和厚朴酚通过UbcH8诱导AML1-ETO蛋白降解  

Honokiol-induced degradation of AML1-ETO oncoprotein through upregulation of UbcH8 in acute myeloid leukemia

作　　者：李海英[1] 周斌[1] 吴建波[1] 邢冲云[2] LI Haiying;ZHOU Bin;WU Jianbo;XING Chongyun(Laboratory of Internal Medicine, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325015, China;Department of Hematology, the First Affiliated Hospital of Wenzhou Medical University, Wenzhou 325015, China)

机构地区：[1]温州医科大学附属第一医院中心实验室,浙江温州325015 [2]温州医科大学附属第一医院血液内科,浙江温州325015

出　　处：《温州医科大学学报》2019年第5期327-332,共6页Journal of Wenzhou Medical University

基　　金：国家自然科学基金青年基金资助项目(81501809)

摘　　要：目的:研究和厚朴酚诱导AML1-ETO蛋白降解的可能机制。方法:用10、20、40 μmol/L的和厚朴酚处理Kasumi-1细胞,分别在24 h和48 h后qRT-PCR检测AML1-ETO、UbcH8 mRNA的表达;Western blot法检测蛋白表达;40 μmol/L和厚朴酚处理24 h,基因芯片分析寻找靶基因;构建UbcH8过表达质粒,通过反转录病毒转染Kasumi-1细胞,检测AML1-ETO蛋白;通过短发夹RNA(sh RNA)敲除Ubc H8的表达,40 μmol/L和厚朴酚处理48 h,同时检测AML1-ETO和UbcH8蛋白;构建异种移植小鼠模型,和厚朴酚处理,检测肿瘤的成瘤性和相关蛋白表达。结果:和厚朴酚减少Kasumi-1细胞中AML1-ETO蛋白的表达,并具有浓度和时间依赖性,但没有影响AML1-ETO mRNA的表达;和厚朴酚处理后的AML1-ETO蛋白稳定性降低;基因芯片分析发现,和厚朴酚将Ubc H8 m RNA的表达提高了约4倍;和厚朴酚处理后,Kasumi-1细胞中的Ubc H8 mRNA和蛋白的表达量均升高;过表达UbcH8的Kasumi-1细胞中,AML1-ETO蛋白表达量减少;用和厚朴酚处理Ubc H8敲除后的Kasumi-1细胞,AML1-ETO的降解被阻断;和厚朴酚抑制了异种移植小鼠模型的肿瘤生长,肿瘤中的AML1-ETO蛋白显著降低,而UBCH8表达升高。结论:和厚朴酚能通过UbcH8降解白血病细胞中的AML1-ETO蛋白。Objective: To investigate the possible mechanism of the degradation of AML1-ETO protein induced by honokiol. Methods: Kasumi-1 cells were treated with 10, 20 and 40 μmol/L honokiol for 24 h and 48 h. The mRNA expression of AML1-ETO and UbcH8 and the protein expression were detected by qRT-PCR and Western blot respectively. Microarry analysis was used to select the target gene. To explore the role of UbcH8 in the degradation of AML1-ETO protein induced by honokiol, the expression of UbcH8 was knocked down by small hairpin RNAs (shRNAs) and overexpressed by MSCV-based vector to detect the related indicators. A xenograft mice model was treated by honokiol to detect tumorigenicity and related protein expression. Results: Honokiol decreased the protein expression of AML1-ETO in a time- and concentration- dependent manner, and reduced the stability of AML1-ETO protein, but did not affect the mRNA expression of AML1-ETO in kasumi-1 cell. We identified UbcH8 as the target gene by honokiol through microarray analysis. Honokiol obviously increased the expression of UbcH8 by 4-fold. Furthermore, overexpression of UbcH8 decreased the expression of AML1-ETO and knockdown of UbcH8 by shRNAs prevented honokiol-induced degradation of AML-ETO, suggesting that UbcH8 plays a critical role in the degradation of AML1-ETO by honokiol. Finally, honokiol decreased xenograft tumor size in a xenograft leukemia mouse model. Meanwhile, the protein of AML1-ETO was significantly reduced, while UBCH8 expression was elevated. Conclusion: Honokiol can degrade AML1-ETO oncoprotein through upregulation of UbcH8 in acute myeloid leukemia.

关 键 词：和厚朴酚 AML1-ETO UbcH8 白血病 

分 类 号：R551.3[医药卫生—血液循环系统疾病]