Search

Long-term results following solid organ transplantation do not mirror the excellent short-term results achieved in recent decades. It is therefore clear that current immunosuppressive maintenance protocols primarily addressing the adaptive immune system no longer meet the required clinical need. Identification of novel targets addressing this shortcoming is urgently needed. There is a growing interest in better understanding the role of the innate immune system in this context. In this review, we focus on macrophages, which are known to prominently infiltrate allografts and, during allograft rejection, to be involved in the surge of the adaptive immune response by expression of pro-inflammatory cytokines and direct cytotoxicity. However, this active participation is janus-faced and unspecific targeting of macrophages may not consider the different subtypes involved. Under this premise, we give an overview on macrophages, including their origins, plasticity, and important markers. We then briefly describe their role in acute allograft rejection, which ranges from sustaining injury to promoting tolerance, as well as the impact of maintenance immunosuppressants on macrophages. Finally, we discuss the observed immunosuppressive role of the vitamin-like compound tetrahydrobiopterin and the recent findings that suggest the innate immune system, particularly macrophages, as its target. [ABSTRACT FROM AUTHOR]

Plasmalogens are an abundant class of glycerophospholipids in the mammalian body, with special occurrence in the brain and in immune cell membranes. Plasmanylethanolamine desaturase (PEDS1) is the final enzyme of plasmalogen biosynthesis, which introduces the characteristic 1-O-alk-1′-enyl double bond. The recent sequence identification of PEDS1 as transmembrane protein 189 showed that its protein sequence is related to a special class of plant desaturases (FAD4), with whom it shares a motif of 8 conserved histidines, which are essential for the enzymatic activity. In the present work, we wanted to gain more insight into the sequence–function relationship of this enzyme and mutated to alanine additional 28 amino acid residues of murine plasmanylethanolamine desaturase including those 20 residues, which are also totally conserved—in addition to the eight-histidine-motif—among the animal PEDS1 and plant FAD4 plant desaturases. We measured the enzymatic activity by transient transfection of tagged murine PEDS1 expression clones to a PEDS1-deficient human HAP1 cell line by monitoring of labeled plasmalogens formed from supplemented 1-O-pyrenedecyl-sn-glycerol in relation to recombinant protein expression. Surprisingly, only a single mutation, namely aspartate 100, led to a total loss of PEDS1 activity. The second strongest impact on enzymatic activity had mutation of phenylalanine 118, leaving only 6% residual activity. A structural model obtained by homology modelling to available structures of stearoyl-CoA reductase predicted that this aspartate 100 residue interacts with histidine 96, and phenylalanine 118 interacts with histidine 187, both being essential histidines assumed to be involved in the coordination of the di-metal center of the enzyme. [ABSTRACT FROM AUTHOR]

Depletion of the enzyme cofactor, tetrahydrobiopterin (BH4), in T-cells was shown to prevent their proliferation upon receptor stimulation in models of allergic inflammation in mice, suggesting that BH4 drives autoimmunity. Hence, the clinically available BH4 drug (sapropterin) might increase the risk of autoimmune diseases. The present study assessed the implications for multiple sclerosis (MS) as an exemplary CNS autoimmune disease. Plasma levels of biopterin were persistently low in MS patients and tended to be lower with high Expanded Disability Status Scale (EDSS). Instead, the bypass product, neopterin, was increased. The deregulation suggested that BH4 replenishment might further drive the immune response or beneficially restore the BH4 balances. To answer this question, mice were treated with sapropterin in immunization-evoked autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. Sapropterin-treated mice had higher EAE disease scores associated with higher numbers of T-cells infiltrating the spinal cord, but normal T-cell subpopulations in spleen and blood. Mechanistically, sapropterin treatment was associated with increased plasma levels of long-chain ceramides and low levels of the poly-unsaturated fatty acid, linolenic acid (FA18:3). These lipid changes are known to contribute to disruptions of the blood–brain barrier in EAE mice. Indeed, RNA data analyses revealed upregulations of genes involved in ceramide synthesis in brain endothelial cells of EAE mice (LASS6/CERS6, LASS3/CERS3, UGCG, ELOVL6, and ELOVL4). The results support the view that BH4 fortifies autoimmune CNS disease, mechanistically involving lipid deregulations that are known to contribute to the EAE pathology. [ABSTRACT FROM AUTHOR]

