Induction of glucosylceramide synthase by synthase inhibitors and ceramide
Induction of glucosylceramide synthase by synthase inhibitors and ceramide
复制标题
DOI：
10.1016/0005-2760(95)00217-0
复制
发表时间：
1996-02-16
期刊：
影响因子：
--
通讯作者：
Shayman, JA
中科院分区：
文献类型：
--
作者：
Abe, A;Radin, NS;Shayman, JA
葡萄糖神经酰胺（GlcCer）合酶作用于鞘脂神经酰胺，从UDP-glc转移葡萄糖部分，从而形成葡萄糖鞘脂大家族的第一个成员。已发现该酶的两种抑制剂，o-苏型-1-苯基-2-癸酰氨基-3-吗啉代-1-丙醇（D-苏型-PdR）和N-丁基脱氧野尻霉素（NBDN）在MDCK细胞中诱导升高的合酶水平。在用20 μ M Pd 4处理的细胞中，然后在吸收的Pd 4被部分稀释的条件下测定合成酶活性，该测定表明，酶的比活性仅在1小时内就显著升高，并在6小时内达到最大值，约为对照活性的3倍。放线菌酮和放线菌素D，翻译和转录蛋白质合成的抑制剂，导致大部分的合酶活性消失在6小时，大概是因为正常的分解代谢破坏。然而，在细胞培养基中同时包含PdR或NBDN减慢了合酶消失的速率。L-Cycloserine阻断了神经酰胺的合成，但却能使PdR提高合成酶的活性。因此，诱导效应似乎至少部分是由于酶-抑制剂复合物对酶降解的正常过程的抗性。GlcCer合酶的另外两种抑制剂，比PdR活性更高，没有产生可检测的诱导，因为它们在细胞洗涤和稀释步骤期间不能与酶解离。产生内源性细胞神经酰胺水平大幅增加的药物（DL-赤藓糖醇、N-乙酰鞘氨醇、酸性细菌鞘磷脂酶）也诱导GlcCer合酶水平升高。后两种试剂在放线菌酮存在下不能保护合酶免受催化剂的破坏。这些发现表明存在酶诱导的第二种机制，由于酶的脂质底物的可用性增加而增强酶的合成。神经酰胺参与细胞信号传导的事件可能部分由鞘糖脂水平的变化介导。
Glucosylceramide (GlcCer) synthase acts on the sphingolipid, ceramide, to transfer a glucose moiety from UDP-glc, thus forming the first member of a large family of glucosphingolipids. Two inhibitors of the enzyme, o-threo-1-phenyl-2-decanoylamino-3-morpholino-1- propanol (D-threo-PDMP) and N-butyldeoxynojirimycin (NBDN), have been found to induce an elevated level of the synthase in MDCK cells. In cells treated with 20 mu M PDMP, then assayed for synthase activity under conditions in which the absorbed PDMP was partially diluted out, the assay showed that the enzyme's specific activity had risen considerably in only 1 h and reached a maximum of about three times the control activity within 6 h. Both cycloheximide and actinomycin D, inhibitors of translational and transcriptional protein synthesis, caused much of the synthase activity to disappear in 6 h, presumably because of normal catabolic destruction. However, simultaneous inclusion of PDMP or NBDN in the cell medium slowed the rate of synthase disappearance. L-Cycloserine, which blocked the synthesis of ceramide, nevertheless allowed PDMP to elevate the synthase activity. Thus the inductive effect appears to be due, in part at least, to resistance of the enzyme-inhibitor complex to the normal process of enzyme degradation. Two other inhibitors of GlcCer synthase, more active than PDMP, did not produce detectable induction because they could not be dissociated from the enzyme during the cell washing and diluting steps. Agents that produced a large increase in endogenous cell ceramide level (DL-erythuo-PDMP, N-acetylsphingosine, acid bacterial sphingomyelinase) also induced an elevated level of GlcCer synthase. The latter two agents did not protect the synthase from catabolism in the presence of cycloheximide. These findings suggest the existence of a second mechanism of enzyme induction, enhanced synthesis of the enzyme due to the increased availability of the enzyme's lipoidal substrate. The possibility is raised that events involving ceramide in cell signalling may be mediated in part by changes in glucosphingolipid levels.