Opposite effects of dihydrosphingosine 1-phosphate and sphingosine 1-phosphate on transforming growth factor-beta/Smad signaling are mediated through the PTEN/PPM1A-dependent pathway

Transforming growth factor-beta (TGF-beta) is an important regulator of physiological connective tissue biosynthesis and plays a central role in pathological tissue fibrosis. Previous studies have established that a biologically active lipid mediator, sphingosine 1-phosphate (S1P), mimics some of the profibrotic functions of TGF-beta through cross-activation of Smad signaling. Here we report that another product of sphingosine kinase, dihydrosphingosine 1-phosphate (dhS1P), has an opposite role in the regulation of TGF-beta signaling. In contrast to S1P, dhS1P inhibits TGF-beta-induced Smad2/3 phosphorylation and up-regulation of collagen synthesis. The effects of dhS1P require a lipid phosphatase, PTEN, a key modulator of cell growth and survival. dhS1P stimulates phosphorylation of the C-terminal domain of PTEN and its subsequent translocation into the nucleus. We demonstrate a novel function of nuclear PTEN as a co-factor of the Smad2/3 phosphatase, PPM1A. Complex formation of PTEN with PPM1A does not require the lipid phosphatase activity but depends on phosphorylation of the serine/threonine residues located in the C-terminal domain of PTEN. Upon complex formation with PTEN, PPM1A is protected from degradation induced by the TGF-beta signaling. Consequently, overexpression of PTEN abrogates TGF-beta-induced Smad2/3 phosphorylation. This study establishes a novel role for nuclear PTEN in the stabilization of PPM1A. PTEN-mediated cross-talk between the sphingolipid and TGF-beta signaling pathways may play an important role in physiological and pathological TGF-beta signaling.     

Sphingolipid metabolism is a crucial determinant of cellular fate in nonstimulated proliferating Madin-Darby canine kidney (MDCK) cells

The present report was addressed to study the influence of sphingolipid metabolism in determining cellular fate. In nonstimulated proliferating Madin-Darby canine kidney (MDCK) cells, sphingolipid de novo synthesis is branched mainly to a production of sphingomyelin and ceramide, with a minor production of sphingosylphosphocholine, ceramide 1-phosphate, and sphingosine 1-phosphate. Experiments with (32)P as a radioactive precursor showed that sphingosine 1-phosphate is produced mainly by a de novo independent pathway. Enzymatic inhibition of the de novo pathway and ceramide synthesis affected cell number and viability only slightly, without changing sphingosine 1-phosphate production. By contrast, inhibition of sphingosine kinase-1 activity provoked a significant reduction in both cell number and viability in a dose-dependent manner. When sphingolipid metabolism was studied, an increase in de novo formed ceramide was found, which correlated with the concentration of enzyme inhibitor and the reduction in cell number and viability. Knockdown of sphingosine kinase-1 expression also induced an accumulation of de novo synthesized ceramide, provoking a slight reduction in cell number and viability similar to that induced by a low concentration of the sphingosine kinase inhibitor. Taken together, our results indicate that the level of de novo formed ceramide is controlled by the synthesis of sphingosine 1-phosphate, which appears to occur through a de novo synthesis-independent pathway, most probably the salvage pathway, that is responsible for the MDCK cell fate, suggesting that under proliferating conditions, a dynamic interplay exists between the de novo synthesis and the salvage pathway.     

Does cytokine signaling link sphingolipid metabolism to host defense and immunity against virus infections?

Sphingosine 1-phosphate (S1P)-metabolizing enzymes regulate the level of bioactive sphingolipids that have curative potential. Recently, S1P-metabolizing enzymes such as sphingosine kinase 1 and S1P lyase were shown to regulate influenza virus replication and the virus-induced cytopathogenicity. The mechanism appeared to employ a JAK/STAT type I interferon signaling pathway that induces anti-viral status. Further, sphingosine analogs altered cytokine responses upon influenza virus infection. This article focuses on recent discoveries about the sphingolipid system that influences on host protection from viral virulence and the involvement of cytokine signaling in its underlying mechanisms. Deciphering the steps of this pathway could help us envision how the modulation of sphingolipid metabolism can be applied as a therapeutic approach to overcome infectious diseases.     