Background: Genome editing in mice using either classical approaches like homologous recombination or CRISPR/Cas9 has been reported to harbor off target effects (insertion/deletion, frame shifts or gene segment duplications) that lead to mutations not only in close proximity to the target site but also outside. Only the genomes of few engineered mouse strains have been sequenced. Since the role of the ether-lipid cleaving enzyme alkylglycerol monooxygenase (AGMO) in physiology and pathophysiology remains enigmatic, we created a knockout mouse model for AGMO using EUCOMM stem cells but unforeseen genotyping issues that did not agree with Mendelian distribution and enzyme activity data prompted an in-depth genomic validation of the mouse model. Results: We report a gene segment tandem duplication event that occurred during the generation of an Agmo knockout-first allele by homologous recombination. Only low homology was seen between the breakpoints. While a single copy of the recombinant 18 kb cassette was integrated correctly around exon 2 of the Agmo gene, whole genome nanopore sequencing revealed a 94 kb duplication in the Agmo locus that contains Agmo wild-type exons 1–3. The duplication fooled genotyping by routine PCR, but could be resolved using qPCR-based genotyping, targeted locus amplification sequencing and nanopore sequencing. Despite this event, this Agmo knockout mouse model lacks AGMO enzyme activity and can therefore be used to study its physiological role. Conclusions: A duplication event occurred at the exact locus of the homologous recombination and was not detected by conventional quality control filters such as FISH or long-range PCR over the recombination sites. Nanopore sequencing provides a cost convenient method to detect such underrated off-target effects, suggesting its use for additional quality assessment of gene editing in mice and also other model organisms. [ABSTRACT FROM AUTHOR]

A significant fraction of the glycerophospholipids in the human body is composed of plasmalogens, particularly in the brain, cardiac, and immune cell membranes. A decline in these lipids has been observed in such diseases as Alzheimer’s and chronic obstructive pulmonary disease. Plasmalogens contain a characteristic 1-O-alk-1′-enyl ether (vinyl ether) double bond that confers special biophysical, biochemical, and chemical properties to these lipids. However, the genetics of their biosynthesis is not fully understood, since no gene has been identified that encodes plasmanylethanolamine desaturase (E.C. 1.14.99.19), the enzyme introducing the crucial alk-1′-enyl ether double bond. The present work identifies this gene as transmembrane protein 189 (TMEM189). Inactivation of the TMEM189 gene in human HAP1 cells led to a total loss of plasmanylethanolamine desaturase activity, strongly decreased plasmalogen levels, and accumulation of plasmanylethanolamine substrates and resulted in an inability of these cells to form labeled plasmalogens from labeled alkylglycerols. Transient expression of TMEM189 protein, but not of other selected desaturases, recovered this deficit. TMEM189 proteins contain a conserved protein motif (pfam10520) with eight conserved histidines that is shared by an alternative type of plant desaturase but not by other mammalian proteins. Each of these histidines is essential for plasmanylethanolamine desaturase activity. Mice homozygous for an inactivated Tmem189 gene lacked plasmanylethanolamine desaturase activity and had dramatically lowered plasmalogen levels in their tissues. These results assign the TMEM189 gene to plasmanylethanolamine desaturase and suggest that the previously characterized phenotype of Tmem189-deficient mice may be caused by a lack of plasmalogens. [ABSTRACT FROM AUTHOR]

