Publications by authors named "Cungui Mao"

54 Publications

Ceramides and sphingosine-1-phosphate mediate the distinct effects of M1/M2-macrophage infusion on liver recovery after hepatectomy.

Cell Death Dis 2021 Mar 26;12(4):324. Epub 2021 Mar 26.

Division of Hepatobiliopancreatic Surgery, Department of General Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, China.

Post-hepatectomy liver dysfunction is a life-threatening morbidity that lacks efficient therapy. Bioactive lipids involved in macrophage polarization crucially regulate tissue injury and regeneration. Herein, we investigate the key bioactive lipids that mediate the cytotherapeutic potential of polarized-macrophage for post-hepatectomy liver dysfunction. Untargeted lipidomics identified elevation of ceramide (CER) metabolites as signature lipid species relevant to M1/M2 polarization in mouse bone-marrow-derived-macrophages (BMDMs). M1 BMDMs expressed a CER-generation-metabolic pattern, leading to elevation of CER; M2 BMDMs expressed a CER-breakdown-metabolic pattern, resulting in upregulation of sphingosine-1-phosphate (S1P). After infusing M1- or M2-polarized BMDMs into the mouse liver after hepatectomy, we found that M1-BMDM infusion increased M1 polarization and CER accumulation, resulting in exaggeration of hepatocyte apoptosis and liver dysfunction. Conversely, M2-BMDM infusion enhanced M2 polarization and S1P generation, leading to alleviation of liver dysfunction with improved hepatocyte proliferation. Treatment of exogenous CER and S1P or inhibition CER and S1P synthesis by siRNA targeting relevant enzymes further revealed that CER induced apoptosis while S1P promoted proliferation in post-hepatectomy primary hepatocytes. In conclusion, CER and S1P are uncovered as critical lipid mediators for M1- and M2-polarized BMDMs to promote injury and regeneration in the liver after hepatectomy, respectively. Notably, the upregulation of hepatic S1P induced by M2-BMDM infusion may have therapeutic potential for post-hepatectomy liver dysfunction.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41419-021-03616-9DOI Listing
March 2021

Neutral Ceramidase Is Required for the Reproduction of Brown Planthopper, (Stål).

Front Physiol 2021 24;12:629532. Epub 2021 Feb 24.

State Key Laboratory of Rice Biology, Key Laboratory of Molecular Biology of Crop Pathogens and Insects, Ministry of Agriculture, Institute of Insect Sciences, Zhejiang University, Hangzhou, China.

Ceramides are bioactive sphingolipids that have been implicated in insect development; however, their role in insect reproduction remains poorly understood. Here, we report the pivotal role of neutral ceramidase (NCER) in the female reproduction of the brown planthopper (BPH), (Stål), a significant pest in rice cultivation in Asia. LC-MS/MS demonstrated that, among different developmental stages of BPH, the levels of ceramides were highest in 1st instar nymphs and lowest in adults. The transcription of was negatively correlated with the levels of ceramides at different developmental stages of BPH, in that the transcript levels of were the highest, whereas ceramides levels were the lowest in BPH adults. Knocking down through RNA interference (RNAi) increased the levels of ceramides in BPH females and ovaries, which resulted in a delay in oocyte maturation, a reduction in oviposition and egg hatching rate, as well as the production of vulnerable offspring. Transmission electron microscopy (TEM) analysis and TdT-mediated dUTP Nick-End Labeling (TUNEL) assays showed mitochondrial deficiency and apoptosis in -deficient oocytes. Taken together, these results suggest that NCER plays a crucial role in female reproduction in BPH, likely by regulating the levels of ceramides.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fphys.2021.629532DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7943485PMC
February 2021

Transcriptional changes revealed genes and pathways involved in the deficient testis caused by the inhibition of alkaline ceramidase (Dacer) in Drosophila melanogaster.

Arch Insect Biochem Physiol 2021 Mar 15;106(3):e21765. Epub 2021 Feb 15.

State Key Laboratory of Rice Biology, MOA Key Laboratory of Agricultural Entomology, Institute of Insect Science, Zhejiang University, Hangzhou, Zhejiang, China.

Sphingolipids are ubiquitous structural components of eukaryotic cell membranes which are vital for maintaining the integrity of cells. Alkaline ceramidase is a key enzyme in sphingolipid biosynthesis pathway; however, little is known about the role of the enzyme in the male reproductive system of Drosophila melanogaster. To investigate the impact of alkaline ceramidase (Dacer) on male Drosophila, we got Dacer deficiency mutants (MUs) and found they displayed apparent defects in the testis's phenotype. To profile the molecular changes associated with this abnormal phenotype, we performed de novo transcriptome analyses of the MU and wildtype (WT) testes; and revealed 1239 upregulated genes and 1102 downregulated genes. Then, six upregulated DEGs (papilin [Ppn], croquemort [Crq], terribly reduced optic lobes [Trol], Laminin, Wunen-2, collagen type IV alpha 1 [Cg25C]) and three downregulated DEGs (mucin related 18B [Mur18B], rhomboid-7 [Rho-7], CG3168) were confirmed through quantitative real-time polymerase chain reaction in WT and MU samples. The differentially expressed genes were mainly associated with catalytic activity, oxidoreductase activity and transmembrane transporter activity, which significantly contributed to extracellular matrix-receptor interaction, fatty acids biosynthesis as well as glycine, serine, and threonine metabolism. The results highlight the importance of Dacer in the reproductive system of D. melanogaster and provide valuable resources to dig out the specific biological functions of Dacer in insect reproduction.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/arch.21765DOI Listing
March 2021

Sphingosine kinase 1 downregulation is required for adaptation to serine deprivation.

FASEB J 2021 Feb;35(2):e21284

Department of Medicine, Stony Brook University, Stony Brook, NY, USA.