De novo biosynthesis of dihydrosphingosine-1-phosphate by sphingosine kinase 1 in mammalian cells

Sphingosine kinase 1 (SK1) is one of the two known kinases, which generates sphingosine-1-phosphate (S1P), a potent endogenous lipid mediator involved in cell survival, proliferation, and cell-cell interactions. Activation of SK1 and intracellular generation of S1P were suggested to be part of the growth and survival factor-induced signaling, and overexpression of SK1 provoked cell tumorigenic transformation. Using a highly selective and sensitive LC-MS/MS approach, here we show that SK1 overexpression, but not SK2, in different primary cells and cultured cell lines results in predominant upregulation of the synthesis of dihydrosphingosine-1-phosphate (DHS1P) compared to S1P. Stable isotope pulse-labeling experiments in conjunction with LC-MS/MS quantitation of different sphingolipids demonstrated strong interference of overexpressed SK1 with the de novo sphingolipid biosynthesis by deviating metabolic flow of newly formed sphingoid bases from ceramide formation toward the synthesis of DHS1P. On the contrary, S1P biosynthesis was not directly linked to the de novo sphingoid bases transformations and was dependent on catabolic generation of sphingosine from complex sphingolipids. As a result of SK1 overexpression, migration and Ca2+-response of human pulmonary artery endothelial cells (HPAEC) to stimulation with external S1P, but not thrombin, was strongly impaired. In contrast, selective increase in intracellular content of DHS1P or S1P through the uptake and phosphorylation of corresponding sphingoid bases had no effect on S1P-induced signaling or facilitation of wound healing. Furthermore, infection of human bronchial epithelial cells (HBEpC) with RSV A-2 virus increased SK1-mediated synthesis of DHS1P and S1P, whereas TNF-alpha enhanced only S1P production in HPAEC. These findings uncover a new functional role for SK1, which can control survival/death (DHS1P-S1P/ceramides) balance by targeting sphingolipid de novo biosynthesis and selectively generating DHS1P at a metabolic step preceding ceramide formation.     

Sphingosine kinase localization in the control of sphingolipid metabolism

The sphingosine kinases (sphingosine kinase-1 and -2) have been implicated in a variety of physiological functions. Discerning their mechanism of action is complicated because in addition to producing the potent lipid second messenger sphingosine-1-phosphate, sphingosine kinases, both by producing sphingosine-1-phosphate and consuming sphingosine, have profound effects on sphingolipid metabolism. Sphingosine kinase-1 translocates to the plasma membrane upon agonist stimulation and this translocation is essential for the pro-oncogenic properties of this enzyme. Many of the enzymes of sphingolipid metabolism, including the enzymes that degrade sphingosine-1-phosphate, are membrane bound with restricted subcellular distributions. In the work described here we explore how subcellular localization of sphingosine kinase-1 affects the downstream metabolism of sphingosine-1-phosphate and the access of sphingosine kinase to its substrates. We find, surprisingly, that restricting sphingosine kinase to either the plasma membrane or the endoplasmic reticulum has a negligible effect on the rate of degradation of the sphingosine-1-phosphate that is produced. This suggests that sphingosine-1-phosphate is rapidly transported between membranes. However we also find that cytosolic or endoplasmic-reticulum targeted sphingosine kinase expressed at elevated levels produces extremely high levels of dihydrosphingosine-1-phosphate. Dihydrosphingosine is a proximal precursor in ceramide biosynthesis. Our data indicate that sphingosine kinase can divert substrate from the ceramide de novo synthesis pathway. However plasma membrane-restricted sphingosine kinase cannot access the pool of dihydrosphingosine. Therefore whereas sphingosine kinase localization does not affect downstream metabolism of sphingosine-1-phosphate, localization has an important effect on the pools of substrate to which this key signaling enzyme has access.     

