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KIF5A-dependent axonal transport deficiency disrupts autophagic flux in trimethyltin chloride-induced neurotoxicity
Autophagy ( IF 14.6 ) Pub Date : 2020-03-30 , DOI: 10.1080/15548627.2020.1739444
Mengyu Liu 1 , Huifeng Pi 1, 2 , Yu Xi 3 , Liting Wang 4 , Li Tian 1 , Mengyan Chen 1 , Jia Xie 1 , Ping Deng 1 , Tao Zhang 1 , Chao Zhou 1 , Yidan Liang 5 , Lei Zhang 1 , Mindi He 1 , Yonghui Lu 1 , Chunhai Chen 1 , Zhengping Yu 1 , Zhou Zhou 3
Affiliation  

Trimethyltin chloride (TMT) is widely used as a constituent of fungicides and plastic stabilizers in the industrial and agricultural fields, and is generally acknowledged to have potent neurotoxicity, especially in the hippocampus; however, the mechanism of induction of neurotoxicity by TMT remains elusive. Herein, we exposed Neuro-2a cells to different concentrations of TMT (2, 4, and 8 μM) for 24 h. Proteomic analysis, coupled with bioinformatics analysis, revealed the important role of macroautophagy/autophagy-lysosome machinery in TMT-induced neurotoxicity. Further analysis indicated significant impairment of autophagic flux by TMT via suppressed lysosomal function, such as by inhibiting lysosomal proteolysis and changing the lysosomal pH, thereby contributing to defects in autophagic clearance and subsequently leading to nerve cell death. Mechanistically, molecular interaction networks of Ingenuity Pathway Analysis identified a downregulated molecule, KIF5A (kinesin family member 5A), as a key target in TMT-impaired autophagic flux. TMT decreased KIF5A protein expression, disrupted the interaction between KIF5A and lysosome, and impaired lysosomal axonal transport. Moreover, Kif5a overexpression restored axonal transport, increased lysosomal dysfunction, and antagonized TMT-induced neurotoxicity in vitro. Importantly, in TMT-administered mice with seizure symptoms and histomorphological injury in the hippocampus, TMT inhibited KIF5A expression in the hippocampus. Gene transfer of Kif5a enhanced autophagic clearance in the hippocampus and alleviated TMT-induced neurotoxicity in vivo. Our results are the first to demonstrate KIF5A-dependent axonal transport deficiency to cause autophagic flux impairment via disturbance of lysosomal function in TMT-induced neurotoxicity; manipulation of KIF5A may be a therapeutic approach for antagonizing TMT-induced neurotoxicity.

Abbreviations: 3-MA: 3-methyladenine; AAV: adeno-associated virus; ACTB: actin beta; AGC: automatic gain control; ATG: autophagy-related; ATP6V0D1: ATPase H+ transporting lysosomal V0 subunit D1; ATP6V1E1: ATPase H+ transporting lysosomal V1 subunit E1; CA: cornu ammonis; CQ: chloroquine; CTSB: cathepsin B; CTSD: cathepsin D; DCTN1: dynactin subunit 1; DG: dentate gyrus; DYNLL1: dynein light chain LC8-type 1; FBS: fetal bovine serum; GABARAP: GABA type A receptor-associated protein; GABARAPL1: GABA type A receptor associated protein like 1; GABARAPL2: GABA type A receptor associated protein like 2; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; IPA: Ingenuity Pathway Analysis; KEGG: Kyoto Encyclopedia of Genes and Genomes; KIF5A: kinesin family member 5A; LAMP: lysosomal-associated membrane protein; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; NBR1: NBR1 autophagy cargo receptor; OPTN: optineurin; PBS: phosphate-buffered saline; PFA: paraformaldehyde; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PRM: parallel reaction monitoring; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; SYP: synaptophysin; TAX1BP1: Tax1 binding protein 1; TMT: trimethyltin chloride; TUB: tubulin.



中文翻译:


KIF5A 依赖性轴突运输缺陷破坏三甲基氯化锡诱导的神经毒性中的自噬通量



三甲基氯化锡(TMT)在工业和农​​业领域广泛用作杀菌剂和塑料稳定剂的成分,并且普遍认为具有强烈的神经毒性,特别是在海马体中;然而,TMT 诱导神经毒性的机制仍不清楚。在此,我们将 Neuro-2a 细胞暴露于不同浓度的 TMT(2、4 和 8 μM)24 小时。蛋白质组学分析与生物信息学分析相结合,揭示了巨自噬/自噬-溶酶体机制在 TMT 诱导的神经毒性中的重要作用。进一步的分析表明,TMT 通过抑制溶酶体功能,例如通过抑制溶酶体蛋白水解和改变溶酶体 pH 值,显着损害自噬流,从而导致自噬清除缺陷,随后导致神经细胞死亡。从机制上讲,Ingenuity Pathway Analysis 的分子相互作用网络确定了一种下调的分子 KIF5A(驱动蛋白家族成员 5A),作为 TMT 受损的自噬通量的关键靶标。 TMT 降低 KIF5A 蛋白表达,破坏 KIF5A 与溶酶体之间的相互作用,并损害溶酶体轴突运输。此外, Kif5a过表达可恢复轴突运输,增加溶酶体功能障碍,并在体外拮抗 TMT 诱导的神经毒性。重要的是,在给予 TMT 的具有癫痫症状和海马组织形态学损伤的小鼠中,TMT 抑制海马中 KIF5A 的表达。 Kif5a的基因转移增强了海马中的自噬清除并减轻了 TMT 诱导的体内神经毒性。 我们的结果首次证明 KIF5A 依赖性轴突运输缺陷在 TMT 诱导的神经毒性中通过扰乱溶酶体功能而导致自噬流损伤;操纵 KIF5A 可能是对抗 TMT 诱导的神经毒性的一种治疗方法。


缩写:3-MA:3-甲基腺嘌呤; AAV:腺相关病毒; ACTB:肌动蛋白β; AGC:自动增益控制; ATG:自噬相关; ATP6V0D1:ATPase H +转运溶酶体 V0 亚基 D1; ATP6V1E1:ATPase H +转运溶酶体 V1 亚基 E1; CA: 山角; CQ:氯喹; CTSB:组织蛋白酶B; CTSD:组织蛋白酶D; DCTN1: 动力蛋白亚基 1; DG:齿状回; DYNLL1:动力蛋白轻链 LC8 型 1; FBS:胎牛血清; GABARAP:GABA A 型受体相关蛋白; GABARAPL1:GABA A 型受体相关蛋白样 1; GABARAPL2:GABA A 型受体相关蛋白样 2; GAPDH:3-磷酸甘油醛脱氢酶; IPA:独创性路径分析; KEGG:京都基因和基因组百科全书; KIF5A:驱动蛋白家族成员5A; LAMP:溶酶体相关膜蛋白; MAP1LC3B/LC3B:微管相关蛋白1轻链3β; NBR1:NBR1自噬货物受体; OPTN: 奥替神经氨酸; PBS:磷酸盐缓冲盐水; PFA:多聚甲醛; PIK3C3/VPS34:磷脂酰肌醇3-激酶催化亚基3型; PRM:平行反应监测; siRNA:小干扰RNA; SQSTM1/p62: 隔离体 1; SYP: 突触素; TAX1BP1:Tax1结合蛋白1; TMT:三甲基氯化锡; TUB:微管蛋白。

更新日期:2020-03-30
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