Alkylglycerol monooxygenase (AGMO) is the only enzyme known to cleave the O-alkyl bonds of ether lipids (alkylglycerols) which are essential components of cell membranes. A homozygous frameshift variant [p.(Glu324LysfsTer12)] in AGMO has recently been reported in two male siblings with syndromic microcephaly. In this study, we identified rare nonsense, in frame deletion, and missense biallelic variants in AGMO in two unrelated individuals with neurodevelopmental disabilities. We assessed the activity of seven disease associated AGMO variants including the four variants identified in our two affected individuals expressed in human embryonic kidney (HEK293T) cells. We demonstrated significantly diminished enzyme activity for all disease-associated variants, supporting the mechanism as decreased AGMO activity. Future mechanistic studies are necessary to understand how decreased AGMO activity leads to the neurologic manifestations. [ABSTRACT FROM AUTHOR]

GTP cyclohydrolase (GCH1) governs de novo synthesis of the enzyme cofactor, tetrahydrobiopterin (BH4), which is essential for biogenic amine production, bioactive lipid metabolism and redox coupling of nitric oxide synthases. Overproduction of BH4 via upregulation of GCH1 in sensory neurons is associated with nociceptive hypersensitivity in rodents, and neuron‐specific GCH1 deletion normalizes nociception. The translational relevance is revealed by protective polymorphisms of GCH1 in humans, which are associated with a reduced chronic pain. Because myeloid cells constitute a major non‐neuronal source of BH4 that may contribute to BH4‐dependent phenotypes, we studied here the contribution of myeloid‐derived BH4 to pain and itch in lysozyme M Cre‐mediated GCH1 knockout (LysM‐GCH1−/−) and overexpressing mice (LysM‐GCH1‐HA). Unexpectedly, knockout or overexpression in myeloid cells had no effect on nociceptive behaviour, but LysM‐driven GCH1 knockout reduced, and its overexpression increased the scratching response in Compound 48/80 and hydroxychloroquine‐evoked itch models, which involve histamine and non‐histamine dependent signalling pathways. Mechanistically, GCH1 overexpression increased BH4, nitric oxide and hydrogen peroxide, and these changes were associated with increased release of histamine and serotonin and degranulation of mast cells. LysM‐driven GCH1 knockout had opposite effects, and pharmacologic inhibition of GCH1 provided even stronger itch suppression. Inversely, intradermal BH4 provoked scratching behaviour in vivo and BH4 evoked an influx of calcium in sensory neurons. Together, these loss‐ and gain‐of‐function experiments suggest that itch in mice is contributed by BH4 release plus BH4‐driven mediator release from myeloid immune cells, which leads to activation of itch‐responsive sensory neurons. [ABSTRACT FROM AUTHOR]

Current strategies used to quantitatively describe the biological diversity of lipids by mass spectrometry are often limited in assessing the exact structural variability of individual molecular species in detail. A major challenge is represented by the extensive isobaric overlap present among lipids' hampering their accurate identification. This is especially true for cardiolipins, a mitochondria-specific class of phospholipids, which are functionally involved in many cellular functions, including energy metabolism, cristae structure, and apoptosis. Substituted with four fatty acyl side chains, cardiolipins offer a particularly high potential to achieve complex mixtures of molecular species. Here, we demonstrate how systematically generated high-performance liquid chromatography-mass spectral data can be utilized in a mathematical structural modeling approach, to comprehensively analyze and characterize the molecular diversity of mitochondrial cardio-lipin compositions in cell culture and disease models, cardiolipin modulation experiments, and a broad variety of frequently studied model organisms. [ABSTRACT FROM AUTHOR]

