably, ubiquitin-positive aggregates, which were usually present in extracellular space and/or neuropil in SOD1 mice, were preferentially localized to soma and neurites of spinal neurons in CNS-specific SQSTM1 deficient SOD1 mice. Next, to further clarify the function of SQSTM1 in neurons, we investigated the contribution of SQSTM1 to the accumulation of polyubiquitinated proteins in relation to the ubiquitin proteasome system (UPS) and the autophagy-endolysosomal system (APELS) in primary cultured motor neurons (PMNs). Loss of SQSTM1 in PMNs resulted in decreased accumulation of insoluble polyubiquitinated proteins, which was induced by simultaneous treatment with proteasome and lysosome inhibitors, suggesting a pivotal role of SQSTM1 in the formation of insoluble protein aggregates. However, SQSTM1 silencing had a limited impact on the susceptibility to proteasome and/or lysosome inhibitor-induced apoptosis in PMNs. Taken together, neuronal SQSTM1, whose functions are associated with both the UPS and APELS, might primarily regulate the distribution and accumulation of misfolded protein aggregates in the CNS, thereby protecting neurons from degeneration in mice.
肌萎缩侧索硬化症（ALS）是一种进行性神经退行性疾病，其特征是大脑和脊髓中的运动神经元选择性丧失。最近的研究表明，SQSTM1 的突变与 ALS 有关。已经证明，全身性 SQSTM1 的缺失会加剧 ALS 小鼠模型中的疾病表型。然而，目前尚不清楚中枢神经系统（CNS）中的 SQSTM1 是否以及如何特异性调节与 ALS 相关的疾病表型。为了解决这个问题，我们生成了中枢神经系统特异性 Sqstm1 缺陷的 SOD1 转基因小鼠，并使用这些小鼠进行了大体表型分析以及脊髓组织的免疫组织化学和生化检查。中枢神经系统特异性 SQSTM1 缺失加速了疾病的发作并缩短了 SOD1 小鼠的寿命。中枢神经系统特异性 SQSTM1 消融还导致泛素阳性聚集体数量增加，而其大小变得更小。值得注意的是，通常存在于 SOD1 小鼠细胞外空间和/或神经间质中的泛素阳性聚集体，在中枢神经系统特异性 SQSTM1 缺陷的 SOD1 小鼠中优先定位于脊髓神经元的体和突起。接下来，为了进一步阐明 SQSTM1 在神经元中的功能，我们研究了 SQSTM1 对聚泛素化蛋白在与泛素蛋白酶体系统（UPS）和自噬-溶酶体系统（APELS）相关的积累中的贡献在原代培养的运动神经元（PMNs）中。PMNs 中 SQSTM1 的缺失导致同时用蛋白酶体和溶酶体抑制剂诱导的不溶性聚泛素化蛋白积累减少，表明 SQSTM1 在不溶性蛋白聚集体的形成中起关键作用。然而，SQSTM1 沉默对 PMNs 对蛋白酶体和/或溶酶体抑制剂诱导的凋亡的敏感性影响有限。总之，神经元 SQSTM1 的功能与 UPS 和 APELS 相关，可能主要调节 CNS 中错误折叠蛋白聚集体的分布和积累，从而保护神经元免受小鼠变性。
