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Biomedical subjects

James A Shayman

Publications and source records attributed to James A Shayman.

At least 19 recordsLinked to original sources
PubMed2006

Positional specificity of lysosomal phospholipase A2.

Lysosomal phospholipase A(2) (Lpla2) is highly expressed in alveolar macrophages and may mediate the phospholipid metabolism of surfactant. Studies on the properties of this phospholipase are consistent with the presence of both phospholipase A(1) and phospholipase A(2) activities. These activities were studied through the production of O-acyl compounds, produced by the transacylase activity of Lpla2. Liposomes containing POPC and N-acetylsphingosine (NAS) were incubated with the soluble fraction obtained from MDCK cells stably transfected with the mouse Lpla2 gene. Two 1-O-acyl-NASs, 1-O-palmitoyl-NAS and 1-O-oleoyl-NAS, were produced by Lpla2. The formation rate of 1-O-oleoyl-NAS was 2.5-fold that of 1-O-palmitoyl-NAS. When 1-oleoyl-2-palmitoyl-sn-glycero-3-phosphocholine (OPPC) was used, the formation rate of 1-O-oleoyl-NAS was 5-fold higher than that of 1-O-palmitoyl-NAS. Thus, Lpla2 can act on acyl groups at both sn-1 and sn-2 positions of POPC and OPPC. When 1-palmitoyl-2-unsaturated acyl-sn-glycero-3-phosphocholines were used as acyl donors, the transacylation of the acyl group from the sn-2 position to NAS was preferred to that of the palmitoyl group from the sn-1 position. An exception was observed for 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PAPC), for which the formation rate of 1-O-palmitoyl-NAS from PAPC was 4-fold greater than that of 1-O-arachidonoyl-NAS. Thus, Lpla2 has broad positional specificity for the sn-1 and sn-2 acyl groups in phosphatidylcholine and phosphatidylethanolamine.

Animals↗
PubMed2006

Alpha-galactosidase A deficiency leads to increased tissue fibrin deposition and thrombosis in mice homozygous for the factor V Leiden mutation.

BACKGROUND: Factor V Leiden (FVL) is a common genetic risk factor for vascular thrombosis in humans. Fabry disease, an X-linked lysosomal storage disorder attributable to alpha-galactosidase A (GLA) deficiency, is associated with premature vascular events that may be thrombotic in nature. METHODS: To examine a potential interaction between FvL and Gla deficiency in vivo, we analyzed tissue fibrin deposition in mice carrying combined mutations in FvL and Gla. Gla deficiency markedly increased tissue fibrin deposition in mice carrying the FvL mutation (0.33+/-0.03%; n=7) compared with FvL mutation (0.14+/-0.02%; n=10; P<0.0005). CONCLUSIONS: These observations demonstrate a synergistic interaction between Gla deficiency and FvL toward tissue fibrin deposition in mice. Concomitant mutations in these genes may increase the penetrance of vascular thrombotic events in humans.

Animals↗
PubMed2006

Sphingosine kinase 1-mediated inhibition of Fas death signaling in rheumatoid arthritis B lymphoblastoid cells.

OBJECTIVE: It is becoming increasingly apparent that B cells play an important role in the pathogenesis of rheumatoid arthritis (RA). Due to the scarcity of B cells in RA, it has been technically difficult to functionally characterize B cell apoptosis in this disease. As a necessary first step to identify candidate aberrations, we investigated Fas-mediated signaling events in immortalized peripheral blood B lymphoblastoid cell lines (LCLs) from patients with RA and controls. METHODS: Cell death was determined by the MTS assay, and apoptosis was detected by the TUNEL assay and DNA laddering. Proteolytic activation of caspase 3 was determined by immunoblotting, and its enzymatic activity was determined by a fluorometric technique. Messenger RNA (mRNA) expression was quantified by real-time polymerase chain reaction (PCR) analysis. The functional role of sphingosine kinase (SPHK) was determined by measuring its enzymatic activity, by quantifying the levels of its product, sphingosine 1-phosphate (S1P), and by investigating the ability of the SPHK inhibitor N,N-dimethylsphingosine and isozyme-specific small interfering RNA (siRNA) oligonucleotides to reverse signaling aberrations. RESULTS: LCLs from patients with RA displayed disease-specific Fas-mediated signal transduction impairment with consequent resistance to cell death. RA LCLs displayed high constitutive SPHK activity and increased levels of S1P. Real-time PCR analysis showed higher SPHK-1 mRNA expression levels in RA patients compared with paired controls. Increased SPHK-1 (but not SPHK-2) mRNA levels were observed in synovial tissue from RA patients. Competitive inhibitors of SPHK reversed the resistance of RA LCLs to Fas-induced apoptosis. Additionally, resistance to Fas-mediated signaling was reversed by siRNA oligonucleotides specific for SPHK-1 but not by oligonucleotides specific for SPHK-2. CONCLUSION: These findings demonstrate disease-specific resistance to Fas-mediated death signaling in patients with RA and implicate increased SPHK-1 activity as the cause of this aberration.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Lysosomal phospholipase A2 and phospholipidosis.