Tetrahydrobiopterin is a cofactor synthesized from GTP with well-known roles in enzymatic nitric oxide synthesis and aromatic amino acid hydroxylation. It is used to treat mild forms of phenylketonuria. Less is known about the role of tetrahydrobiopterin in lipid metabolism, although it is essential for irreversible ether lipid cleavage by alkylglycerol monooxygenase. Here we found intracellular alkylglycerol monooxygenase activity to be an important regulator of alkylglycerol metabolism in intact murine RAW264.7 macrophage-like cells. Alkylglycerol monooxygenase was expressed and active also in primary mouse bone marrowderived monocytes and "alternatively activated" M2 macrophages obtained by interleukin 4 treatment, but almost missing in M1 macrophages obtained by IFN-γ and lipopolysaccharide treatment. The cellular lipidome of RAW264.7 was markedly changed in a parallel way by modulation of alkylglycerol monooxygenase expression and of tetrahydrobiopterin biosynthesis affecting not only various ether lipid species upstream of alkylglycerol monooxygenase but also other more complex lipids including glycosylated ceramides and cardiolipins, which have no direct connection to ether lipid pathways. Alkylglycerol monooxygenase activity manipulation modulated the IFN-γ/lipopolysaccharide-induced expression of inducible nitric oxide synthase, interleukin-1β, and interleukin 1 receptor antagonist but not transforming growth factor β1, suggesting that alkylglycerolmonooxygenase activity affects IFN-γ/lipopolysaccharide signaling. Our results demonstrate a central role of tetrahydrobiopterin and alkylglycerol monooxygenase in ether lipid metabolism of murine macrophages and reveal that alteration of alkylglycerol monooxygenase activity has a profound impact on the lipidome also beyond the class of ether lipids. [ABSTRACT FROM AUTHOR]

Objective: Aim of this study was to identify the nitric oxide synthase (NOS) isoform involved in early microcirculatory derangements following solid organ transplantation. Background: Tetrahydrobiopterin donor treatment has been shown to specifically attenuate these derangements following pancreas transplantation, and tetrahydrobiopterin-mediated protective effects to rely on its NOS-cofactor activity, rather than on its antioxidant capacity. However, the NOS-isoform mainly involved in this process has still to be defined. Methods: Using a murine pancreas transplantation model, grafts lacking one of the three NOS-isoforms were compared to grafts from wild-type controls. Donors were treated with either tetrahydrobiopterin or remained untreated. All grafts were subjected to 16 h cold ischemia time and transplanted into wild-type recipients. Following 4 h graft reperfusion, microcirculation was analysed by confocal intravital fluorescence microscopy. Recipient survival was monitored for 50 days. Results: Transplantation of the pancreas from untreated wild-type donor mice resulted in microcirculatory damage of the transplanted graft and no recipient survived more than 72 h. Transplanting grafts from untreated donor mice lacking either endothelial or inducible NOS led to similar outcomes. In contrast, donor treatment with tetrahydrobiopterin prevented microcirculatory breakdown enabling long-term survival. Sole exception was transplantation of grafts from untreated donor mice lacking neuronal NOS. It resulted in intact microvascular structure and long-term recipient survival, either if donor mice were untreated or treated with tetrahydrobiopterin. Conclusion: We demonstrate for the first time the crucial involvement of neuronal NOS in early microcirculatory derangements following solid organ transplantation. In this model, protective effects of tetrahydrobiopterin are mediated by targeting this isoform. [ABSTRACT FROM AUTHOR]

Alkylglycerol monooxygenase (E.C. 1.14.16.5), also called glyceryl ether monooxygenase, is a tetrahydrobiopterin-dependent enzyme. It is the only enzyme known to cleave the ether bond of alkylglycerols and lyso-alkylglycerol phospholipids, including lyso-platelet activating factor. Although it has been first described already in 1964, it was not possible so far to purify the protein. It took until 2010 to assign a sequence to this labile integral membrane enzyme by bioinformatic selection of candidate genes, recombinant expression of these, and sensitive monitoring of the enzymatic activity by a fluorescence-based assay. The sequence shows no significant similarity with the other known tetrahydrobiopterin-dependent enzymes but contains the fatty acid hydroxylase protein motif signature. Proteins containing this signature are all labile and catalyze reactions similar to the alkylglycerol monooxygenase reaction. They are thought to use a di-iron centre for catalysis. Site directed mutagenesis of alkylglycerol monooxygenase defined a region of the active site and a conserved glutamate residue important for tetrahydrobiopterin interaction. Current research now focuses on defining a physiological role of this enzyme which occurs not only in mammals but also in commonly used model organisms such as zebrafish and the nematode Caenorhabditis elegans. © 2013 IUBMB Life 65(4):366-372, 2013. [ABSTRACT FROM AUTHOR]

