Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

This laboratory first provided evidence for a potential signal transduction pathway involving sphingomyelin and its derivatives (Kolesnick, R.N., and Clegg, S. (1988) J. Biol. Chem. 263, 6534-6537). Recently, this laboratory demonstrated the existence of the novel sphingolipid ceramide 1-phosphate in human leukemia (HL-60) cells. Ceramide 1-phosphate was synthesized from ceramide derived from sphingomyelin but not glycosphingolipids. This suggested that a specific pathway extended from sphingomyelin to ceramide 1-phosphate. The present studies provide additional support for this notion by demonstrating the existence of a ceramide kinase activity distinct from diacylglycerol (DG) kinase in HL-60 cells. Microsomal membranes contained a kinase activity that phosphorylated ceramide but not 1,2-DG in the presence of physiologic and higher Ca2+ concentrations (60 nM-3 mM). Kinetic analyses demonstrated an apparent Vmax for ceramide and ATP of 70 pmol.min-1.mg protein-1; apparent Km values were 45 and 25 microM, respectively. The pH optimum was within the physiologic range (pH 6-8). Magnesium but not other divalent cations (Mn2+, Ba2+, Cd2+, Zn2+) also stimulated ceramide phosphorylation. Magnesium also induced 1,2-DG phosphorylation. Since DG kinase is a Mg2(+)-stimulable enzyme that may utilize ceramide as substrate, additional studies separated calcium-dependent ceramide kinase from DG kinase activity. 1,2-DGs competitively inhibited magnesium- but not calcium-dependent ceramide phosphorylation. Hence, calcium-dependent ceramide kinase activity neither utilized DG as substrate nor was inhibited by DG. These activities were physically separable. Both activities were solubilized by n-octyl-beta-D-glucopyranoside and stabilized by glycerol. Ceramide kinase activity bound weakly to a DEAE-cellulose anion exchange column and eluted with 4-fold purification as a single peak of activity in the flow-through and 0.05 M NaCl elutions. In contrast, the majority of DG kinase activity bound more tightly and was recovered as a broad peak in the 0.2-0.35 M NaCl elutions. These studies demonstrate the existence of a ceramide kinase activity in HL-60 cells which is functionally and physically separable from DG kinase. These studies provide further support for the notion of a specific pathway from sphingomyelin to ceramide 1-phosphate.
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PMID:Characterization of a ceramide kinase activity from human leukemia (HL-60) cells. Separation from diacylglycerol kinase activity. 217 34

Ceramide (Cer) is the precursor for sphingolipids and functions as a second messenger in a variety of cellular processes including apoptosis. However, no direct target of Cer leading to apoptosis has been identified. Understanding the movement and trafficking of Cer is important for fully understanding Cer signaling. In this study, we identified, for the first time, the transbilayer movement of Cer in the plasma membrane (PM) of living cells. We developed a new method to monitor transbilayer Cer movement using ceramide kinase activity. To produce Cer on the extracellular leaflet of the PM, bacterial sphingomyelinase (SMase) was added to rat basophilic leukemia cells. Interestingly, the dramatic elevation of ceramide 1-phosphate (C1P), the product of CerK, was observed following the increase of Cer induced by SMase treatment. Since we determined that both the protein and catalytic activity of CerK exists in the intracellular compartment, the all conversion of Cer to C1P by CerK should be occur intracellularly. This result indicates the rapid transbilayer movement of Cer from the outer leaflet to the inner leaflet of the PM of living cells. Furthermore, protease digestion of membrane proteins, inhibition of ABC transporters (by glibencramide) and of cation channels (by carbonyl cyanide m-chlorophenylhydrozone), and modification of cholesterol content did not affect the transbilayer movement of Cer. Thus, this movement might occur spontaneously. Our findings indicate not only Cer movement in the PM, but also identify an intrinsic property of Cer enabling Cer signaling.
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PMID:Transbilayer movement of ceramide in the plasma membrane of live cells. 1755 61