scholarly journals Effect of a membrane-targeted sphingosine kinase 1 on cell proliferation and survival

2005 ◽  
Vol 388 (3) ◽  
pp. 827-834 ◽  
Author(s):  
Farida SAFADI-CHAMBERLAIN ◽  
Li-Ping WANG ◽  
Shawn G. PAYNE ◽  
Chang-Uk LIM ◽  
Suzanne STRATFORD ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Plasma Membrane ◽  
Phorbol Esters ◽  
Cell Function ◽  
Sphingosine Kinase ◽  
Subcellular Location ◽  
Inhibitory Effect ◽  
Potent Inhibitory Effect ◽  
Sphingosine 1 Phosphate ◽  
Extracellular Stimuli

Numerous extracellular stimuli activate SK1 (sphingosine kinase type 1) to catalyse the production of sphingosine 1-phosphate, a bioactive lipid that functions as both an extracellular ligand for a family of G-protein-linked receptors and as a putative intracellular messenger. Phorbol esters, calcium or immunoglobulin receptors stimulate SK1 by promoting its translocation to the plasma membrane, which brings it into proximity both to its substrate (i.e. sphingosine) and to activating acidic phospholipids (e.g. phosphatidylserine). To evaluate the consequence of SK translocation, we generated an SK1-derivative tagged with a myristoylation sequence (Myr-SK1) on its N-terminus and overexpressed the construct in 3T3-L1 fibroblasts using recombinant retrovirus. Myr-SK1 overexpression increased SK activity by more than 50-fold in crude membranes, while only stimulating cytoplasmic SK activity by 4-fold. In contrast, the overexpression of WT-SK1 (wild-type SK1), as well as that of a construct containing a false myristoylation sequence (A2-Myr-SK1), markedly increased SK activity in both membrane and cytoplasmic compartments. Immunofluorescence confirmed that Myr-SK1 preferentially localized at the plasma membrane, whereas WT-SK1 and A2-Myr-SK1 partitioned in cytoplasmic/perinuclear cellular regions. Surprisingly, Myr-SK1 overexpression significantly decreased the rates of cell proliferation by delaying exit from G0/G1 phase. Moreover, expression of Myr-SK1 but not WT-SK1 or A2-Myr-SK1 protected cells from apoptosis induced by serum withdrawal. Collectively, these findings reveal that altering the subcellular location of SK1 has marked effects on cell function, with plasma membrane-associated SK having a potent inhibitory effect on the G1–S phase transition.

Autoregulation of muscarinic and gastrin receptors on gastric parietal cells

1989 ◽  
Vol 256 (2) ◽  
pp. G356-G363 ◽  
Author(s):  
T. Chiba ◽  
S. K. Fisher ◽  
B. W. Agranoff ◽  
T. Yamada
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Parietal Cell ◽  
Phorbol Esters ◽  
Cell Function ◽  
Inhibitory Effect ◽  
Parietal Cells ◽  
Kinase C ◽  
Protein Kinase C Activity ◽  
Inositol Phospholipids

In previous studies we demonstrated that parietal cell stimulation with gastrin and carbamoylcholine (carbachol) is accompanied by increased turnover of membrane inositol phospholipids. We conducted the present studies to examine whether membrane-associated protein kinase C activity is enhanced as a consequence of these events and to explore the role of this enzyme in regulating parietal cell function. We observed that carbachol and gastrin dose dependently increased membrane-associated protein kinase C activity while histamine did not. Furthermore, compounds such as phorbol esters and diacylglycerol, which are known to be direct stimulants of protein kinase C activity, also stimulated parietal cell aminopyrine uptake. In contrast, the phorbol ester 12-O-tetradecanoylphorbol-13-acetate and the synthetic diacylglycerol 1-oleoyl-2-acetyl-sn-glycerol inhibited both aminopyrine uptake and membrane inositol phospholipid turnover in parietal cells induced by carbachol and gastrin. The inhibitory effect appeared to result from reduction in the quantity of muscarinic and gastrin receptors without alterations in their specific affinities. These data suggest that protein kinase C mediates stimulation of parietal cells by gastrin and carbachol but also activates an autoregulatory mechanism via downregulation of muscarinic and gastrin receptors.

scholarly journals Sphingosine kinase 1 overexpression stimulates intestinal epithelial cell proliferation through increased c-Myc translation

