Homoharringtonine binds to and increases myosin-9 in myeloid leukaemia
Homoharringtonine binds to and increases myosin-9 in myeloid leukaemia
复制标题
高三尖杉酯碱结合并增加髓系白血病中的肌球蛋白 9
DOI：
10.1111/bph.13359
复制
发表时间：
2016-01-01
影响因子：
7.3
通讯作者：
Jin, Jie
中科院分区：
文献类型：
--
作者：
Zhang, Ting;Shen, Shuijie;Jin, Jie
背景和目的高三尖杉酯碱 (HHT) 是从多种三尖杉属物种中分离出来的天然生物碱。 HHT已用于治疗急性粒细胞白血病（AML）、慢性粒细胞白血病（CML）、慢性淋巴细胞白血病和骨髓增生异常综合征。尽管临床前研究中HHT抑制蛋白质合成并促进白血病细胞凋亡，但其分子靶蛋白仍不清楚。本研究的目的是鉴定HHT的靶蛋白。实验方法我们合成了生物素化亲和柱，用它来鉴定HHT的靶蛋白，并通过MS和Western blots证实了结果。我们还通过蛋白质印迹、流式细胞术和逆转录病毒转染检查了 HHT 对靶蛋白的影响，并确定了靶蛋白在 HHT 抗白血病活性中的作用。 关键结果肌球蛋白超家族的成员 Myosin-9 被确定为 HHT 的直接相互作用因子。此外，HHT 以时间依赖性方式上调 AML 和 CML 细胞系中 myosin-9 的表达水平。因此，HHT诱导的白血病细胞凋亡在6小时内开始，并持续增加24小时。上调的 myosin-9 表达水平与 HHT 介导的凋亡细胞百分比增加呈正相关。过表达myosin-9可以增加白血病细胞对HHT细胞毒性的敏感性，并使细胞阻滞于S期和G2/M期。结论与意义我们的研究结果表明，myosin-9是HHT的靶蛋白，在HHT诱导的白血病细胞凋亡中发挥重要作用。
Background and PurposeHomoharringtonine (HHT) is a natural alkaloid isolated from various Cephalotaxus species. HHT has been used to treat acute myeloid leukaemia (AML), chronic myeloid leukaemia (CML), chronic lymphocyte leukaemia and myelodysplastic syndromes. Although HHT inhibits protein synthesis and promotes apoptosis of leukaemia cells in preclinical studies, its molecular target proteins remain unknown. The aim of this study was to identify target proteins of HHT.Experimental ApproachWe have synthesized a biotinylated affinity column and used it to identify targets of HHT and confirmed the results by MS and Western blots. We also examined the effects of HHT on the target protein and determined roles of the target protein in anti-leukaemia activities of HHT through Western blots, flow cytometry and retrovirus transfection.Key ResultsMyosin-9, a member of the myosin super-family, was identified as a direct interactor of HHT. Furthermore, HHT up-regulated the expression level of myosin-9 in both AML and CML cell lines in a time-dependent manner. Thus, HHT-induced apoptosis of leukaemia cells begins in 6h and continues to increase for 24h. There is a positive correlation between up-regulated myosin-9 expression level and increased percentage of apoptotic cells mediated by HHT. Overexpression of myosin-9 could increase the sensitivity of the leukaemia cells to the cytotoxicity of HHT and arrest cells in S and G2/M phases.Conclusions and ImplicationsOur results indicated that myosin-9 was the target protein of HHT and played an important role in the HHT-induced apoptosis of leukaemia cells.