Short-Chain Ubiquitination Is Associated with the Degradation Rate of a Cell-Surface-Resident Bile Salt Export Pump (BSEP/ABCB11)
Short-Chain Ubiquitination Is Associated with the Degradation Rate of a Cell-Surface-Resident Bile Salt Export Pump (BSEP/ABCB11)
复制标题
DOI：
10.1124/mol.108.049288
复制
发表时间：
2009-01-01
影响因子：
3.6
通讯作者：
Sugiyama, Yuichi
中科院分区：
文献类型：
--
作者：
Hayashi, Hisamitsu;Sugiyama, Yuichi
小管膜上胆盐输出泵（BSEP/ABCB 11）的表达减少与胆汁淤积诱导的肝毒性有关，这是由于胆汁酸在肝细胞中蓄积所致。我们以前证明，4-苯基丁酸（4PBA）处理，一个U。S.美国食品药品监督管理局批准用于治疗尿素循环障碍的药物，通过延长细胞表面驻留BSEP的降解速率诱导BSEP的细胞表面表达。另一方面，在进行性家族性肝内胆汁淤积症2型（PFIC 2）中发现的BSEP突变E297 G和D482 G通过缩短细胞表面驻留BSEP的降解速率来降低它。因此，为了帮助胆汁淤积的药物治疗的发展，我们研究了4PBA和PFIC 2型突变影响BSEP从细胞表面降解的潜在机制，重点是短链泛素化。在表达BSEP和大鼠泪小管膜囊泡的Madin-Darby犬肾II（MDCK II）细胞中，成熟形式的BSEP/Bsep的分子量在泛蛋白修饰后从170 kDa变为190 kDa（分子量，8 kDa）。BSEP/Bsep的泛素化敏感性在体外和体内通过4PBA处理降低，相反，通过BSEP突变E297 G和D482 G增强。此外，使用MDCK II细胞的生物素标记的研究表明，细胞表面驻留的嵌合蛋白融合泛素BSEP的降解速度比BSEP本身。总之，BSEP/Bsep被两到三个泛素修饰，并且其泛素化通过4PBA处理和PFIC 2型突变来调节。短链泛素化的调节可以调节4PBA处理和PFIC 2型突变引起的细胞表面驻留BSEP降解速率的变化。
The reduced expression of the bile salt export pump (BSEP/ABCB11) at the canalicular membrane is associated with cholestasis-induced hepatotoxicity due to the accumulation of bile acids in hepatocytes. We demonstrated previously that 4-phenylbutyrate (4PBA) treatment, a U. S. Food and Drug Administration-approved drug for the treatment of urea cycle disorders, induces the cell-surface expression of BSEP by prolonging the degradation rate of cell-surface-resident BSEP. On the other hand, BSEP mutations, E297G and D482G, found in progressive familial intrahepatic cholestasis type 2 (PFIC2), reduced it by shortening the degradation rate of cell-surface-resident BSEP. Therefore, to help the development of the medical treatment of cholestasis, we investigated the underlying mechanism by which 4PBA and PFIC2-type mutations affect the BSEP degradation from cell surface, focusing on short-chain ubiquitination. In Madin-Darby canine kidney II (MDCK II) cells expressing BSEP and rat canalicular membrane vesicles, the molecular mass of the mature form of BSEP/Bsep shifted from 170 to 190 kDa after ubiquitin modification (molecular mass, 8 kDa). Ubiquitination susceptibility of BSEP/Bsep was reduced in vitro and in vivo by 4PBA treatment and, conversely, was enhanced by BSEP mutations E297G and D482G. Moreover, biotin-labeling studies using MDCK II cells demonstrated that the degradation of cell-surface-resident chimeric protein fusing ubiquitin to BSEP was faster than that of BSEP itself. In conclusion, BSEP/Bsep is modified with two to three ubiquitins, and its ubiquitination is modulated by 4PBA treatment and PFIC2-type mutations. Modulation of short-chain ubiquitination can regulate the change in the degradation rate of cell-surface-resident BSEP by 4PBA treatment and PFIC2-type mutations.