Department of Chemico-Pharmacological Sciences, Graduate School of Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan.
Department of Neurology, University of Michigan, Ann Arbor, MI, USA.
J Pharmacol Sci. 2023 Jul;152(3):182-192. doi: 10.1016/j.jphs.2023.05.002. Epub 2023 May 11.
Missense mutations of ubiquilin 2 (UBQLN2) have been identified to cause X-linked amyotrophic lateral sclerosis (ALS). Proteasome-mediated protein degradation is reported to be impaired by ALS-associated mutations of UBQLN2. However, it remains unknown how these mutations affect autophagy-lysosome protein degradation, which consists of macroautophagy (MA), microautophagy (mA), and chaperone-mediated autophagy (CMA). Using a CMA/mA fluorescence reporter we found that overexpression of wild-type UBQLN2 impairs CMA. Conversely, knockdown of endogenous UBQLN2 increases CMA activity, suggesting that normally UBQLN2 negatively regulates CMA. ALS-associated mutant forms of UBQLN2 exacerbate this impairment of CMA. Using cells stably transfected with wild-type or ALS-associated mutant UBQLN2, we further determined that wild-type UBQLN2 increased the ratio of LAMP2A (a CMA-related protein) to LAMP1 (a lysosomal protein). This could represent a compensatory reaction to the impairment of CMA by wild-type UBQLN2. However, ALS-associated mutant UBQLN2 failed to show this compensation, exacerbating the impairment of CMA by mutant UBQLN2. We further demonstrated that ALS-associated mutant forms of UBQLN2 also impair MA, but wild-type UBQLN2 does not. These results support the view that ALS-associated mutant forms of UBQLN2 impair both CMA and MA which may contribute to the neurodegeneration observed in patients with UBQLN2-mediated ALS.
泛素结合酶 2（UBQLN2）的错义突变已被鉴定为导致 X 连锁肌萎缩侧索硬化症（ALS）的原因。据报道，蛋白酶体介导的蛋白质降解被 ALS 相关的 UBQLN2 突变所损害。然而，这些突变如何影响自噬溶酶体蛋白降解仍然未知，自噬溶酶体蛋白降解由巨自噬（MA）、微自噬（mA）和伴侣介导的自噬（CMA）组成。使用 CMA/MA 荧光报告物，我们发现野生型 UBQLN2 的过表达会损害 CMA。相反，内源性 UBQLN2 的敲低会增加 CMA 活性，表明正常情况下 UBQLN2 负调节 CMA。ALS 相关的突变型 UBQLN2 加剧了这种 CMA 的损害。使用稳定转染野生型或 ALS 相关突变型 UBQLN2 的细胞，我们进一步确定野生型 UBQLN2 增加了 LAMP2A（一种 CMA 相关蛋白）与 LAMP1（一种溶酶体蛋白）的比值。这可能代表了野生型 UBQLN2 对 CMA 损害的代偿反应。然而，ALS 相关的突变型 UBQLN2 未能表现出这种代偿作用，加剧了突变型 UBQLN2 对 CMA 的损害。我们进一步证明，ALS 相关的突变型 UBQLN2 也会损害 MA，但野生型 UBQLN2 不会。这些结果支持了 ALS 相关的突变型 UBQLN2 损害 CMA 和 MA 的观点，这可能导致 UBQLN2 介导的 ALS 患者中观察到的神经退行性变。