It has been well-established that cancer cells often display altered metabolic profiles, and recent work has concentrated on how cancer cells adapt to serine removal. Serine can be either taken exogenously or synthesized from glucose, and its regulation forms an important mechanism for nutrient integration. One of the several important metabolic roles for serine is in the generation of bioactive sphingolipids since it is the main substrate for serine palmitoyltransferase, the initial and rate-limiting enzyme in the synthesis of sphingolipids. Previously, serine deprivation has been connected to the action of the tumor suppressor p53, and we have previously published on a role for p53 regulating sphingosine kinase 1 (SK1), an enzyme that phosphorylates sphingosine to form sphingosine-1-phosphate (S1P). SK1 is a key enzyme in sphingolipid synthesis that functions in pro-survival and tumor-promoting pathways and whose expression is also often elevated in cancers. Here we show that SK1 was degraded during serine starvation in a time and dose-dependent manner, which led to sphingosine accumulation. This was independent of effects on p53 but required the action of the proteasome. Furthermore, we show that overexpression of SK1, to compensate for SK1 loss, was detrimental to cell growth under conditions of serine starvation, demonstrating that the suppression of SK1 under these conditions is adaptive. Mitochondrial oxygen consumption decreased in response to SK1 degradation, and this was accompanied by an increase in intracellular reactive oxygen species (ROS). Suppression of ROS with N-acteylcysteine resulted in suppression of the metabolic adaptations and in decreased cell growth under serine deprivation. The effects of SK1 suppression on ROS were mimicked by D-erythro-sphingosine, whereas S1P was ineffective, suggesting that the effects of loss of SK1 were due to the accumulation of its substrate sphingosine. This study reveals a new mechanism for regulating SK1 levels and a link of SK1 to serine starvation as well as mitochondrial function.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.202001814RRDOI Listing
February 2021

Alkaline ceramidase family: The first two decades.

Cell Signal 2021 Feb 1;78:109860. Epub 2020 Dec 1.

Department of Medicine, Stony Brook University, Stony Brook, New York 11794, USA; Stony Brook Cancer Center, Stony Brook University, Stony Brook, New York 11794, USA. Electronic address:

Ceramidases are a group of enzymes that catalyze the hydrolysis of ceramide, dihydroceramide, and phytoceramide into sphingosine (SPH), dihydrosphingosine (DHS), and phytosphingosine (PHS), respectively, along with a free fatty acid. Ceramidases are classified into the acid, neutral, and alkaline ceramidase subtypes according to the pH optima for their catalytic activity. YPC1 and YDC1 were the first alkaline ceramidase genes to be identified and cloned from the yeast Saccharomyces cerevisiae two decades ago. Subsequently, alkaline ceramidase genes were identified from other species, including one Drosophila melanogaster ACER gene (Dacer), one Arabidopsis thaliana ACER gene (AtACER), three Mus musculus ACER genes (Acer1, Acer2, and Acer3), and three Homo sapiens ACER genes (ACER1, ACER2, and ACER3). The protein products of these genes constitute a large protein family, termed the alkaline ceramidase (ACER) family. All the biochemically characterized members of the ACER family are integral membrane proteins with seven transmembrane segments in the Golgi complex or endoplasmic reticulum, and they each have unique substrate specificity. An increasing number of studies suggest that the ACER family has diverse roles in regulating sphingolipid metabolism and biological processes. Here we discuss the discovery of the ACER family, the biochemical properties, structures, and catalytic mechanisms of its members, and its role in regulating sphingolipid metabolism and biological processes in yeast, insects, plants, and mammals.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cellsig.2020.109860DOI Listing
February 2021

Transcriptional Regulation of Sphingosine Kinase 1.

Cells 2020 11 8;9(11). Epub 2020 Nov 8.

Department of Pharmacology, Stony Brook University, Stony Brook, NY 11794, USA.

Once thought to be primarily structural in nature, sphingolipids have become increasingly appreciated as second messengers in a wide array of signaling pathways. Sphingosine kinase 1, or SK1, is one of two sphingosine kinases that phosphorylate sphingosine into sphingosine-1-phosphate (S1P). S1P is generally pro-inflammatory, pro-angiogenic, immunomodulatory, and pro-survival; therefore, high SK1 expression and activity have been associated with certain inflammatory diseases and cancer. It is thus important to develop an understanding of the regulation of SK1 expression and activity. In this review, we explore the current literature on SK1 transcriptional regulation, illustrating a complex system of transcription factors, cytokines, and even micro-RNAs (miRNAs) on the post transcriptional level.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/cells9112437DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7695205PMC
November 2020

Maternal and fetal alkaline ceramidase 2 is required for placental vascular integrity in mice.

FASEB J 2020 Nov 22;34(11):15252-15268. Epub 2020 Sep 22.

Department of Medicine, Stony Brook University, Stony Brook, NY, USA.

Sphingolipids have been implicated in mammalian placental development and function, but their regulation in the placenta remains unclear. Herein we report that alkaline ceramidase 2 (ACER2) plays a key role in sustaining the integrity of the placental vasculature by regulating the homeostasis of sphingolipids in mice. The mouse alkaline ceramidase 2 gene (Acer2) is highly expressed in the placenta between embryonic day (E) 9.5 and E12.5. Acer2 deficiency in both the mother and fetus decreases the placental levels of sphingolipids, including sphingoid bases (sphingosine and dihydrosphingosine) and sphingoid base-1-phosphates (sphingosine-1-phosphate and dihydrosphingosine-1-phosphate) and results in the in utero death of ≈50% of embryos at E12.5 whereas Acer2 deficiency in either the mother or fetus has no such effects. Acer2 deficiency causes hemorrhages from the maternal vasculature in the junctional and/or labyrinthine zones in E12.5 placentas. Moreover, hemorrhagic but not non-hemorrhagic Acer2-deficient placentas exhibit an expansion of parietal trophoblast giant cells with a concomitant decrease in the area of the fetal blood vessel network in the labyrinthine zone, suggesting that Acer2 deficiency results in embryonic lethality due to the atrophy of the fetal blood vessel network in the placenta. Taken together, these results suggest that ACER2 sustains the integrity of the placental vasculature by controlling the homeostasis of sphingolipids in mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.202001104RDOI Listing
November 2020

Generation of sphingosine-1-phosphate by sphingosine kinase 1 protects nonalcoholic fatty liver from ischemia/reperfusion injury through alleviating reactive oxygen species production in hepatocytes.