Palmitate increases sphingosine-1-phosphate in C2C12 myotubes via upregulation of sphingosine kinase message and activity

Studies in skeletal muscle demonstrate that elevation of plasma FFAs increases the sphingolipid ceramide. We aimed to determine the impact of FFA oversupply on total sphingolipid profiles in a skeletal muscle model. C2C12 myotubes were treated with palmitate (PAL). Lipidomics analysis revealed pleiotropic effects of PAL on cell sphingolipids not limited to ceramides. ¹³C labeling demonstrated that PAL activated several branches of sphingolipid synthesis by distinct mechanisms. Intriguingly, PAL increased sphingosine-1-phosphate independently of de novo synthesis. Quantitative real-time PCR demonstrated that PAL increased sphingosine kinase 1 (SK1) mRNA by approximately 4-fold. This was accompanied by a 2.3-fold increase in sphingosine kinase enzyme activity. This upregulation did not occur upon treatment with oleate, suggesting some level of specificity for PAL. These findings were recapitulated in the diet-induced obesity mouse model, in which high-fat feeding increased SK1 message in skeletal muscle over 2.3-fold. These data suggest that the impact of elevated FFA on sphingolipids reaches beyond ceramides and de novo sphingolipid synthesis. Moreover, these findings identify PAL as a novel regulatory stimulus for SK1.     

Role of sphingosine kinase localization in sphingolipid signaling

The sphingosine kinases, SK1 and SK2, produce the potent signaling lipid sphingosine-1-phosphate (S1P). These enzymes have garnered increasing interest for their roles in tumorigenesis, inflammation, vascular diseases, and immunity, as well as other functions. The sphingosine kinases are considered signaling enzymes by producing S1P, and their activity is acutely regulated by a variety of agonists. However, these enzymes are also key players in the control of sphingolipid metabolism. A variety of sphingolipids, such as sphingosine and the ceramides, are potent signaling molecules in their own right. The role of sphingosine kinases in regulating sphingolipid metabolism is potentially a critical aspect of their signaling function. A central aspect of signaling lipids is that their hydrophobic nature constrains them to membranes. Most enzymes of sphingolipid metabolism, including the enzymes that degrade S1P, are membrane enzymes. Therefore the localization of the sphingosine kinases and S1P is likely to be important in S1P signaling. Sphingosine kinase localization affects sphingolipid signaling in several ways. Translocation of SK1 to the plasma membrane promotes extracellular secretion of S1P. SK1 and SK2 localization to specific sites appears to direct S1P to intracellular protein effectors. SK localization also determines the access of these enzymes to their substrates. This may be an important mechanism for the regulation of ceramide biosynthesis by diverting dihydrosphingosine, a precursor in the ceramide biosynthetic pathway, from the de novo production of ceramide.     

Sphingosine 1-phosphate (S1P) lyase deficiency increases sphingolipid formation via recycling at the expense of de novo biosynthesis in neurons

Sphingosine 1-phosphate lyase (S1P lyase) irreversibly cleaves sphingosine 1-phosphate (S1P) in the final step of sphingolipid catabolism. As sphingoid bases and their 1-phosphate are not only metabolic intermediates but also highly bioactive lipids that modulate a wide range of physiological processes, it would be predicted that their elevation might induce adjustments in other facets of sphingolipid metabolism and/or alter cell behavior. Indeed, we have previously reported that S1P lyase deficiency causes neurodegeneration and other adverse symptoms. We next asked the question whether and how S1P lyase deficiency affects the metabolism of (glyco)sphingolipids and cholesterol, two lipid classes that might be involved in the neurodegenerative processes observed in S1P lyase-deficient mice. As predicted, there was a considerable increase in free and phosphorylated sphingoid bases upon elimination of S1P lyase, but to our surprise, rather than increasing, the mass of (glyco)sphingolipids persisted at wild type levels. This was discovered to be due to reduced de novo sphingoid base biosynthesis and a corresponding increase in the recycling of the backbones via the salvage pathway. There was also a considerable increase in cholesterol esters, although free cholesterol persisted at wild type levels, which might be secondary to the shifts in sphingolipid metabolism. All in all, these findings show that accumulation of free and phosphorylated sphingoid bases by loss of S1P lyase causes an interesting readjustment of the balance between de novo biosynthesis and recycling to maintain (glyco)sphingolipid homeostasis. These changes, and their impact on the metabolism of other cellular lipids, should be explored as possible contributors to the neurodegeneration in S1P lyase deficiency.     