A lysosomal phospholipase A2, LPLA2, was recently characterized and shown to have substrate specificity for phosphatidylcholine and phosphatidylethanolamine. LPLA2 is ubiquitously expressed but is most highly expressed in alveolar macrophages. Double conditional gene targeting was employed to elucidate the function of LPLA2. LPLA2-deficient mice (Lpla2-/-) were generated by the systemic deletion of exon 5 of the Lpla2 gene, which encodes the lipase motif essential for the phospholipase A2 activity. The survival of the Lpla2-/- mice was normal. Lpla2-/- mouse mating pairs yielded normal litter sizes, indicating that the gene deficiency did not impair fertility or fecundity. Alveolar macrophages from wild-type but not Lpla2-/- mice readily degraded radiolabeled phosphatidylcholine. A marked accumulation of phospholipids, in particular phosphatidylethanolamine and phosphatidylcholine, was found in the alveolar macrophages, the peritoneal macrophages, and the spleens of Lpla2-/- mice. By 1 year of age, Lpla2-/- mice demonstrated marked splenomegaly and increased lung surfactant phospholipid levels. Ultrastructural examination of Lpla2-/- mouse alveolar and peritoneal macrophages revealed the appearance of foam cells with lamellar inclusion bodies, a hallmark of cellular phospholipidosis. Thus, a deficiency of lysosomal phospholipase A2 results in foam cell formation, surfactant lipid accumulation, splenomegaly, and phospholipidosis in mice.

Alleles↗
PubMed2005

Structure and function of lysosomal phospholipase A2: identification of the catalytic triad and the role of cysteine residues.

Lysosomal phospholipase A2 (LPLA2) is an acidic phospholipase that is highly expressed in alveolar macrophages and that may play a role in the catabolism of pulmonary surfactant. The primary structure found in LCAT is conserved in LPLA2, including three amino acid residues potentially required for catalytic activity and four cysteine residues. LPLA2 activity was measured in COS-7 cells transfected with c-myc-conjugated mouse LPLA2 (mLPLA2) or mutated LPLA2. Single alanine substitutions in the catalytic triad resulted in the elimination of LPLA2 activity. Four cysteine residues (C65, C89, C330, and C371), conserved between LPLA2 and LCAT, were replaced with alanine. Quadruple mutations at C65, C89, C330, and C371, double mutations at C65 and C89, and a single mutation at C65 or C89 resulted in the elimination of activity. Double mutations at C330 and C371 and a single mutation at C330 or C371 resulted in a partial reduction of activity. Thus, the presence of a disulfide bond between C330 and C371 is not required for LPLA2 activity. We propose that one disulfide bond between C65 and C89 and free cysteine residues at C330 and C371 and the triad, serine-198, aspartic acid-360, and histidine-392, are required for the full expression of mLPLA2 activity.

Alanine↗
PubMed2005

An in vitro model of Fabry disease.

Fabry disease is an X-linked inherited loss of alpha-galactosidase A (alpha-Gal A). Affected patients experience complications that include neuropathy, renal failure, and cardiovascular disease. Although the genetic and biochemical basis of this sphingolipidosis is well studied, the basis for the vascular disease remains poorly understood. In an attempt to create a suitable in vitro model of this disease, conditions for the growth of primary cultures of aortic endothelial cells from wild-type and alpha-Gal A -/0 mice were established. The cultured cells demonstrated CD-31 expression by flow cytometry and LDL binding by immunofluorescence. The glycolipid expression patterns were compared between wild-type and alpha-Gal A null cells. Importantly, cells from alpha-Gal A -/0 mice but not alpha-Gal A +/0 mice expressed high levels of the globo-series glycosphingolipid globotriaosylceramide (Gb3). The age-dependent elevation in Gb3 was measured. By 4 mo of age, alpha-Gal A -/0 mouse aortic endothelial cells achieved their peak Gb3 levels. The ability to lower Gb3 levels pharmacologically was assessed next. The glucosylceramide synthase inhibitor ethylenedioxyphenyl-P4 significantly lowered but did not eliminate Gb3 levels by 96 h of treatment. Gb3 synthesis was completely blocked as measured by [14C]galactose labeling. Recombinant alpha-Gal A more significantly lowered Gb3 levels by 48 h but had a more limited effect on de novo synthesis. Together, both agents eliminated detectable Gb3. In summary, primary cultures of aortic endothelial cells from Fabry mice retain the phenotype of elevated globo-series glycosphingolipids. These cells provide a useful model for comparing pharmacologic agents used for glycolipid reduction.

