Regulatory Effect of Phorbol Esters on Sphingosine Kinase in BALB/C 3T3 Fibroblasts (Variant A31): Demonstration of Cell Type-Specific Response - A Preliminary Note

Mazurek, Nachman; Megidish, Tamar; Hakomori, S; Igarashi, Yasuyuki

doi:10.1006/bbrc.1994.1001

Cited by 86 publications

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“…It is known that S1P levels in the cell are largely mediated by its formation from sphingosine by the activity of SK, and to a lesser extent by its degradation by endoplasmic reticulum-associated S1P lyase [13,14] and S1P phosphatase [2]. Basal levels of S1P in the cell are generally low [2,15], but can increase rapidly and transiently when cells are exposed to various agonists, including TNFα [4,10], plateletderived growth factor [15], nerve growth factor [16] and phorbol esters [17,18]. This response appears to be invariably correlated with an increase in SK activity in the cytosol and can be prevented by addition of SK inhibitors.…”

Section: Introductionmentioning

confidence: 99%

Human sphingosine kinase: purification, molecular cloning and characterization of the native and recombinant enzymes

Pitson

D’Andrea

Vandeleur

et al. 2000

Biochemical Journal

140

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Sphingosine 1-phosphate (S1P) is a novel lipid messenger that has important roles in a wide variety of mammalian cellular processes including growth, differentiation and death. Basal levels of S1P in mammalian cells are generally low, but can increase rapidly and transiently when cells are exposed to mitogenic agents and other stimuli. This increase is largely due to increased activity of sphingosine kinase (SK), the enzyme that catalyses its formation. In the current study we have purified, cloned and characterized the first human SK to obtain a better understanding of its biochemical activity and possible activation mechanisms. The enzyme was purified to homogeneity from human placenta using ammonium sulphate precipitation, anion-exchange chromatography, calmodulin-affinity chromatography and gel-filtration chromatography. This resulted in a purification of over 106-fold from the original placenta extract. The enzyme was cloned and expressed in active form in both HEK-293T cells and Escherichia coli, and the recombinant E. coli-derived SK purified to homogeneity. To establish whether post-translational modifications lead to activation of human SK activity we characterized both the purified placental enzyme and the purified recombinant SK produced in E. coli, where such modifications would not occur. The premise for this study was that post-translational modifications are likely to cause conformational changes in the structure of SK, which may result in detectable changes in the physico-chemical or catalytic properties of the enzyme. Thus the enzymes were characterized with respect to substrate specificity and kinetics, inhibition kinetics and various other physico-chemical properties. In all cases, both the native and recombinant SKs displayed remarkably similar properties, indicating that post-translational modifications are not required for basal activity of human SK.

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Section: Introductionmentioning

confidence: 99%

Human sphingosine kinase: purification, molecular cloning and characterization of the native and recombinant enzymes

Pitson

D’Andrea

Vandeleur

et al. 2000

Biochemical Journal

140

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“…Olivera and Spiegel claimed that Sph-l-P is a novel second messenger in platelet-derived growth factor-and serum-dependent cell growth [10]. Sph kinase activity is highly stimulated by phorbol esters in BALB/c 3T3 (A31) cells but not in Swiss 3T3 cells [11]. Here we show that the motility-inhibitory effect of Sph-l-P is based on inhibition of at/in nucleation, polymerization, and reorganization of pseudopodium formatior, *Corresponding author.…”

Section: Introductionmentioning

confidence: 99%

Sphingosine‐1‐phosphate inhibits actin nucleation and pseudopodium formation to control cell motility of mouse melanoma cells

Yamamura¹,

Sadahira²,

Ruan

et al. 1996

FEBS Letters

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Sphingosine-l-phosphate (Sph-l-P), the initial product of.~phingosine (Sph) catabolism, has been reported to inhibit motility of mouse melanoma B16/F1 and other types of cells at very low coneemraClens (10-]00 nM). Sph-l-P (100 nM-I pM) inhibited pseudopodlum formation by bloekJnl~ ~olymerization and reorganization of actin filaments in newl~t formed pseudopodia, and reduced F-aetin by ,~2.~% in FI cells, A pyrenelabeled aetin nucleation assay revealed that Sph-I-P (100 aM) inhibits actln nucleation mediated by t'~l cell plasma membranes. These results suggest that Sph-I-P int~vacts with molecules associated with aetin nucleation to inhiblt reorg.~r,!z.~t!an of pseudopodlum formation and cell motility.

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“…Sphingosine kinases are activated by stimuli such as treatment with platelet-derived growth factor (18), tumor necrosis factor ␣ (19), or phorbol ester (20) as well as the cross-linking of Fc␥R1 (21) or Fc⑀R1 (22). Additionally, SPHK1 is known to be regulated by protein-protein interactions (23)(24)(25)(26)(27), phosphorylation (28,29), and translocation to the plasma membrane (28,29).…”

mentioning

confidence: 99%

Mouse Sphingosine Kinase Isoforms SPHK1a and SPHK1b Differ in Enzymatic Traits Including Stability, Localization, Modification, and Oligomerization

Kihara

Anada

Igarashi

2006

Journal of Biological Chemistry

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Sphingosine kinases catalyze the production of the bioactive lipid molecule sphingosine 1-phosphate. Mice have two isoforms of sphingosine kinase type 1, SPHK1a and SPHK1b. In addition to the previously reported difference in their enzyme activities, we have found that these isoforms differ in several enzymatic characteristics. First, SPHK1b is unstable, whereas SPHK1a is highly stable. Degradation of SPHK1b occurs at the membrane and is inhibited by a proteasome inhibitor. Second, only SPHK1b exhibits abnormal mobility on SDS-PAGE, probably due to its SDS-resistant structure. Third, SPHK1a and SPHK1b are predominantly detected in the soluble and membrane fractions, respectively, when their degradation is inhibited. Fourth, only SPHK1b is modified with lipid, on its unique Cys residues (Cys-4 and Cys-5). Site-directed mutagenesis at these Cys residues resulted in increased sphingosine kinase activity, suggesting that the modification is inhibitory to the enzyme. Finally, SPHK1b tends to form homo-oligomers, whereas most SPHK1a is presented as monomers. We have also determined that the lipid modification of SPHK1b is involved in its homo-oligomerization. Thus, although these two proteins differ only in a few N-terminal amino acid residues, their enzymatic traits are extremely different.

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Regulatory Effect of Phorbol Esters on Sphingosine Kinase in BALB/C 3T3 Fibroblasts (Variant A31): Demonstration of Cell Type-Specific Response - A Preliminary Note

Cited by 86 publications

References 0 publications

Human sphingosine kinase: purification, molecular cloning and characterization of the native and recombinant enzymes

Human sphingosine kinase: purification, molecular cloning and characterization of the native and recombinant enzymes

Sphingosine‐1‐phosphate inhibits actin nucleation and pseudopodium formation to control cell motility of mouse melanoma cells

Mouse Sphingosine Kinase Isoforms SPHK1a and SPHK1b Differ in Enzymatic Traits Including Stability, Localization, Modification, and Oligomerization

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