The control of the balance between ceramide and sphingosine-1-phosphate by sphingosine kinase: oxidative stress and the seesaw of cell survival and death

Sphingolipids are components of all eukaryotic cells that play important roles in a wide variety of biological processes. Ceramides and sphingosine-1-phosphate (S1P) are signaling molecules that regulate cell fate decisions in a wide array of species including yeast, plants, vertebrates, and invertebrates. Ceramides favor anti-proliferative and cell death pathways such as senescence and apoptosis, whereas S1P stimulates cell proliferation and survival pathways. The control of cell fate by these two interconvertible lipids has been called the sphingolipid rheostat or sphingolipid biostat. Sphingosine kinase, the enzyme that synthesizes S1P, is a crucial enzyme in regulation of the balance of these sphingolipids. Sphingosine kinase has been shown to play dynamic roles in the responses of cells to stress, leading to modulation of cell fate through a variety of signaling pathways impinging on the processes of cell proliferation, apoptosis, autophagy and senescence. This review summarizes the roles of sphingosine kinase signaling in these processes and the mechanisms mediating these responses. In addition, we discuss the evidence tying sphingosine kinase-mediated stress responses to the process of aging.     

TORC1-regulated protein kinase Npr1 phosphorylates Orm to stimulate complex sphingolipid synthesis

The evolutionarily conserved Orm1 and Orm2 proteins mediate sphingolipid homeostasis. However, the homologous Orm proteins and the signaling pathways modulating their phosphorylation and function are incompletely characterized. Here we demonstrate that inhibition of nutrient-sensitive target of rapamycin complex 1 (TORC1) stimulates Orm phosphorylation and synthesis of complex sphingolipids in Saccharomyces cerevisiae. TORC1 inhibition activates the kinase Npr1 that directly phosphorylates and activates the Orm proteins. Npr1-phosphorylated Orm1 and Orm2 stimulate de novo synthesis of complex sphingolipids downstream of serine palmitoyltransferase. Complex sphingolipids in turn stimulate plasma membrane localization and activity of the nutrient scavenging general amino acid permease 1. Thus activation of Orm and complex sphingolipid synthesis upon TORC1 inhibition is a physiological response to starvation.     

Identification of two lipid phosphatases that regulate sphingosine-1-phosphate cellular uptake and recycling

Sphingosine-1-phosphate (S1P) is a sphingolipid metabolite that serves as a potent extracellular signaling molecule. Metabolic regulation of extracellular S1P levels impacts key cellular activities through altered S1P receptor signaling. Although the pathway through which S1P is degraded within the cell and thereby eliminated from reuse has been previously described, the mechanism used for S1P cellular uptake and the subsequent recycling of its sphingoid base into the sphingolipid synthesis pathway is not completely understood. To identify the genes within this S1P uptake and recycling pathway, we performed a genome-wide CRISPR/Cas9 KO screen using a positive-selection scheme with Shiga toxin, which binds a cell-surface glycosphingolipid receptor, globotriaosylceramide (Gb3), and causes lethality upon internalization. The screen was performed in HeLa cells with their sphingolipid de novo pathway disabled so that Gb3 cell-surface expression was dependent on salvage of the sphingoid base of S1P taken up from the medium. The screen identified a suite of genes necessary for S1P uptake and the recycling of its sphingoid base to synthesize Gb3, including two lipid phosphatases, PLPP3 (phospholipid phosphatase 3) and SGPP1 (S1P phosphatase 1). The results delineate a pathway in which plasma membrane–bound PLPP3 dephosphorylates extracellular S1P to sphingosine, which then enters cells and is rephosphorylated to S1P by the sphingosine kinases. This rephosphorylation step is important to regenerate intracellular S1P as a branch-point substrate that can be routed either for dephosphorylation to salvage sphingosine for recycling into complex sphingolipid synthesis or for degradation to remove it from the sphingolipid synthesis pathway.     