Free Radic Biol Med 2020 11 30;159:136-149. Epub 2020 Jul 30.

Division of Hepatobiliopancreatic Surgery, Department of General Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, China. Electronic address:

Background: Nonalcoholic fatty liver (NAFL) is emerging as a leading risk factor of hepatic ischemia/reperfusion (I/R) injury lacking of effective therapy. Lipid dyshomeostasis has been implicated in the hepatopathy of NAFL. Herein, we investigate the bioactive lipids that critically regulate I/R injury in NAFL.

Methods: Lipidomics were performed to identify dysregulated lipids in mouse and human NAFL with I/R injury. The alteration of corresponding lipid-metabolizing genes was examined. The effects of the dysregulated lipid metabolism on I/R injury in NAFL were evaluated in mice and primary hepatocytes.

Results: Sphingolipid metabolic pathways responsible for the generation of sphingosine-1-phosphate (S1P) were uncovered to be substantially activated by I/R in mouse NAFL. Sphingosine kinase 1 (Sphk1) was found to be essential for hepatic S1P generation in response to I/R in hepatocytes of NAFL mice. Sphk1 knockdown inhibited the hepatic S1P rise while accumulating ceramides in hepatocytes of NAFL mice, leading to aggressive hepatic I/R injury with upregulation of oxidative stress and increase of reactive oxygen species (ROS). In contrast, administration of exogenous S1P protected hepatocytes of NAFL mice from hepatic I/R injury. Clinical study revealed a significant activation of S1P generation by I/R in liver specimens of NAFL patients. In vitro studies on the L02 human hepatocytes consolidated that inhibiting the generation of S1P by knocking down SPHK1 exaggerated I/R-induced damage and oxidative stress in human hepatocytes of NAFL.

Conclusions: Generation of S1P by SPHK1 is important for protecting NAFL from I/R injury, which may serve as therapeutic targets for hepatic I/R injury in NAFL.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.freeradbiomed.2020.07.004DOI Listing
November 2020

Elusive Roles of the Different Ceramidases in Human Health, Pathophysiology, and Tissue Regeneration.

Cells 2020 06 2;9(6). Epub 2020 Jun 2.

Department of Periodontology, College of Dental Medicine, Nova Southeastern University, Fort Lauderdale, FL 33324, USA.

Ceramide and sphingosine are important interconvertible sphingolipid metabolites which govern various signaling pathways related to different aspects of cell survival and senescence. The conversion of ceramide into sphingosine is mediated by ceramidases. Altogether, five human ceramidases-named acid ceramidase, neutral ceramidase, alkaline ceramidase 1, alkaline ceramidase 2, and alkaline ceramidase 3-have been identified as having maximal activities in acidic, neutral, and alkaline environments, respectively. All five ceramidases have received increased attention for their implications in various diseases, including cancer, Alzheimer's disease, and Farber disease. Furthermore, the potential anti-inflammatory and anti-apoptotic effects of ceramidases in host cells exposed to pathogenic bacteria and viruses have also been demonstrated. While ceramidases have been a subject of study in recent decades, our knowledge of their pathophysiology remains limited. Thus, this review provides a critical evaluation and interpretive analysis of existing literature on the role of acid, neutral, and alkaline ceramidases in relation to human health and various diseases, including cancer, neurodegenerative diseases, and infectious diseases. In addition, the essential impact of ceramidases on tissue regeneration, as well as their usefulness in enzyme replacement therapy, is also discussed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/cells9061379DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349419PMC
June 2020

Correction: Targeting alkaline ceramidase 3 alleviates the severity of nonalcoholic steatohepatitis by reducing oxidative stress.

Cell Death Dis 2020 Mar 17;11(3):191. Epub 2020 Mar 17.

Department of Medicine and Cancer Center, the State University of New York at Stony Brook, Stony Brook, New York, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41419-020-2396-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7078179PMC
March 2020

Targeting alkaline ceramidase 3 alleviates the severity of nonalcoholic steatohepatitis by reducing oxidative stress.

Cell Death Dis 2020 01 16;11(1):28. Epub 2020 Jan 16.

Department of Medicine and Cancer Center, the State University of New York at Stony Brook, Stony Brook, New York, USA.

Overload of palmitic acids is linked to the dysregulation of ceramide metabolism in nonalcoholic steatohepatitis (NASH), and ceramides are important bioactive lipids mediating the lipotoxicity of palmitic acid in NASH. However, much remains unclear about the role of ceramidases that catalyze the hydrolysis of ceramides in NASH. By analyzing the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database, we found that alkaline ceramidase 3 (ACER3) is upregulated in livers of patients with NASH. Consistently, we found that Acer3 mRNA levels and its enzymatic activity were also upregulated in mouse livers with NASH induced by a palmitate-enriched Western diet (PEWD). Moreover, we demonstrated that palmitate treatment also elevated Acer3 mRNA levels and its enzymatic activity in mouse primary hepatocytes. In order to investigate the function of Acer3 in NASH, Acer3 null mice and their wild-type littermates were fed a PEWD to induce NASH. Knocking out Acer3 was found to augment PEWD-induced elevation of C-ceramide and alleviate early inflammation and fibrosis but not steatosis in mouse livers with NASH. In addition, Acer3 deficiency attenuated hepatocyte apoptosis in livers with NASH. These protective effects of Acer3 deficiency were found to be associated with suppression of hepatocellular oxidative stress in NASH liver. In vitro studies further revealed that loss of ACER3/Acer3 increased C-ceramide and inhibited apoptosis and oxidative stress in mouse primary hepatocytes and immortalized human hepatocytes induced by palmitic-acid treatment. These results suggest that ACER3 plays an important pathological role in NASH by mediating palmitic-acid-induced oxidative stress.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41419-019-2214-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6965144PMC
January 2020

Identification of as a First Susceptibility Gene for Lithium-Induced Nephrogenic Diabetes Insipidus in Mice.

J Am Soc Nephrol 2019 12 26;30(12):2322-2336. Epub 2019 Sep 26.

