Generation of diacylglycerol and ceramide during homologous complement activation.
Formation of sublytic terminal complement complexes (TCC) on nucleated cells produces transient increase in [Ca2+]i and activates protein kinase C. The present study is to evaluate whether TCC can generate endogenous signal messengers other than Ca2+ that regulate cell activities by measuring mass-levels of sn-1,2-diacylglycerol (DAG) and ceramide. As targets, lymphoblastoid human B cell lines JY25 and its mutant JY5 were used. JY5, cells deficient in glycosylphosphatidylinositol-anchored proteins with higher lytic susceptibility to human complement, are four times more efficient in forming C5b-9. When cells sensitized with limited anti-class II IgG were exposed to human serum to generate sublytic TCC, a sustained increase in DAG and ceramide was observed with a maximum 3.6-fold DAG increase over basal level in JY25 and 2.8-fold in JY5, and 6.3-fold ceramide increase in JY25 and 2.8-fold in JY5. The effect of TCC was evaluated with C7-deficient human serum (C7D) +/- C7 and also with C5b6, C7, C8, and C9 proteins. The DAG and ceramide increase by C7D + C7 over C7D control were 1.6- and 1.8-fold, respectively, in JY25, and 2.3-, and two-fold in JY5. TCC activation also induced an increased hydrolysis of sphyingomyelin and phosphatidylcholine. In addition, DAG increase by TCC was primarily achieved by C5b-7 and preincubation of cells with pertussis toxininhibited DAG increase, suggesting an involvement of a pertussis toxin-sensitive GTP-binding protein. As important signal transduction molecules, DAG and ceramide generated in response to TCC assembly, could participate in cell activation during inflammation and repair.
Niculescu
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University of Maryland, School of Medicine, Department of Pathology, Baltimore 21201.
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Shin
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Lang
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D000818:Animals; D002461:Cell Line, Transformed; D002518:Ceramides / Q000096:biosynthesis*; D003167:Complement Activation; D003186:Complement C9 / Q000378:metabolism; D004075:Diglycerides / Q000096:biosynthesis*; D006801:Humans; D051379:Mice; D008970:Molecular Weight; D037342:Pertussis Toxin; D010713:Phosphatidylcholines / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism; D013379:Substrate Specificity; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D002518:Ceramides; D003186:Complement C9; D004075:Diglycerides; D010713:Phosphatidylcholines; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D011493:Protein Kinase C