Comparative quantification of sphingolipids and analogs in biological samples by high-performance liquid chromatography after chloroform extraction

Sphingosine 1-phosphate (S1P) is an extra- and intracellular messenger that specifically activates five G-protein-coupled cell surface receptors designated S1P_1-5. The S1P₁ receptor is particularly important for the maintenance of immune surveillance by regulating egress of lymphocytes from thymus and secondary lymphoid organs. S1P is generated through phosphorylation of sphingosine which is catalyzed by sphingosine kinase types 1 and 2. The immunosuppressant and sphingosine analog Fingolimod (2-amino-2-(2-[4-octylphenyl]ethyl)-1,3-propanediol, FTY720) can also be phosphorylated and induces lymphopenia by downregulating cell surface expression of the S1P₁ receptor on lymphocytes. To analyze the role of S1P in lymphocyte circulation and distribution we established a high-performance-liquid-chromatography-based method for parallel detection and quantification of Fingolimod, sphingosine, and dihydrosphingosine together with their phosphorylated derivatives Fingolimod-phosphate, S1P, and dihydrosphingosine 1-phosphate. Phosphorylated and nonphosphorylated lipids were efficiently isolated from biological samples such as cells, tissues, serum, plasma, and media by simple chloroform extraction. Fluorescence labeling with 9-fluorenylmethyl chloroformiate ensured high selectivity and enhanced sensitivity for sphingolipid detection. The described method provides an accurate approach to investigate phosphorylation, dephosphorylation, hydrolyzation, and dehydrolyzation of sphingolipids and analogs. In addition it works independently from enzymatic conversions, measuring actual concentrations rather than enzymatic activities.     

Neutral ceramidase encoded by the Asah2 gene is essential for the intestinal degradation of sphingolipids

Complex sphingolipids are abundant as eukaryotic cell membrane components, whereas their metabolites, in particular ceramide, sphingosine, and sphingosine 1-phosphate, are involved in diverse cell signaling processes. In mammals, degradation of ceramide by ceramidase yields sphingosine, which is phosphorylated by the action of sphingosine kinase to generate sphingosine 1-phosphate. Therefore, ceramidases are key enzymes in the regulation of the cellular levels of ceramide, sphingosine, and sphingosine 1-phosphate. To explore the physiological functions of a neutral ceramidase with diverse cellular locations, we disrupted the Asah2 gene in mice. Asah2 null mice have a normal life span and do not show obvious abnormalities or major alterations in total ceramide levels in tissues. The Asah2-encoded neutral ceramidase is highly expressed in the small intestine along the brush border, suggesting that the neutral ceramidase may be involved in a pathway for the digestion of dietary sphingolipids. Indeed, Asah2 null mice were deficient in the intestinal degradation of ceramide. Thus, the results indicate that the Asah2-encoded neutral ceramidase is a key enzyme for the catabolism of dietary sphingolipids and regulates the levels of bioactive sphingolipid metabolites in the intestinal tract.     

Involvement of sphingoid bases in mediating reactive oxygen intermediate production and programmed cell death in Arabidopsis

Sphingolipids have been suggested to act as second messengers for an array of cellular signaling activities in plant cells, including stress responses and programmed cell death （PCD）. However, the mechanisms underpinning these processes are not well understood. Here, we report that an Arabidopsis mutant, fumonisin B1 r_esistant11-1 （/br11-1）, which fails to generate reactive oxygen intermediates （ROIs）, is incapable of initiating PCD when the mutant is challenged by fumonisin B l （FB0, a specific inhibitor of ceramide synthase. Molecular analysis indicated that FBR11 encodes a long-chain base 1 （LCB 1） subunit of serine palmitoyltransferase （SPT）, which catalyzes the first rate-limiting step of de novo sphingolipid synthesis. Mass spectrometric analysis of the sphingolipid concentrations revealed that whereas the fbr11-1 mutation did not affect basal levels of sphingoid bases, the mutant showed attenuated formation of sphingoid bases in response to FBl. By a direct feeding experiment, we show that the free sphingoid bases dihydrosphingosine, phytosphingosine and sphingosine efficiently induce ROI generation followed by cell death. Conversely, ROI generation and cell death induced by dihydrosphingosine were specifically blocked by its phosphorylated form dihydrosphingosine- 1-phosphate in a dosedependent manner, suggesting that the maintenance of homeostasis between a free sphingoid base and its phosphorylated derivative is critical to determining the cell fate. Because alterations of the sphingolipid level occur prior to the ROI production, we propose that the free sphingoid bases are involved in the control of PCD in Arabidopsis, presumably through the regulation of the ROI level upon receiving different developmental or environmental cues.     