The Jackson Laboratory, Bar Harbor, Maine;

Background: Lithium, mainstay treatment for bipolar disorder, causes nephrogenic diabetes insipidus and hypercalcemia in about 20% and 10% of patients, respectively, and may lead to acidosis. These adverse effects develop in only a subset of patients treated with lithium, suggesting genetic factors play a role.

Methods: To identify susceptibility genes for lithium-induced adverse effects, we performed a genome-wide association study in mice, which develop such effects faster than humans. On day 8 and 10 after assigning female mice from 29 different inbred strains to normal chow or lithium diet (40 mmol/kg), we housed the animals for 48 hours in metabolic cages for urine collection. We also collected blood samples.

Results: In 17 strains, lithium treatment significantly elevated urine production, whereas the other 12 strains were not affected. Increased urine production strongly correlated with lower urine osmolality and elevated water intake. Lithium caused acidosis only in one mouse strain, whereas hypercalcemia was found in four strains. Lithium effects on blood pH or ionized calcium did not correlate with effects on urine production. Using genome-wide association analyses, we identified eight gene-containing loci, including a locus containing , which encodes a ceramidase and is specifically expressed in the collecting duct. Knockout of led to increased susceptibility for lithium-induced diabetes insipidus development.

Conclusions: We demonstrate that genome-wide association studies in mice can be used successfully to identify susceptibility genes for development of lithium-induced adverse effects. We identified as a first susceptibility gene for lithium-induced diabetes insipidus in mice.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1681/ASN.2018050549DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6900794PMC
December 2019

DEGS1 variant causes neurological disorder.

Eur J Hum Genet 2019 11 11;27(11):1668-1676. Epub 2019 Jun 11.

The Morris Kahn Laboratory of Human Genetics, National Institute for Biotechnology in the Negev and Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, 84105, Israel.

Sphingolipidoses are monogenic lipid storage diseases caused by variants in enzymes of lipid synthesis and metabolism. We describe an autosomal recessive complex neurological disorder affecting consanguineous kindred. All four affected individuals, born at term following normal pregnancies, had mild to severe intellectual disability, spastic quadriplegia, scoliosis and epilepsy in most, with no dysmorphic features. Brain MRI findings were suggestive of leukodystrophy, with abnormal hyperintense signal in the periventricular perioccipital region and thinning of the body of corpus callosum. Notably, all affected individuals were asymptomatic at early infancy and developed normally until the age of 8-18 months, when deterioration ensued. Homozygosity mapping identified a single 8.7 Mb disease-associated locus on chromosome 1q41-1q42.13 between rs1511695 and rs537250 (two-point LOD score 2.1). Whole exome sequencing, validated through Sanger sequencing, identified within this locus a single disease-associated homozygous variant in DEGS1, encoding C4-dihydroceramide desaturase, an enzyme of the ceramide synthesis pathway. The missense variant, segregating within the family as expected for recessive heredity, affects an evolutionary-conserved amino acid of all isoforms of DEGS1 (c.656A>G, c.764A>G; p.(N219S), p.(N255S)) and was not found in a homozygous state in ExAC and gnomAD databases or in 300 ethnically matched individuals. Lipidomcs analysis of whole blood of affected individuals demonstrated augmented levels of dihydroceramides, dihydrosphingosine, dihydrosphingosine-1-phosphate and dihydrosphingomyelins with reduced levels of ceramide, sphingosine, sphingosine-1-phosphate and monohexosylceramides, as expected in malfunction of C4-dihydroceramide desaturase. Thus, we describe a sphingolipidosis causing a severe regressive neurological disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41431-019-0444-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6871177PMC
November 2019

New fluorogenic probes for neutral and alkaline ceramidases.

J Lipid Res 2019 06 29;60(6):1174-1181. Epub 2019 Mar 29.

Spanish National Research Council (CSIC), Institute for Advanced Chemistry of Catalonia (IQAC-CSIC), Department of Biological Chemistry, Research Unit on Bioactive Molecules (RUBAM), 08034 Barcelona, Spain

New fluorogenic ceramidase substrates derived from the -acyl modification of our previously reported probes (RBM14) are reported. While none of the new probes were superior to the known RBM14C12 as acid ceramidase substrates, the corresponding nervonic acid amide (RBM14C24:1) is an efficient and selective substrate for the recombinant human neutral ceramidase, both in cell lysates and in intact cells. A second generation of substrates, incorporating the natural 2-(-acylamino)-1,3-diol-4-ene framework (compounds RBM15) is also reported. Among them, the corresponding fatty acyl amides with an unsaturated -acyl chain can be used as substrates to determine alkaline ceramidase (ACER)1 and ACER2 activities. In particular, compound RBM15C18:1 has emerged as the best fluorogenic probe reported so far to measure ACER1 and ACER2 activities in a 96-well plate format.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1194/jlr.D092759DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6547634PMC
June 2019

Alkaline ceramidase 2 is essential for the homeostasis of plasma sphingoid bases and their phosphates.

FASEB J 2018 06 22;32(6):3058-3069. Epub 2018 Jan 22.

Department of Medicine, State University of New York at Stony Brook, Stony Brook, New York, USA.

Sphingosine-1-phosphate (S1P) plays important roles in cardiovascular development and immunity. S1P is abundant in plasma because erythrocytes-the major source of S1P-lack any S1P-degrading activity; however, much remains unclear about the source of the plasma S1P precursor, sphingosine (SPH), derived mainly from the hydrolysis of ceramides by the action of ceramidases that are encoded by 5 distinct genes, acid ceramidase 1 ( ASAH1)/ Asah1, ASAH2/ Asah2, alkaline ceramidase 1 ( ACER1)/ Acer1, ACER2/ Acer2, and ACER3/ Acer3, in humans/mice. Previous studies have reported that knocking out Asah1 or Asah2 failed to reduce plasma SPH and S1P levels in mice. In this study, we show that knocking out Acer1 or Acer3 also failed to reduce the blood levels of SPH or S1P in mice. In contrast, knocking out Acer2 from either whole-body or the hematopoietic lineage markedly decreased the blood levels of SPH and S1P in mice. Of interest, knocking out Acer2 from whole-body or the hematopoietic lineage also markedly decreased the levels of dihydrosphingosine (dhSPH) and dihydrosphingosine-1-phosphate (dhS1P) in blood. Taken together, these results suggest that ACER2 plays a key role in the maintenance of high plasma levels of sphingoid base-1-phosphates-S1P and dhS1P-by controlling the generation of sphingoid bases-SPH and dhSPH-in hematopoietic cells.-Li, F., Xu, R., Low, B. E., Lin, C.-L., Garcia-Barros, M., Schrandt, J., Mileva, I., Snider, A., Luo, C. K., Jiang, X.-C., Li, M.-S., Hannun, Y. A., Obeid, L. M., Wiles, M. V., Mao, C. Alkaline ceramidase 2 is essential for the homeostasis of plasma sphingoid bases and their phosphates.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1096/fj.201700445RRDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5956249PMC
June 2018

A neutral ceramidase, NlnCDase, is involved in the stress responses of brown planthopper, Nilaparvata lugens (Stål).