Animals↗
PubMed2005

Phagocytic signaling molecules in lipid rafts of COS-1 cells transfected with FcgammaRIIA.

COS-1 cells bearing FcgammaRIIA were used as a model to demonstrate co-localization of several enzymes previously shown to regulate neutrophil phagocytosis. In COS-1 cells, phospholipase D (PLD) in the membrane fraction was activated during phagocytosis. PLD was found almost exclusively in lipid rafts, along with RhoA and ARF1. Protein kinase C-delta (PKCdelta) and Raf-1 translocated to lipid rafts. In neutrophils, ceramide levels increase during phagocytosis, indicating that FcgammaRIIA engagement initiates ceramide generation. Applying this model, we transfected COS-1 cells with FcgammaRIIA that had been mutated in the ITAM region, rendering them unable to ingest particles. When the mutant receptors were engaged, ceramide was generated and MAPK was activated normally, thus these processes did not require actual ingestion of particles. These results indicate that signaling proteins for phagocytosis are either constitutively present in, or are recruited to, lipid rafts where they are readily available to activate one another.

Animals↗
PubMed2005

Ceramide 1-phosphate, a mediator of phagocytosis.

The agonist-stimulated metabolism of membrane lipids produces potent second messengers that regulate phagocytosis. We studied whether human ceramide kinase (hCERK) activity and ceramide 1-phosphate formation could lead to enhanced phagocytosis through a mechanism involving modulation of the membrane-structural order parameter. hCERK was stably transfected into COS-1 cells that were stably transfected with the FcgammaRIIA receptor. hCERK-transfected cells displayed a significant increase in phagocytic index in association with increased ceramide kinase activation and translocation to lipid rafts after activation with opsonized erythrocytes. When challenged with opsonized erythrocytes, hCERK-transfected cells increased phagocytosis by 1.5-fold compared with vector control and simultaneously increased ceramide 1-phosphate levels 2-fold compared with vector and unstimulated control cells. Control and hCERK-transfected cells were subjected to cellular fractionation. Utilizing an antibody against hCERK, we observed that CERK translocates during activation from the cytosol to a lipid raft fraction. The plasma membrane-structural order parameter of the transfectants was measured by labeling cells with Laurdan. Cells transfected with hCERK showed a higher liquid crystalline order than control cells with stimulation, conditions that are favorable for the promotion of membrane fusion at the sites of phagocytosis. The change in the structural order parameter of the lipid rafts probably contributes to phagocytosis by promoting phagosome formation.

2-Naphthylamine↗
PubMed2005

Alpha-galactosidase A deficiency accelerates atherosclerosis in mice with apolipoprotein E deficiency.

BACKGROUND: Alpha-galactosidase A (Gla) deficiency leads to widespread tissue accumulation of neutral glycosphingolipids and is associated with premature vascular complications such as myocardial infarction and stroke. Glycosphingolipids have been shown to accumulate in human atherosclerotic lesions, although their role in atherogenesis is unclear. METHODS AND RESULTS: To determine whether Gla affects the progression of atherosclerosis, mice were generated with combined deficiencies of apolipoprotein E and Gla. At 45 weeks of age, Gla-deficient mice had developed more atherosclerosis than mice with normal Gla expression (25.1+/-14.0 versus 12.3+/-9.3 mm2 of total lesion area, P<0.02). This increase in atherosclerosis was associated with the presence of increased Gb3, enhanced inducible nitric oxide synthase expression, and increased nitrotyrosine staining. CONCLUSIONS: These findings suggest that deficiency of Gla leads to increased inducible nitric oxide synthase expression and accelerated atherosclerosis.

Animals↗
PubMed2004

Lysosomal phospholipase A2 is selectively expressed in alveolar macrophages.

