β-arrestin1/2抑制SphK1蛋白质表达研究  

Inhibition of SphK1 expression by β-arrestin1/2

作　　者：刘永健 卞金俊[3] 花金宝 吴跃军[4] 陈妍[1,5] 殷武[2] 

机构地区：[1]中国中医科学院广安门医院,北京100053 [2]南京大学生命科学学院,南京210023 [3]第二军医大学长海医院麻醉与重症医学科,上海200433 [4]江苏省中医院检验科,南京210029 [5]江苏省肿瘤医院,南京210009

出　　处：《南京大学学报（自然科学版）》2017年第6期1203-1211,共9页Journal of Nanjing University（Natural Science）

基　　金：国家自然科学基金(81473293;81171843;81673462;81671939;91540119);上海市科委项目(15411963200);中国博士后科学基金(2016M591365);江苏省重点研发项目(BE2017712);江苏省中医管理局(LZ13230)

摘　　要：神经鞘氨醇激酶1(SphK1)在鞘脂类代谢中起关键作用.研究表明,SphK1在肝癌等多种肿瘤细胞中过度表达,促进肿瘤细胞生长与转移,而阻断SphK1促进肿瘤细胞凋亡.因此,SphK1可以作为肝癌治疗的潜在靶点,但到目前为止,SphK1蛋白质降解调控机制并不清楚.采用Western blot、RT-PCR和siRNA干扰等方法研究SphK1蛋白质降解机制,发现β-arrestin1/2可能通过泛素化途径促进SphK1蛋白质降解.蛋白酶体抑制剂MG132能明显上调外源性和内源性SphK1蛋白水平,过表达β-arrestin1/2显著下调人源胚胎肾细胞HEK293T细胞和人肝癌细胞HepG2细胞中SphK1蛋白水平,且呈剂量依赖性,MG132可以缓解β-arrestin1/2对SphK1的降解作用;β-arrestin1/2干扰片段处理后,HEK293与HepG2细胞内内源性SphK1蛋白表达显著升高.证明β-arrestin1/2可以通过蛋白质泛素化方式促进SphK1降解,这为设计SphK1特异性抑制剂提供理论依据.Sphingosine kinase type 1（SphK1）is a critical regulator of sphingolipid metabolites.Increasing evidence indicates that SphK1 is highly expressed in human liver cancer,and up regulation of SphK1 expression promotes cancer cell growth and metastasis.In contrast,down regulation of SphK1 expression triggers cancer cell apoptosis suggests the SphK1 could be a potential target for liver cancer therapy.However,the underlying mechanism forSphK1 degradation remains unclear.Here,western blotting,quantitative RT-PCR,and small interfering RNA（siRNA）technologies were used to investigate the mechanism of SphK1 degradation.As a result,the expression of both exogenous and endogenous SphK1 were significantly up-regulated when HEK293 Tcells and HepG2 cells were treated with the proteasome inhibitor MG132.Moreover,the forced over-expression ofβ-arrestin1/2 dose-dependently suppressed SphK1 protein level,and this effect could be attenuated by MG132.Knockdown ofβ-arrestin1/2 expressions in HEK293 Tcells and HepG2 cells led to a significant increase in cellular SphK1 protein level.In conclusion,β-arrestin1/2 could promote SphK1 degradation probably through ubiquitin-mediated pathway,the result of which offers theoretical basis for the development of specific SphK1 inhibitor.

关 键 词：β-arrestin1/2 神经鞘氨醇激酶1(SphK1) 泛素化 蛋白质降解 

分 类 号：Q5-33[生物学—生物化学]