Sci Rep 2018 01 18;8(1):1130. Epub 2018 Jan 18.

State Key Laboratory of Rice Biology, Key Laboratory of Molecular Biology of Crop Pathogens and Insects, Ministry of Agriculture, and Institute of Insect Sciences, Zhejiang University, Hangzhou, Zhejiang, 310058, China.

Ceramidases (CDases) are vital enzymes involved in the biosynthesis of sphingolipids, which are essential components of eukaryotic membranes. The function of these enzymes in insects, however, is poorly understood. We identified a neutral ceramidase (NlnCDase) from the brown planthopper, Nilaparvata lugens, one of the most destructive hemipteran pests of rice. The C12-ceramide was the most preferred substrate for the NlnCDase enzyme. The activity of the NlnCDase enzyme was highest in the neutral-pH range (pH 6.0). It was inhibited by EGTA, Cs and Fe, while stimulated by EDTA and Ca. Moreover, the NlnCDase has higher transcript level and activity in adults than in eggs and nymphs, and in the reproductive organs (ovaries and spermaries) than in other tissues (i.e. heads, thorax, legs, midguts), which suggested that the NlnCDase might be elevated to mediate developmental process. In addition, transcripts and activity of the NlnCDase were up-regulated under abiotic stresses including starvation, abnormal temperature, and insecticides, and biotic stress of resistant rice varieties. Knocking down NlnCDase by RNA interference increased female survival under starvation and temperature stresses, suggesting that NlnCDase might be involved in the stress response in N. lugens.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-018-19219-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5773612PMC
January 2018

Tumor suppressor p53 links ceramide metabolism to DNA damage response through alkaline ceramidase 2.

Cell Death Differ 2018 05 11;25(5):841-856. Epub 2017 Dec 11.

Department of Medicine, State University of New York (SUNY), Stony Brook, NY, 11794, USA.

p53 mediates the DNA damage response (DDR) by regulating the expression of genes implicated in cell cycle arrest, senescence, programmed cell death (PCD), and metabolism. Herein we demonstrate that human alkaline ceramidase 2 (ACER2) is a novel transcriptional target of p53 and that its transactivation by p53 mediates the DDR. We found that p53 overexpression or its activation by ionizing radiation (IR) upregulated ACER2 in cells. Two putative p53 responsive elements (p53REs) were found in its first intron of the ACER2 gene, and Chromatin Immunoprecipitation (ChIP) assays in combination with promoter activity assays demonstrated that these p53REs are the bona fide p53 binding sites that mediate ACER2 transactivation by p53. As ACER2 catalyzes the hydrolysis of ceramides into sphingosine, which in turn is phosphorylated to form sphingosine-1-phosphate (S1P), ACER2 upregulation increased the levels of both sphingosine and S1P while decreasing the levels of ceramides in cells. A moderate upregulation of ACER2 inhibited cell cycle arrest and cellular senescence in response to low-level expression of p53 or low-dose IR by elevating S1P, a pro-proliferative and pro-survival bioactive lipid, and/or decreasing ceramides whereas its robust upregulation mediated PCD in response to high-level expression of p53 or high-dose IR likely by accumulating cellular sphingosine, a pro-death bioactive lipid. ACER2 is frequently inactivated in various cancers due to its deletion or mutations, and restoring its expression inhibited the growth of tumor xenografts in mice. These results suggest that p53 mediates DDR and exerts its tumor suppressive role in part by regulating the expression of ACER2, which in turn regulates the bioactive sphingolipid lipids.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41418-017-0018-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5943524PMC
May 2018

Alkaline Ceramidase 1 Protects Mice from Premature Hair Loss by Maintaining the Homeostasis of Hair Follicle Stem Cells.

Stem Cell Reports 2017 11 19;9(5):1488-1500. Epub 2017 Oct 19.

Department of Medicine, Stony Brook University, Stony Brook, NY, USA; Department of Medicine and Stony Brook Cancer Center, Stony Brook University, HSC T15-023, Stony Brook, NY 11794, USA; Graduate Program in Molecular and Cellular Biology, Stony Brook University, Stony Brook, NY, USA; Department of Dermatology, Stony Brook University, Stony Brook, NY, USA. Electronic address:

Ceramides and their metabolites are important for the homeostasis of the epidermis, but much remains unknown about the roles of specific pathways of ceramide metabolism in skin biology. With a mouse model deficient in the alkaline ceramidase (Acer1) gene, we demonstrate that ACER1 plays a key role in the homeostasis of the epidermis and its appendages by controlling the metabolism of ceramides. Loss of Acer1 elevated the levels of various ceramides and sphingoid bases in the skin and caused progressive hair loss in mice. Mechanistic studies revealed that loss of Acer1 widened follicular infundibulum and caused progressive loss of hair follicle stem cells (HFSCs) due to reduced survival and stemness. These results suggest that ACER1 plays a key role in maintaining the homeostasis of HFSCs, and thereby the hair follicle structure and function, by regulating the metabolism of ceramides in the epidermis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.stemcr.2017.09.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5829345PMC
November 2017

Anticancer actions of lysosomally targeted inhibitor, LCL521, of acid ceramidase.

PLoS One 2017 14;12(6):e0177805. Epub 2017 Jun 14.

Department of Medicine, Stony Brook University, Stony Brook, New York, United States of America.