Lung surfactant is the surface-active agent comprised of phospholipids and proteins that lines pulmonary alveoli. Surfactant stabilizes the alveolar volume by reducing surface tension. Previously, we identified a lysosomal phospholipase A2, termed LPLA2, with specificity toward phosphatidylcholine and phosphatidylethanolamine. The phospholipase is localized to lysosomes, is calcium-independent, has an acidic pH optimum, and transacylates ceramide. Here, we demonstrate that LPLA2 is selectively expressed in alveolar macrophages but not in peritoneal macrophages, peripheral blood monocytes, or other tissues. Other macrophage-associated phospholipase A2s do not show a comparable distribution. LPLA2 is of high specific activity and recognizes disaturated phosphatidylcholine as a substrate. The lysosomal phospholipase A2 activity is six times lower in alveolar macrophages from mice with a targeted deletion of the granulocyte macrophage colony-stimulating factor (GM-CSF), a model of impaired surfactant catabolism, compared with those from wild-type mice. However, LPLA2 activity and protein levels are measured in GM-CSF null mice in which GM-CSF is expressed as a transgene under the control of the surfactant protein C promoter. Thus LPLA2 may be a major enzyme of pulmonary surfactant phospholipid degradation by alveolar macrophages and may be deficient in disorders of surfactant metabolism.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed2004

Induction of lysosomal phospholipase A2 through the retinoid X receptor in THP-1 cells.

An acidic phospholipase A(2) (LPLA(2)) was recently purified and cloned. THP-1 cells were used to characterize the gene induction of LPLA(2). THP-1 cells were stimulated with several differentiation agents. The LPLA(2) mRNA and activity increased in cells treated with phorbol ester but not with vitamin D3, interferon-gamma, or granulocyte macrophage colony-stimulating factor. All-trans-retinoic acid enhanced mRNA expression and enzyme activity in a dose- and time-dependent manner. The natural 9-cis and 13-cis isomers of retinoic acid enhanced transcription and activity. Two classes of nuclear receptors, the retinoic acid receptor (RAR) and the retinoid X receptor (RXR), mediate retinoic acid signaling. Specific RAR and RXR agonists were used to identify the nuclear receptor responsible for LPLA(2) induction by retinoic acid. Treatment with the RAR agonist 4-[E-2-tetrahydro-5,5,8,8-tetra-methyl-2-naphthalenyl]1-propenyl benzoic acid (TTNPB) resulted in a small and statistically significant increase of the mRNA expression and activity of LPLA(2). The RXR agonist methoprene acid worked as well as all-trans-retinoic acid at increasing both mRNA and enzyme activity. The methoprene acid and TTNPB effects were not synergistic. The peroxisome proliferator-activated receptor gamma agonists 15-deoxy-Delta(12,14)-prostaglandin J(2) and troglitazone failed to induce LPLA(2) activity and mRNA. Thus, an RXR-dependent pathway controls LPLA(2) gene activation by retinoic acid in THP-1 cells.

Cell Differentiation↗
PubMed2004

A turn in the road: How studies on the pharmacology of glucosylceramide synthase inhibitors led to the identification of a lysosomal phospholipase A2 with ceramide transacylase activity.

A series of inhibitors of glucosylceramide synthesis, the PDMP based family of compounds, has been developed as a tool for the study of sphingolipid biochemistry and biology. During the course of developing more active glucosylceramide synthase inhibitors, we identified a second site of inhibitory activity for PDMP and its structural homologues that accounted for the ability of the inhibitors to raise cell and tissue ceramide levels. This inhibitory activity was directed against a previously unknown pathway for ceramide metabolism, viz. the formation of 1- O -acylceramide. In this pathway the addition of a fatty acyl group to the primary hydroxyl of ceramide occurs through a transacylation with either phosphatidylethanolamine or phosphatidylcholine as a substrate. However, both in the absence and presence of ceramide, water serves as an acceptor for the fatty acid. Thus the enzyme may be considered to be a phospholipase A2. The enzyme is unique in that it has an acidic pH optimum and is localized to lysosomes by cell fractionation. More recently, the 1- O -acylceramide synthase has been purified, sequenced, and cloned. This phospholipase A2 was discovered to be structurally homologous to lecithin cholesterol acyltransferase (LCAT). However, this phospholipase A2 does not recognize cholesterol and lacks the defined lipoprotein-binding domain present in LCAT. We now refer to this enzyme as lysosomal phospholipase A2 (LPLA2). Although acidic phospholipase A2 activities have been previously identified, LPLA2 appears to be the first lysosomal PLA2 to have been sequenced. This new phospholipase A2 lacks an obvious and proven biological function.