2013 ◽  
Vol 304 (12) ◽  
pp. C1187-C1197 ◽  
Author(s):  
Ping Jiang ◽  
Alexis D. Smith ◽  
Ruiyun Li ◽  
Jaladanki N. Rao ◽  
Lan Liu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cellular Proliferation ◽  
Sphingosine Kinase ◽  
Intestinal Cell ◽  
Intestinal Epithelial ◽  
Sphingosine Kinase 1 ◽  
Epithelial Homeostasis ◽  
Sphingosine 1 Phosphate ◽  
Enhanced Expression ◽  
Rate Limiting

Sphingosine-1-phosphate (S1P), through mechanisms that are not completely understood, is shown to modulate cellular proliferation, which is critically important for maintaining the integrity of intestinal epithelium. Here, we show that increased S1P promotes proliferation in intestinal epithelial cells. We found that overexpression of sphingosine kinase 1 (SphK1), the rate-limiting enzyme for S1P synthesis, significantly increased cell proliferation and that this occurred through enhanced expression of c-Myc. Further, we found that the increased pattern of expression of c-Myc occurred predominantly due to its increased translation. The overexpressed SphK1 led to increased checkpoint kinase 2 and enhanced HuR phosphorylation which allowed for increased translation of c-Myc mRNA through HuR binding at the 3′-untranslated regions. Our findings demonstrate that S1P modulates intestinal cell proliferation and provides new insights as to the mechanistic actions of SphK1 and S1P in maintaining intestinal epithelial homeostasis.

scholarly journals Phospholipase C and Protein Kinase C-β 2 Mediate Insulin-Like Growth Factor II-Dependent Sphingosine Kinase 1 Activation

2011 ◽  
Vol 25 (12) ◽  
pp. 2144-2156 ◽  
Author(s):  
Hesham M. El-Shewy ◽  
Souzan A. Abdel-Samie ◽  
Abdelmohsen M. Al Qalam ◽  
Mi-Hye Lee ◽  
Kazuyuki Kitatani ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Fluorescent Protein ◽  
Sphingosine Kinase ◽  
Sphingosine Kinase 1 ◽  
Membrane Translocation ◽  
Kinase C ◽  
Sphingosine 1 Phosphate

Abstract We recently reported that IGF-II binding to the IGF-II/mannose-6-phosphate (M6P) receptor activates the ERK1/2 cascade by triggering sphingosine kinase 1 (SK1)-dependent transactivation of G protein-coupled sphingosine 1-phosphate (S1P) receptors. Here, we investigated the mechanism of IGF-II/M6P receptor-dependent sphingosine kinase 1 (SK1) activation in human embryonic kidney 293 cells. Pretreating cells with protein kinase C (PKC) inhibitor, bisindolylmaleimide-I, abolished IGF-II-stimulated translocation of green fluorescent protein (GFP)-tagged SK1 to the plasma membrane and activation of endogenous SK1, implicating PKC as an upstream regulator of SK1. Using confocal microscopy to examine membrane translocation of GFP-tagged PKCα, β1, β2, δ, and ζ, we found that IGF-II induced rapid, transient, and isoform-specific translocation of GFP-PKCβ2 to the plasma membrane. Immunoblotting of endogenous PKC phosphorylation confirmed PKCβ2 activation in response to IGF-II. Similarly, IGF-II stimulation caused persistent membrane translocation of the kinase-deficient GFP-PKCβ2 (K371R) mutant, which does not dissociate from the membrane after translocation. IGF-II stimulation increased diacylglycerol (DAG) levels, the established activator of classical PKC. Interestingly, the polyunsaturated fraction of DAG was increased, indicating involvement of phosphatidyl inositol/phospholipase C (PLC). Pretreating cells with the PLC inhibitor, U73122, attenuated IGF-II-dependent DAG production and PKCβ2 phosphorylation, blocked membrane translocation of the kinase-deficient GFP-PKCβ2 (K371R) mutant, and reduced sphingosine 1-phosphate production, suggesting that PLC/PKCβ2 are upstream regulators of SK1 in the pathway. Taken together, these data provide evidence that activation of PLC and PKCβ2 by the IGF-II/M6P receptor are required for the activation of SK1.

scholarly journals Sphingosine and Sphingosine Kinase 1 Involvement in Endocytic Membrane Trafficking

2017 ◽  
Vol 292 (8) ◽  
pp. 3074-3088 ◽  
Author(s):  
Santiago Lima ◽  
Sheldon Milstien ◽  
Sarah Spiegel
Keyword(s):  
Plasma Membrane ◽  
Membrane Trafficking ◽  
Sphingosine Kinase ◽  
Cell Types ◽  
Membrane Cholesterol ◽  
Vesicle Formation ◽  
Sphingosine Kinase 1 ◽  
Late Endosomes ◽  
Sphingosine 1 Phosphate ◽  
Plasma Membrane Cholesterol