Acid ceramidase, which catalyzes ceramide hydrolysis to sphingosine and free fatty acid mainly in the lysosome, is being recognized as a potential therapeutic target for cancer. B13 is an effective and selective acid ceramidase inhibitor in vitro, but not as effective in cells due to poor access to the lysosomal compartment. In order to achieve targeting of B13 to the lysosome, we designed lysosomotropic N, N-dimethyl glycine (DMG)-conjugated B13 prodrug LCL521 (1,3-di-DMG-B13). Our previous results indicated the efficient delivery of B13 to the lysosome resulted in augmented effects of LCL521 on cellular acid ceramidase as evaluated by effects on substrate/product levels. Our current studies indicate that functionally, this translated into enhanced inhibition of cell proliferation. Moreover, there were greater synergistic effects of LCL521 with either ionizing radiation or Tamoxifen. Taken together, these results clearly indicate that compartmental targeting for the inhibition of acid ceramidase is an efficient and valuable therapeutic strategy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0177805PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5470663PMC
September 2017

Tricyclic Antidepressants Promote Ceramide Accumulation to Regulate Collagen Production in Human Hepatic Stellate Cells.

Sci Rep 2017 03 21;7:44867. Epub 2017 Mar 21.

Gastrointestinal Unit, Massachusetts General Hospital, Harvard Medical School, Boston, MA USA.

Activation of hepatic stellate cells (HSCs) in response to injury is a key step in hepatic fibrosis, and is characterized by trans-differentiation of quiescent HSCs to HSC myofibroblasts, which secrete extracellular matrix proteins responsible for the fibrotic scar. There are currently no therapies to directly inhibit hepatic fibrosis. We developed a small molecule screen to identify compounds that inactivate human HSC myofibroblasts through the quantification of lipid droplets. We screened 1600 compounds and identified 21 small molecules that induce HSC inactivation. Four hits were tricyclic antidepressants (TCAs), and they repressed expression of pro-fibrotic factors Alpha-Actin-2 (ACTA2) and Alpha-1 Type I Collagen (COL1A1) in HSCs. RNA sequencing implicated the sphingolipid pathway as a target of the TCAs. Indeed, TCA treatment of HSCs promoted accumulation of ceramide through inhibition of acid ceramidase (aCDase). Depletion of aCDase also promoted accumulation of ceramide and was associated with reduced COL1A1 expression. Treatment with B13, an inhibitor of aCDase, reproduced the antifibrotic phenotype as did the addition of exogenous ceramide. Our results show that detection of lipid droplets provides a robust readout to screen for regulators of hepatic fibrosis and have identified a novel antifibrotic role for ceramide.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep44867DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5359599PMC
March 2017

Deficiency of the sphingosine-1-phosphate lyase SGPL1 is associated with congenital nephrotic syndrome and congenital adrenal calcifications.

Hum Mutat 2017 04 6;38(4):365-372. Epub 2017 Mar 6.

Department of Pediatrics I, Medical University of Innsbruck, Innsbruck, Austria.

We identified two unrelated consanguineous families with three children affected by the rare association of congenital nephrotic syndrome (CNS) diagnosed in the first days of life, of hypogonadism, and of prenatally detected adrenal calcifications, associated with congenital adrenal insufficiency in one case. Using exome sequencing and targeted Sanger sequencing, two homozygous truncating mutations, c.1513C>T (p.Arg505*) and c.934delC (p.Leu312Phefs*30), were identified in SGPL1-encoding sphingosine-1-phosphate (S1P) lyase 1. SGPL1 catalyzes the irreversible degradation of endogenous and dietary S1P, the final step of sphingolipid catabolism, and of other phosphorylated long-chain bases. S1P is an intracellular and extracellular signaling molecule involved in angiogenesis, vascular maturation, and immunity. The levels of SGPL1 substrates, S1P, and sphingosine were markedly increased in the patients' blood and fibroblasts, as determined by liquid chromatography-tandem mass spectrometry. Vascular alterations were present in a patient's renal biopsy, in line with changes seen in Sgpl1 knockout mice that are compatible with a developmental defect in vascular maturation. In conclusion, loss of SGPL1 function is associated with CNS, adrenal calcifications, and hypogonadism.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/humu.23192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5384969PMC
April 2017

Molecular Characterization of Rice Gene and Functional Analysis of its Role in Insect Resistance.

Front Plant Sci 2016 1;7:1789. Epub 2016 Dec 1.

State Key Laboratory of Rice Biology, Key Laboratory of Agricultural Entomology, Ministry of Agriculture and Institute of Insect Sciences, Zhejiang University Hangzhou, China.

In plants, sphingolipids, such as long-chain bases (LCBs), act as bioactive molecules in stress responses. Until now, it is still not clear if these lipids are involved in biotic stress responses to herbivore. Herein we report that a rice LCB gene, encoding a subunit of serine palmitoyltransferase (SPT), a key enzyme responsible for the biosynthesis of sphingolipids, plays a critical role in plant defense response to the brown planthopper (BPH) attack and that its up-regulation protects plants from herbivore infestation. Transcripts of gene in rice seedlings were increased at 4 h, but decreased at 8-24 h after BPH attack. Sphingolipid measurement profiling revealed that overexpression of in increased trihydroxylated LCB phytosphingosine (t18:0) and phytoceramide by 1.7 and 1.3-fold, respectively, compared with that of wild type (WT) plants. Transgenic Arabidopsis plants also showed higher callose and wax deposition in leaves than that of WT. Overexpression of gene in reduced the population size of green peach aphid (). Moreover, the electrical penetration graph (EPG) results indicated that the aphids encounter resistance factors while reaching for the phloem on the transgenic plants. The defense response genes related to salicylic acid signaling pathway, remained uplgulated in the -overexpressing transgenic plants. Our data highlight the key functions of in biotic stress response in plants.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fpls.2016.01789DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5130998PMC
December 2016

Ceramidases, roles in sphingolipid metabolism and in health and disease.

Adv Biol Regul 2017 Jan 11;63:122-131. Epub 2016 Oct 11.

