The ectodomain shedding of angiotensin-converting enzyme is independent of its localisation in lipid rafts
The ectodomain shedding of angiotensin-converting enzyme is independent of its localisation in lipid rafts
复制标题
血管紧张素转换酶的胞外域脱落与其在脂筏中的定位无关
作者：
E. Parkin;F. Tan;R. Skidgel;A. Turner;N. Hooper
血管紧张素转换酶 (ACE) 是一种 I 型整合膜蛋白，在血管活性肽代谢中起主要作用，通过近膜柄内的蛋白水解裂解从质膜上脱落。为了研究这种脱落是否受到富含胆固醇的脂筏中横向分离的调节，中国仓鼠卵巢细胞和人神经母细胞瘤SH-SY5Y细胞被野生型ACE（WT-ACE）或用糖基磷脂酰肌醇（GPI）锚附着信号取代跨膜和胞质结构域（GPI-ACE）的构建体转染。在这两种细胞类型中，GPI-ACE（而不是 WT-ACE）被隔离在小窝蛋白或富含浮菌素的脂筏中，并通过磷脂酰肌醇特异性磷脂酶 C 处理从细胞表面释放。当用蛋白激酶 C 信号级联激活剂（肉豆蔻酸佛波酯或卡巴胆碱）处理细胞时，GPI-ACE 的脱落受到与 WT-ACE 相似程度的刺激。一系列基于异羟肟酸盐的锌金属蛋白酶抑制剂以相同的方式抑制细胞中 WT-ACE 和 GPI-ACE 的释放。菲律宾素处理对脂筏的破坏不会改变 GPI-ACE 的脱落，佛波酯处理不会改变 WT-ACE 或 GPI-ACE 在筏膜和非筏膜区室之间的分布。这些数据清楚地表明，蛋白激酶 C 刺激的 ACE 脱落不需要蛋白质的跨膜或胞质区域，并且脂筏中的隔离不调节蛋白质的脱落。
Angiotensin-converting enzyme (ACE), a type I integral membrane protein that plays a major role in vasoactive peptide metabolism, is shed from the plasma membrane by proteolytic cleavage within the juxtamembrane stalk. To investigate whether this shedding is regulated by lateral segregation in cholesterol-rich lipid rafts, Chinese hamster ovary cells and human neuroblastoma SH-SY5Y cells were transfected with either wild-type ACE (WT-ACE) or a construct with a glycosylphosphatidylinositol (GPI) anchor attachment signal replacing the transmembrane and cytosolic domains (GPI-ACE). In both cell types, GPI-ACE, but not WT-ACE, was sequestered in caveolin or flotillin-enriched lipid rafts and was released from the cell surface by treatment with phosphatidylinositol-specific phospholipase C. When cells were treated with activators of the protein kinase C signalling cascade (phorbol myristate acetate or carbachol) the shedding of GPI-ACE was stimulated to a similar extent to that of WT-ACE. The release of WT-ACE and GPI-ACE from the cells was inhibited in an identical manner by a range of hydroxamate-based zinc metalloprotease inhibitors. Disruption of lipid rafts by filipin treatment did not alter the shedding of GPI-ACE, and phorbol ester treatment did not alter the distribution of WT-ACE or GPI-ACE between raft and non-raft membrane compartments. These data clearly show that the protein kinase C-stimulated shedding of ACE does not require the transmembrane or cytosolic regions of the protein, and that sequestration in lipid rafts does not regulate the shedding of the protein.
DOI：
10.1016/s0021-9258(17)42144-2
发表时间：
1994-01
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
R. Ramchandran;G. Sen;K. Misono;I. Sen
通讯作者：
R. Ramchandran;G. Sen;K. Misono;I. Sen
DOI：
10.1073/pnas.86.20.7741
发表时间：
1989-10-01
影响因子：
11.1
作者：
EHLERS, MRW;FOX, EA;RIORDAN, JF
通讯作者：
RIORDAN, JF
Madin-Darby 犬肾细胞中的羧肽酶 M。
DOI：
--
发表时间：
1990
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Deddish,PA;Skidgel,RA;Kriho,VB;Li,XY;Becker,RP;Erdös,EG
通讯作者：
Erdös,EG