Wang F, Buckley N E, Olivera A, Goodemote K A, Su Y, Spiegel S
Department of Biochemistry and Molecular Biology, Georgetown University Medical Center, Washington, DC 20007, USA.
Glycoconj J. 1996 Dec;13(6):937-45. doi: 10.1007/BF01053189.
The B subunit of cholera toxin, which binds specifically to ganglioside GM1, is mitogenic for quiescent Swiss 3T3 fibroblasts. Recently, sphingolipids metabolites, ceramide, sphingosine and sphingosine-1-phosphate, have been implicated as second messengers in cell growth regulation and differentiation. In this paper, we examined the possibility that interaction of the B subunit with membrane GM1 leads to alterations in metabolism of glycosphingolipids and that increased levels of sphingolipids metabolites may mediate the biological effects of the B subunit. While the B subunit did not induce a change in the level of ceramide or sphingosine, the level of sphingosine-1-phosphate was rapidly and transiently increased. The B subunit also transiently activated cytosolic sphingosine kinase activity, which catalyzes the phosphorylation of the primary hydroxyl group of sphingosine to produce sphingosine-1-phosphate. To determine whether the increase in sphingosine-1-phosphate level plays a role in B subunit-induced mitogenicity, we used a competitive inhibitor of sphingosine kinase, D,L-threo-dihydrosphingosine. D,L-thereo-Dihydrosphingosine not only inhibited B subunit-induced DNA synthesis by 26%, it also reduced its ability to stimulate DNA-binding activity of the transcription factor AP-1. This sphingosine kinase inhibitor also inhibited B subunit-induced increases in the activity of cell cycle-regulated, cyclin-dependent serine/threonine kinases, cdk2 and p34cdc2. These findings suggest that sphingosine-1-phosphate may play a role in the signal transduction pathways activated by binding of the B subunit to endogenous ganglioside GM1.
霍乱毒素的B亚基可特异性结合神经节苷脂GM1，对静止的瑞士3T3成纤维细胞具有促有丝分裂作用。最近，鞘脂代谢产物神经酰胺、鞘氨醇和1-磷酸鞘氨醇被认为是细胞生长调节和分化中的第二信使。在本文中，我们研究了B亚基与膜GM1相互作用导致鞘糖脂代谢改变以及鞘脂代谢产物水平升高可能介导B亚基生物学效应的可能性。虽然B亚基未引起神经酰胺或鞘氨醇水平的变化，但1-磷酸鞘氨醇的水平迅速且短暂升高。B亚基还短暂激活了胞质鞘氨醇激酶活性，该酶催化鞘氨醇伯羟基磷酸化生成1-磷酸鞘氨醇。为了确定1-磷酸鞘氨醇水平的升高是否在B亚基诱导的促有丝分裂中起作用，我们使用了鞘氨醇激酶的竞争性抑制剂D,L-苏式-二氢鞘氨醇。D,L-苏式-二氢鞘氨醇不仅抑制B亚基诱导的DNA合成达26%，还降低了其刺激转录因子AP-1 DNA结合活性的能力。这种鞘氨醇激酶抑制剂还抑制B亚基诱导的细胞周期调节的细胞周期蛋白依赖性丝氨酸/苏氨酸激酶cdk2和p34cdc2活性的增加。这些发现表明，1-磷酸鞘氨醇可能在B亚基与内源性神经节苷脂GM1结合激活的信号转导途径中起作用。