Health Science Center, Stony Brook University, 100 Nicolls Road, L4, 182, 11794, Stony Brook, NY, USA. Electronic address:

Over the past three decades, extensive research has been able to determine the biologic functions for the main bioactive sphingolipids, namely ceramide, sphingosine, and sphingosine 1-phosphate (S1P) (Hannun, 1996; Hannun et al., 1986; Okazaki et al., 1989). These studies have managed to define the metabolism, regulation, and function of these bioactive sphingolipids. This emerging body of literature has also implicated bioactive sphingolipids, particularly S1P and ceramide, as key regulators of cellular homeostasis. Ceramidases have the important role of cleaving fatty acid from ceramide and producing sphingosine, thereby controlling the interconversion of these two lipids. Thus far, five human ceramidases encoded by five different genes have been identified: acid ceramidase (AC), neutral ceramidase (NC), alkaline ceramidase 1 (ACER1), alkaline ceramidase 2 (ACER2), and alkaline ceramidase 3 (ACER3). These ceramidases are classified according to their optimal pH for catalytic activity. AC, which is localized to the lysosomal compartment, has been associated with Farber's disease and is involved in the regulation of cell viability. Neutral ceramidase, which is localized to the plasma membrane and primarily expressed in the small intestine and colon, is involved in digestion, and has been implicated in colon carcinogenesis. ACER1 which can be found in the endoplasmic reticulum and is highly expressed in the skin, plays an important role in keratinocyte differentiation. ACER2, localized to the Golgi complex and highly expressed in the placenta, is involved in programed cell death in response to DNA damage. ACER3, also localized to the endoplasmic reticulum and the Golgi complex, is ubiquitously expressed, and is involved in motor coordination-associated Purkinje cell degeneration. This review seeks to consolidate the current knowledge regarding these key cellular players.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jbior.2016.10.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5330250PMC
January 2017

Aging-related elevation of sphingoid bases shortens yeast chronological life span by compromising mitochondrial function.

Oncotarget 2016 Apr;7(16):21124-44

Department of Medicine, Stony Brook University, Stony Brook, NY, USA.

Sphingoid bases (SBs) as bioactive sphingolipids, have been implicated in aging in yeast. However, we know neither how SBs are regulated during yeast aging nor how they, in turn, regulate it. Herein, we demonstrate that the yeast alkaline ceramidases (YPC1 and YDC1) and SB kinases (LCB4 and LCB5) cooperate in regulating SBs during the aging process and that SBs shortens chronological life span (CLS) by compromising mitochondrial functions. With a lipidomics approach, we found that SBs were increased in a time-dependent manner during yeast aging. We also demonstrated that among the enzymes known for being responsible for the metabolism of SBs, YPC1 was upregulated whereas LCB4/5 were downregulated in the course of aging. This inverse regulation of YPC1 and LCB4/5 led to the aging-related upregulation of SBs in yeast and a reduction in CLS. With the proteomics-based approach (SILAC), we revealed that increased SBs altered the levels of proteins related to mitochondria. Further mechanistic studies demonstrated that increased SBs inhibited mitochondrial fusion and caused fragmentation, resulting in decreases in mtDNA copy numbers, ATP levels, mitochondrial membrane potentials, and oxygen consumption. Taken together, these results suggest that increased SBs mediate the aging process by impairing mitochondrial structural integrity and functions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.8195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5008273PMC
April 2016

Alkaline ceramidase 2 and its bioactive product sphingosine are novel regulators of the DNA damage response.

Oncotarget 2016 Apr;7(14):18440-57

Department of Medicine, State University of New York at Stony Brook, Stony Brook, NY 11794, USA.

Human cells respond to DNA damage by elevating sphingosine, a bioactive sphingolipid that induces programmed cell death (PCD) in response to various forms of stress, but its regulation and role in the DNA damage response remain obscure. Herein we demonstrate that DNA damage increases sphingosine levels in tumor cells by upregulating alkaline ceramidase 2 (ACER2) and that the upregulation of the ACER2/sphingosine pathway induces PCD in response to DNA damage by increasing the production of reactive oxygen species (ROS). Treatment with the DNA damaging agent doxorubicin increased both ACER2 expression and sphingosine levels in HCT116 cells in a dose-dependent manner. ACER2 overexpression increased sphingosine in HeLa cells whereas knocking down ACER2 inhibited the doxorubicin-induced increase in sphingosine in HCT116 cells, suggesting that DNA damage elevates sphingosine by upregulating ACER2. Knocking down ACER2 inhibited an increase in the apoptotic and necrotic cell population and the cleavage of poly ADP ribose polymerase (PARP) in HCT116 cells in response to doxorubicin as well as doxorubicin-induced release of lactate dehydrogenase (LDH) from these cells. Similar to treatment with doxorubicin, ACER2 overexpression induced an increase in the apoptotic and necrotic cell population and PARP cleavage in HeLa cells and LDH release from cells, suggesting that ACER2 upregulation mediates PCD in response to DNA damage through sphingosine. Mechanistic studies demonstrated that the upregulation of the ACER2/sphingosine pathway induces PCD by increasing ROS levels. Taken together, these results suggest that the ACER2/sphingosine pathway mediates PCD in response to DNA damage through ROS production.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.18632/oncotarget.7825DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4951300PMC
April 2016

Deficiency of the alkaline ceramidase ACER3 manifests in early childhood by progressive leukodystrophy.

J Med Genet 2016 06 20;53(6):389-96. Epub 2016 Jan 20.

The Monique and Jacques Roboh Department of Genetic Research, Hadassah, Hebrew University Medical Center, Jerusalem, Israel.

Background/aims: Leukodystrophies due to abnormal production of myelin cause extensive morbidity in early life; their genetic background is still largely unknown. We aimed at reaching a molecular diagnosis in Ashkenazi-Jewish patients who suffered from developmental regression at 6-13 months, leukodystrophy and peripheral neuropathy.

Methods: Exome analysis, determination of alkaline ceramidase activity catalysing the conversion of C18:1-ceramide to sphingosine and D-ribo-C12-N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) (NBD)-phytoceramide to NBD-C12-fatty acid using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and thin layer chromatography, respectively, and sphingolipid analysis in patients' blood by LC-MS/MS.