The balance between cholesterol and sphingolipids within the plasma membrane has long been implicated in endocytic membrane trafficking. However, in contrast to cholesterol functions, little is still known about the roles of sphingolipids and their metabolites. Perturbing the cholesterol/sphingomyelin balance was shown to induce narrow tubular plasma membrane invaginations enriched with sphingosine kinase 1 (SphK1), the enzyme that converts the bioactive sphingolipid metabolite sphingosine to sphingosine-1-phosphate, and suggested a role for sphingosine phosphorylation in endocytic membrane trafficking. Here we show that sphingosine and sphingosine-like SphK1 inhibitors induced rapid and massive formation of vesicles in diverse cell types that accumulated as dilated late endosomes. However, much smaller vesicles were formed in SphK1-deficient cells. Moreover, inhibition or deletion of SphK1 prolonged the lifetime of sphingosine-induced vesicles. Perturbing the plasma membrane cholesterol/sphingomyelin balance abrogated vesicle formation. This massive endosomal influx was accompanied by dramatic recruitment of the intracellular SphK1 and Bin/Amphiphysin/Rvs domain-containing proteins endophilin-A2 and endophilin-B1 to enlarged endosomes and formation of highly dynamic filamentous networks containing endophilin-B1 and SphK1. Together, our results highlight the importance of sphingosine and its conversion to sphingosine-1-phosphate by SphK1 in endocytic membrane trafficking.

scholarly journals Synthesis and Biological Evaluation of BODIPY-PF-543

Molecules ◽  
2019 ◽  
Vol 24 (23) ◽  
pp. 4408
Author(s):  
Shrestha ◽  
Hwang ◽  
Lee ◽  
Kim ◽  
Oh ◽  
...  
Keyword(s):  
Anticancer Activity ◽  
Cancer Cells ◽  
Intracellular Localization ◽  
Sphingosine Kinase ◽  
Inhibitory Effect ◽  
Biological Evaluation ◽  
Boron Dipyrromethene ◽  
Sphingosine 1 Phosphate ◽  
Synthesis And Biological Evaluation

Sphingosine-1-phosphate (S1P) regulates the proliferation of various cells and promotes the growth of cancer cells. Sphingosine kinase (SK), which transforms sphingosine into S1P, has two isotypes: SK1 and SK2. To date, both isotypes are known to be involved in the proliferation of cancer cells. PF-543, an SK1 inhibitor developed by Pfizer, strongly inhibits SK1. However, despite its strong SK1 inhibitory effect, PF-543 shows low anticancer activity in vitro. Therefore, additional biological evidence on the anticancer activity of SK1 inhibitor is required. The present study aimed to investigate the intracellular localization of PF-543 and identify its association with anticancer activity by introducing a fluoroprobe into PF-543. Boron–dipyrromethene (BODIPY)-introduced PF-543 has a similar SK1 inhibitory effect as PF-543. These results indicate that the introduction of BODIPY does not significantly affect the inhibitory effect of SK1. In confocal microscopy after BODIPY-PF-543 treatment, the compound was mainly located in the cytosol of the cells. This study demonstrated the possibility of introducing fluorescent material into an SK inhibitor and designing a synthesized compound that is permeable to cells while maintaining the SK inhibitory effect.

Sphingosine 1-phosphate is a novel inhibitor of T-cell proliferation

Blood ◽  
2003 ◽  
Vol 101 (12) ◽  
pp. 4909-4915 ◽  
Author(s):  
Yixin Jin ◽  
Eirunn Knudsen ◽  
Ling Wang ◽  
Yenan Bryceson ◽  
Bassam Damaj ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
Dendritic Cells ◽  
T Cell ◽  
Interleukin 2 ◽  
Inhibitory Effect ◽  
Interferon Γ ◽  
T Cell Proliferation ◽  
Inhibitory Molecule ◽  
Sphingosine 1 Phosphate