Acylation↗
PubMed2003

Ceramide inhibition of phospholipase D and its relationship to RhoA and ARF1 translocation in GTP gamma S-stimulated polymorphonuclear leukocytes.

Phospholipase D (PLD) regulates the polymorphonuclear leukocyte (PMN) functions of phagocytosis, degranulation, and oxidant production. Ceramide inhibition of PLD suppresses PMN function. In streptolysin O-permeabilized PMNs, PLD was directly activated by guanosine 5'-[gamma-thio]triphosphate (GTP gamma S) stimulation of adenosine diphosphate (ADP)-ribosylation factor (ARF) and Rho, stimulating release of lactoferrin from specific granules of permeabilized PMNs; PLD activation and degranulation were inhibited by C2-ceramide but not dihydro-C2-ceramide. To investigate the mechanism of ceramide's inhibitory effect on PLD, we used a cell-free system to examine PLD activity and translocation from cytosol to plasma membrane of ARF, protein kinase C (PKC)alpha and beta, and RhoA, all of which can activate PLD. GTP gamma S-activated cytosol stimulated PLD activity and translocation of ARF, PKC alpha and beta, and RhoA when recombined with cell membranes. Prior incubation of PMNs with 10 microM C2-ceramide inhibited PLD activity and RhoA translocation, but not ARF1, ARF6, PKC alpha, or PKC beta translocation. However, in intact PMNs stimulated with N-formyl-1-methionyl-1-leucyl-1-phenylalamine (FMLP) or permeabilized PMNs stimulated with GTP gamma S, C2-ceramide did not inhibit RhoA translocation. Exogenous RhoA did not restore ceramide-inhibited PLD activity but bound to membranes despite ceramide treatment. These observations suggest that, although ceramide may affect RhoA in some systems, ceramide inhibits PLD through another mechanism, perhaps related to the ability of ceramide to inhibit phosphatidylinositol-bisphosphate (PIP2) interaction with PLD.

ADP-Ribosylation Factor 1↗
PubMed2003

Src kinase mediates the regulation of phospholipase C-gamma activity by glycosphingolipids.

Glucosylceramide-based glycosphingolipids have been previously demonstrated to regulate negatively the formation of inositol 1,4,5-trisphosphate by phospholipase C-gamma1. In the present study, the depletion of endogenous glucosylceramide by D-t-EtDO-P4 in cultured ECV304 cells induced autophosphorylation of Src kinase at tyrosine residue 418 within the catalytic loop and dephosphorylation of Src kinase at tyrosine residues 529 within the carboxyl-terminal regulatory region. Phosphotransferase activities of Src kinase were also induced in the glucosylceramide-depleted cells. c-Src kinase activity and phosphorylations at Src Tyr-418 and epidermal growth factor (EGF) receptor Tyr-1068 were significantly enhanced by bradykinin in response to 100 nm D-t-EtDO-P4 compared with control cells. The phosphorylation and dephosphorylation on Tyr-418 and Tyr-529 residues of c-Src were reversed by treatment of 4-amino-5-(4-chlorophenyl)-7-t-butyl(pyrazolo)[3,4-d]pyrimidine (PP2), an inhibitor of Src kinase, in control cells. Glucosylceramide-depleted cells resisted treatment with PP2, and both phosphorylation of Tyr-418 and dephosphorylation of Tyr-529 induced by depletion of glucosylceramide were maintained. Compared with untreated cells, tyrosine phosphorylation of phospholipase C-gamma1 was enhanced by EGF stimulation in glucosylceramide-depleted cells, associated with enhanced tyrosine phosphorylation of the EGF receptor at Tyr-1068 and Tyr-1086 stimulated by EGF. The Src inhibitor, PP2, significantly blocked EGF-induced tyrosine phosphorylation of phospholipase C-gamma1 in control cells, whereas in glucosylceramide-depleted cells, suppression of Src kinase activity by PP2 toward EGF-induced tyrosine phosphorylation of phospholipase C-gamma1 was less significant. Thus the activation of Src kinase by depletion of glucosylceramide-based glycosphingolipids in cultured ECV304 cells is a critical up-stream event in the activation of phospholipase C-gamma1.

Amino Acid Sequence↗
PubMed2003

Fabry disease in mice is associated with age-dependent susceptibility to vascular thrombosis.