Rapid turnover of sphingosine synthesized de novo from [14C]serine by Chinese hamster ovary cells

It has been hypothesized that complex sphingolipids may serve as another "lipid second messenger" system via their hydrolysis to free sphingosine, which inhibits protein kinase C and affects multiple cellular functions. To investigate sphingolipid turnover, Chinese hamster ovary cells were pulse labelled with [14C]serine and the [14C]sphingosine in cellular sphingolipids was determined over time. Much of the radiolabelled sphingosine was initially seen in ceramides and was incorporated into sphingomyelin during the 5-hour chase. A major portion of the radiolabel that was initially seen in other sphingolipids disappeared over time. Overall, about half of the total long-chain bases made during this pulse were degraded within 2 to 5 h, depending on the method of analysis. Hence, a substantial portion of the sphingosine synthesized de novo by these cells is turned over fairly quickly. Since the doubling time of these cells is 12 h, this rapid turnover may reflect the remodelling of the cell surface, or the utilization of the free sphingosine derived from sphingolipid turnover, as part of the control of cell growth and division.     

Altered sphingolipid metabolism in human endometrial cancer

There is a growing body of evidence indicating that bioactive sphingolipids play a key role in cancer development, progression and metastasis. However, sphingolipid metabolism in malignant tumors is poorly investigated. Therefore, the aim of the present study was to examine the content of selected intermediates of ceramide metabolism and the activity of key enzymes of ceramide de novo synthesis and sphingosine-1-phosphate (S1P) production in the endometrial cancer. The specimens of cancer tissue and healthy endometrium were obtained from women undergoing surgery because of the cancer (n = 23) and because of myomas (n = 18), respectively. The content of sphinganine, dihydroceramide, ceramide, sphingosine and S1P was measured using high pressure liquid chromatography. The activity of the enzymes was determined using radioactive substrates. It has been found that the content of each examined sphingolipid was markedly elevated in the cancer tissue compared with the healthy endometrium. Namely, sphinganine, sphingosine and dihydroceramide by 3–4.6-fold, ceramide and S1P by 1.9- and 1.6-fold, respectively. Interestingly, the ratio of S1P to ceramide remained stable. The activity of serine palmitoyltransferase and sphingosine kinase 1 was increased by 2.3- and 2.6-fold, respectively. We conclude that endometrial carcinoma is characterized by profound changes in sphingolipid metabolism that likely contribute to its progression and chemoresistance.     

Gene Expression Alterations in the Sphingolipid Metabolism Pathways during Progression of Dementia and Alzheimer’s Disease: A Shift Toward Ceramide Accumulation at the Earliest Recognizable Stages of Alzheimer’s Disease?

There is mounting evidence linking Aβ₄₂ generation in Alzheimer’s disease (AD) with sphingomyelin catabolism. Using microarray technology to study 17 brain regions from subjects with varying severity of AD and dementia we detected multiple gene expression abnormalities of the key enzymes that control sphingolipid metabolism. These changes were correlated with the progression of clinical dementia. The upregulation of gene expression of the enzymes controlling synthesis de novo of Cer and the downregulation of the enzymes involved in glycosphingolipid synthesis was evident as early in disease progression as in mild dementia. Together these changes suggest a shift in sphingolipid metabolism towards accumulation of Cer, depletion of glycosphingolipids and the reduction of synthesis of the anti-apoptosis signaling lipid—sphingosine 1-phosphate as a function of disease progression. This disrupted balance within the sphingolipid metabolism may trigger signaling events promoting neurodegeneration across cortical regions. This potential mechanism may provide a link between lipid metabolism disturbance and AD. Special issue dedicated to John P. Blass.     