Abstract Sphingosine 1-phosphate (S1P) is a pleiotropic lysosphingophospholipid stored and secreted by platelets. Using reverse transcription–polymerase chain reaction and flow cytometric analyses, we determined the expression of S1P receptors (S1P1, S1P3, S1P4, and S1P5) in peripheral blood T cells. T cells were induced to proliferate in the presence of phorbol 12-myristate 13-acetate (PMA) plus ionomycin, anti-CD3 plus anti-CD28, and allogeneic immature or mature dendritic cells. This activity was inhibited by the addition of S1P. Enhanced T-cell proliferation was observed when these cells were stimulated with the same stimuli, but were incubated in serum-free media (SFM). Addition of S1P to SFM inhibited the stimulation of T cells induced by T-cell stimuli, suggesting that S1P is an important inhibitory molecule present in the serum. T-cell proliferation was also inhibited by the addition of dihydrosphingosine 1-phosphate (DHS1P), sphingosine, and ceramide; however, the latter 2 sphingolipids required higher concentrations than S1P. Pretreatment of T cells with pertussis toxin (PTX) blocked the inhibitory effect of S1P on activation with PMA plus ionomycin, but not on activation with anti-CD3 plus anti-CD28. This is corroborated with the down-regulation of S1P1 in T cells stimulated with anti-CD3 plus anti-CD28. Similarly, PTX did not affect the inhibitory effect of S1P on T-cell proliferation when dendritic cells were used as stimuli. Further, S1P or DHS1P but not ceramide or sphingosine enhanced rather than decreased secretion of interleukin 2 and interferon γ by T cells stimulated with anti-CD3 plus anti-CD28. These results show differential effects of S1P on polyclonal T-cell proliferation and cytokine secretion.

scholarly journals Spatial diversity: Ras trafficking and signalling

2003 ◽  
Vol 25 (3) ◽  
pp. 22-24 ◽  
Author(s):  
Ian A. Prior
Keyword(s):  
Cell Proliferation ◽  
Plasma Membrane ◽  
G Proteins ◽  
Spatial Diversity ◽  
Ras Proteins ◽  
Cell Proliferation And Differentiation ◽  
Proliferation And Differentiation ◽  
Extracellular Stimuli ◽  
New Research ◽  
Ras Isoforms

Ras proteins are small monomeric G-proteins that play a central role in linking extracellular stimuli with the internal kinase cascades that are primarily involved in modulating cell proliferation and differentiation. Constitutively activated mutants of these proteins are found in many human cancers. Differences in the trafficking and plasma membrane localization of Ras isoforms play critical roles in regulating their function. New research is trying to understand the mechanisms behind, and consequences of, these differences.

Inhalation of sphingosine kinase inhibitor attenuates airway inflammation in asthmatic mouse model

2008 ◽  
Vol 294 (6) ◽  
pp. L1085-L1093 ◽  
Author(s):  
Teruaki Nishiuma ◽  
Yoshihiro Nishimura ◽  
Taro Okada ◽  
Emi Kuramoto ◽  
Yoshikazu Kotani ◽  
...  
Keyword(s):  
Mouse Model ◽  
Bronchial Asthma ◽  
Kinase Inhibitor ◽  
Mrna Level ◽  
Bronchial Hyperresponsiveness ◽  
Sphingosine Kinase ◽  
Inhibitory Effect ◽  
Eosinophil Infiltration ◽  
Cell Hyperplasia ◽  
Sphingosine 1 Phosphate

Sphingosine 1-phosphate (S1P) produced by sphingosine kinase (SPHK) is implicated in acute immunoresponses, however, mechanisms of SPHK/S1P signaling in the pathogenesis of bronchial asthma are poorly understood. In this study, we hypothesized that SPHK inhibition could ameliorate lung inflammation in ovalbumin (OVA)-challenged mouse lungs. Six- to eight-week-old C57BL/6J mice were sensitized and exposed to OVA for 3 consecutive days. Twenty-four hours later, mice lungs and bronchoalveolar lavage (BAL) fluid were analyzed. For an inhibitory effect, either of the two different SPHK inhibitors, N, N-dimethylsphingosine (DMS) or SPHK inhibitor [SK-I; 2-( p-hydroxyanilino)-4-( p-chlorophenyl) thiazole], was nebulized for 30 min before OVA inhalation. OVA inhalation caused S1P release into BAL fluid and high expression of SPHK1 around bronchial epithelial walls and inflammatory areas. DMS or SK-I inhalation resulted in a decrease in S1P amounts in BAL fluid to basal levels, accompanied by decreased eosinophil infiltration and peroxidase activity. The extent of inhibition caused by DMS inhalation was higher than that caused by SK-I. Like T helper 2 (Th2) cytokine release, OVA inhalation-induced increase in eotaxin expression was significantly suppressed by DMS pretreatment both at protein level in BAL fluid and at mRNA level in lung homogenates. Moreover, bronchial hyperresponsiveness to inhaled methacholine and goblet cell hyperplasia were improved by SPHK inhibitors. These data suggest that the inhibition of SPHK affected acute eosinophilic inflammation induced in antigen-challenged mouse model and that targeting SPHK may provide a novel therapeutic tool to treat bronchial asthma.

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