Results: The patients were homozygous for p.E33G in the ACER3, which encodes a C18:1-alkaline ceramidase and C20:1-alkaline ceramidase. The mutation abolished ACER3 catalytic activity in the patients' cells and failed to restore alkaline ceramidase activity in yeast mutant strain. The levels of ACER3 substrates, C18:1-ceramides and dihydroceramides and C20:1-ceramides and dihydroceramides and other long-chain ceramides and dihydroceramides were markedly increased in the patients' plasma, along with that of complex sphingolipids, including monohexosylceramides and lactosylceramides.

Conclusions: Homozygosity for the p.E33G mutation in the ACER3 gene results in inactivation of ACER3, leading to the accumulation of various sphingolipids in blood and probably in brain, likely accounting for this new form of childhood leukodystrophy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5068917PMC
http://dx.doi.org/10.1136/jmedgenet-2015-103457DOI Listing
June 2016

Alkaline Ceramidase 3 Deficiency Results in Purkinje Cell Degeneration and Cerebellar Ataxia Due to Dyshomeostasis of Sphingolipids in the Brain.

PLoS Genet 2015 Oct 16;11(10):e1005591. Epub 2015 Oct 16.

Department of Medicine, Stony Brook University, Stony Brook, New York, United States of America; Stony Brook Cancer Center, Stony Brook, New York, United States of America.

Dyshomeostasis of both ceramides and sphingosine-1-phosphate (S1P) in the brain has been implicated in aging-associated neurodegenerative disorders in humans. However, mechanisms that maintain the homeostasis of these bioactive sphingolipids in the brain remain unclear. Mouse alkaline ceramidase 3 (Acer3), which preferentially catalyzes the hydrolysis of C18:1-ceramide, a major unsaturated long-chain ceramide species in the brain, is upregulated with age in the mouse brain. Acer3 knockout causes an age-dependent accumulation of various ceramides and C18:1-monohexosylceramide and abolishes the age-related increase in the levels of sphingosine and S1P in the brain; thereby resulting in Purkinje cell degeneration in the cerebellum and deficits in motor coordination and balance. Our results indicate that Acer3 plays critically protective roles in controlling the homeostasis of various sphingolipids, including ceramides, sphingosine, S1P, and certain complex sphingolipids in the brain and protects Purkinje cells from premature degeneration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pgen.1005591DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4608763PMC
October 2015

Tumor Necrosis Factor-α (TNFα)-induced Ceramide Generation via Ceramide Synthases Regulates Loss of Focal Adhesion Kinase (FAK) and Programmed Cell Death.

J Biol Chem 2015 Oct 28;290(42):25356-73. Epub 2015 Aug 28.

From the Stony Brook Cancer Center, the Department of Medicine, and the Northport Veterans Affairs Medical Center, Northport, New York 11768

Ceramide synthases (CerS1-CerS6), which catalyze the N-acylation of the (dihydro)sphingosine backbone to produce (dihydro)ceramide in both the de novo and the salvage or recycling pathway of ceramide generation, have been implicated in the control of programmed cell death. However, the regulation of the de novo pathway compared with the salvage pathway is not fully understood. In the current study, we have found that late accumulation of multiple ceramide and dihydroceramide species in MCF-7 cells treated with TNFα occurred by up-regulation of both pathways of ceramide synthesis. Nevertheless, fumonisin B1 but not myriocin was able to protect from TNFα-induced cell death, suggesting that ceramide synthase activity is crucial for the progression of cell death and that the pool of ceramide involved derives from the salvage pathway rather than de novo biosynthesis. Furthermore, compared with control cells, TNFα-treated cells exhibited reduced focal adhesion kinase and subsequent plasma membrane permeabilization, which was blocked exclusively by fumonisin B1. In addition, exogenously added C6-ceramide mimicked the effects of TNFα that lead to cell death, which were inhibited by fumonisin B1. Knockdown of individual ceramide synthases identified CerS6 and its product C16-ceramide as the ceramide synthase isoform essential for the regulation of cell death. In summary, our data suggest a novel role for CerS6/C16-ceramide as an upstream effector of the loss of focal adhesion protein and plasma membrane permeabilization, via the activation of caspase-7, and identify the salvage pathway as the critical mechanism of ceramide generation that controls cell death.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M115.658658DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4646185PMC
October 2015

Activity of neutral and alkaline ceramidases on fluorogenic N-acylated coumarin-containing aminodiols.

J Lipid Res 2015 Oct 18;56(10):2019-28. Epub 2015 Aug 18.

Consejo Superior de Investigaciones Científicas (CSIC), Institut de Química Avançada de Catalunya (IQAC-CSIC), Research Unit on Bioactive Molecules (RUBAM), 08034 Barcelona, Spain.

Ceramidases catalyze the cleavage of ceramides into sphingosine and fatty acids. Previously, we reported on the use of the RBM14 fluorogenic ceramide analogs to determine acidic ceramidase activity. In this work, we investigated the activity of other amidohydrolases on RBM14 compounds. Both bacterial and human purified neutral ceramidases (NCs), as well as ectopically expressed mouse neutral ceramidase hydrolyzed RBM14 with different selectivity, depending on the N-acyl chain length. On the other hand, microsomes from alkaline ceramidase (ACER)3 knockdown cells were less competent at hydrolyzing RBM14C12, RBM12C14, and RBM14C16 than controls, while microsomes from ACER2 and ACER3 overexpressing cells showed no activity toward the RBM14 substrates. Conversely, N-acylethanolamine-hydrolyzing acid amidase (NAAA) overexpressing cells hydrolyzed RBM14C14 and RBM14C16 at acidic pH. Overall, NC, ACER3, and, to a lesser extent, NAAA hydrolyze fluorogenic RBM14 compounds. Although the selectivity of the substrates toward ceramidases can be modulated by the length of the N-acyl chain, none of them was specific for a particular enzyme. Despite the lack of specificity, these substrates should prove useful in library screening programs aimed at identifying potent and selective inhibitors for NC and ACER3.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1194/jlr.D061564DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4583077PMC
October 2015