Metabolism and biological functions of two phosphorylated sphingolipids, sphingosine 1-phosphate and ceramide 1-phosphate

Sphingolipids are major lipid constituents of the eukaryotic plasma membrane. Without certain sphingolipids, cells and/or embryos cannot survive, indicating that sphingolipids possess important physiological functions that are not substituted for by other lipids. One such role may be signaling. Recent studies have revealed that some sphingolipid metabolites, such as long-chain bases (LCBs; sphingosine (Sph) in mammals), long-chain base 1-phosphates (LCBPs; sphingosine 1-phosphate (S1P) in mammals), ceramide (Cer), and ceramide 1-phosphate (C1P), act as signaling molecules. The addition of phosphate groups to LCB/Sph and Cer generates LCBP/S1P and C1P, respectively. These phospholipids exhibit completely different functions than those of their precursors. In this review, we describe recent advances in understanding the functions of LCBP/S1P and C1P in mammals and in the yeast Saccharomyces cerevisiae. Since LCB/Sph, LCBP/S1P, Cer, and C1P are mutually convertible, regulation of not only the total amount of the each lipid but also of the overall balance in cellular levels is important. Therefore, we describe in detail their metabolic pathways, as well as the genes involved in each reaction.     

A role for the de novo sphingolipids in apoptosis of photosensitized cells

Sphingolipids have been implicated in apoptosis after various stress inducers. To assess the involvement of the de novo sphingolipid pathway in apoptosis, photodynamic therapy (PDT) with the photosensitizer Pc 4 was used as a novel stress inducer. Here we provide biochemical and genetic evidence of the role of the de novo sphingolipids in apoptosis post-Pc 4-PDT. In Jurkat cells PDT-induced intracellular sphinganine accumulation, DEVDase activation, PARP cleavage, and apoptosis were suppressed by the de novo sphingolipid synthesis inhibitor ISP-1 (Myriocin). Coincubation with sphinganine, sphingosine, or C16-ceramide specifically reversed the antiapoptotic actions of ISP-1 or the singlet oxygen scavenger L-histidine. PDT-induced cytochrome c release from mitochondria into the cytosol was inhibited by L-histidine, but not by ISP-1. Cotreatment with sphinganine did not reverse the inhibitory effect of L-histidine. In addition, PDT-induced sphinganine accumulation and apoptosis were ISP-1-sensitive in A431 human epidermoid and HT29 human carcinoma cells. Furthermore, in LY-B cells, CHO-derived mutants deficient in the de novo sphingolipid synthesis enzyme serine palmitoyltransferase (SPT) activity, DEVDase activation and apoptosis were delayed and suppressed post-PDT. Hence, the data are consistent with the partial involvement of the de novo sphingolipid pathway in apoptosis via DEVDase activation downstream of mitochondrial cytochrome c release post-Pc 4-PDT.     

S1P metabolism in cancer and other pathological conditions

Nearly two decades ago, the sphingolipid metabolite sphingosine 1-phosphate was discovered to function as a lipid mediator and regulator of cell proliferation. Since that time, sphingosine 1-phosphate has been shown to mediate a diverse array of fundamental biological processes including cell proliferation, migration, invasion, angiogenesis, vascular maturation and lymphocyte trafficking. Sphingosine 1-phosphate acts primarily via signaling through five ubiquitously expressed G protein-coupled receptors. Intracellular sphingosine 1-phosphate molecules are transported extracellularly and gain access to cognate receptors for autocrine and paracrine signaling and for signaling at distant sites reached through blood and lymphatic circulation systems. Intracellular pools of sphingosine 1-phosphate available for signaling are tightly regulated primarily by three enzymes: sphinosine kinase, S1P lyase and S1P phosphatase. Alterations in sphingosine 1-phosphate as well as the enzymes involved in its synthesis and catabolism have been observed in many types of malignancy. These enzymes are being evaluated for their role in mediating cancer formation and progression, as well as their potential to serve as targets of anti-cancer therapeutics. In this review, the impact of sphingosine 1-phosphate, its cognate receptors, and the enzymes of sphingosine 1-phosphate metabolism on cell survival, apoptosis, autophagy, cellular transformation, invasion, angiogenesis and hypoxia in relation to cancer biology and treatment are discussed.