Fabry disease is an X-linked lysosomal storage disorder due to deficiency of alpha-galactosidase A (GLA) activity that results in the widespread accumulation of neutral glycosphingolipids. Renal failure, neuropathy, premature myocardial infarction, and stroke occur in patients with this condition primarily due to deposition of glycosphingolipids in vascular endothelial cells. The clinical consequences of Fabry disease suggest that vascular thrombosis may play a prominent role in the pathogenesis of this disease; however, the vasculopathy associated with Fabry disease has not been extensively studied. To determine if mice genetically deficient in Gla are susceptible to vascular thrombosis, a photochemical carotid injury model was used to induce occlusive thrombosis. In this model, Gla-/0 mice displayed a progressive age-dependent shortening of the time to occlusive thrombosis after vascular injury that correlated with progressive accumulation of globotriasylceramide (Gb3) in the arterial wall. Bone marrow transplantation from Gla-/0 to Gla+/0 mice and from Gla+/0 to Gla-/0 mice did not change the thrombotic phenotype of the host. These studies reveal a potent vascular prothrombotic phenotype in Gla-deficient mice and suggest that antithrombotic therapies as well as therapies designed to reduce the vascular accumulation of Gb3 may have beneficial effects on thrombotic complications in patients with Fabry disease.

Aging↗
PubMed2002

Enhanced phagocytosis through inhibition of de novo ceramide synthesis.

Fcgamma receptors are important mediators of the binding of IgG to and induction of phagocytosis in neutrophils. COS-1 cells provide a potentially useful model for studying these receptors because transfection with the FcgammaRIIA renders these cells phagocytic. During FcgammaRIIA-mediated phagocytosis in COS-1 cells, endogenous ceramide levels increased 52% by 20 min (p < 0.01). Phospholipase D activity increased by 62% (p < 0.01). Correspondingly, the phagocytic index increased by 3.7-fold by 20 min. Two inhibitors of ceramide formation were used to assess the consequences of reduced ceramide generation. l-Cycloserine, an inhibitor that blocks serine palmitoyltransferase activity, lowered both sphingosine and ceramide levels. Under these conditions, the phagocytic index increased 100% in the presence of 2 mm l-cycloserine. The formation of ceramide resulting from the N-acylation of dihydrosphingosine or sphingosine by ceramide synthase is inhibited by the fungal toxin fumonisin B(1). When cells were treated with 5-50 microm fumonisin B(1), the cellular level of ceramide decreased in a concentration-dependent manner, while simultaneously the phagocytic index increased by 52%. Concomitantly, three indirect measures of FcgammaRIIA activity were altered with the fall in ceramide levels. Syk phosphorylation, phospholipase D activity, and mitogen-activated protein (MAP) kinase phosphorylation were increased at 30 min. When Syk phosphorylation was blocked with piceatannol and cells were similarly challenged, phosphatidylinositol 3-kinase activation was blocked, but no changes in either ceramide accumulation or MAP kinase activation were observed. Ceramide formation and MAP kinase activation are therefore not dependent on Syk kinase activity in this system. These results indicate that COS-1 cells provide a useful model for the recapitulation of sphingolipid signaling in the study of phagocytosis. Ceramide formed by de novo synthesis may represent an important mechanism in the regulation of phagocytosis.

Animals↗
PubMed2002

Disruption of the glucosylceramide biosynthetic pathway in Aspergillus nidulans and Aspergillus fumigatus by inhibitors of UDP-Glc:ceramide glucosyltransferase strongly affects spore germination, cell cycle, and hyphal growth.

The opportunistic mycopathogen Aspergillus fumigatus expresses both glucosylceramide and galactosylceramide (GlcCer and GalCer), but their functional significance in Aspergillus species is unknown. We here identified and characterized a GlcCer from Aspergillus nidulans, a non-pathogenic model fungus. Involvement of GlcCer in fungal development was tested on both species using a family of compounds known to inhibit GlcCer synthase in mammals. Two analogs, D-threo-1-phenyl-2-palmitoyl-3-pyrrolidinopropanol (P4) and D-threo-3',4'-ethylenedioxy-P4, strongly inhibited germination and hyphal growth. Neutral lipids from A. fumigatus cultured in the presence of these inhibitors displayed a significantly reduced GlcCer/GalCer ratio. These results suggest that synthesis of GlcCer is essential for normal development of A. fumigatus and A. nidulans.

Antifungal Agents↗