Tetrahydrobiopterin has been shown to efficiently abrogate ischemia reperfusion injury (IRI). However, it is unclear, whether its beneficial action relies on cofactor activity of one of the five known tetrahydrobiopterin-dependent reactions or on its antioxidative capacity. We therefore compared tetrahydrobiopterin with the pterin derivate tetrahydroneopterin (similar biochemical properties, but no nitric oxide synthase cofactor activity) and the antioxidants vitamin C and 5-methyltetrahydrofolate. Donor mice were pretreated with tetrahydrobiopterin, tetrahydroneopterin, vitamin C, or 5-methyltetrahydrofolate. Pancreatic grafts were subjected to 16-h cold ischemia time and implanted in syngeneic recipients. Untreated and nontransplanted animals served as controls. Following 2-h reperfusion, microcirculation was analyzed by intravital fluorescence microscopy. Graft damage was assessed by histology and nitrotyrosine immunostaining, and tetrahydrobiopterin levels were determined by HPLC. Recipient survival served as ultimate readout. Prolonged cold ischemia time resulted in microcirculatory breakdown. Only tetrahydrobiopterin pretreatment succeeded to preserve the capillary net, whereas all other compounds showed no beneficial effects. Along with increased intragraft tetrahydrobiopterin levels during recovery and implantation, only tetrahydrobiopterin pretreatment led to significant reduction of IRI-related parenchymal damage enabling recipient survival. These results show a striking superiority of tetrahydrobiopterin in preventing lethal IRI compared with related compounds and suggest nitric oxide synthases as treatment target. [ABSTRACT FROM AUTHOR]

Alkylglycerol mono-oxygenase (EC 1.14.16.5) forms a third, distinct, class among tetrahydrobiopterin-dependent enzymes in addition to aromatic amino acid hydroxylases and nitric oxide synthases. Its protein sequence contains the fatty acid hydroxylase motif, a signature indicative of a di-iron centre, which contains eight conserved histidine residues. Membrane enzymes containing this motif, including alkylglycerol monooxygenase, are especially labile and so far have not been purified to homogeneity in active form. To obtain a first insight into structure-function relationships of this enzyme, we performed site-directed mutagenesis of 26 selected amino acid residues and expressed wild-type and mutant proteins containing a Cterminal Myc tag together with fatty aldehyde dehydrogenase in Chinese-hamster ovary cells. Among all of the acidic residues within the eight-histidinemotif, onlymutation of Glu137 to alanine led to an 18-fold increase in the Michaelis-Menten constant for tetrahydrobiopterin, suggesting a role in tetrahydrobiopterin interaction. A ninth additional histidine residue essential for activity was also identified. Nine membrane domains were predicted by four programs: ESKW, TMHMM, MEMSAT and Phobius. Prediction of a part of the structure using the Rosetta membrane ab initio method led to a plausible suggestion for a structure of the catalytic site of alkylglycerol mono-oxygenase. [ABSTRACT FROM AUTHOR]

Alkylglycerol monooxygenase (glyceryl-ether monooxygenase, EC 1.14.16.5) is the only enzyme known to cleave the 0-alkyl bond of ether lipids which are essential components of brain membranes, protect the eye from cataract, interfere or mediate signalling processes, and are required for spermatogenesis. Along with phenylalanine hydroxylase, tyrosine hydroxylase, tryptophan hydroxylase; and nitric oxide synthase, alkylglycerol monooxygenase is one of five known enzymatic reactions which depend on tetrahydrobiopterin. Although first described in 1964, no sequence had been assigned to this enzyme so far since it lost activity upon protein purification attempts. A functional library screen using pools of plasmids of a rat liver expression library transfected to CHO cells was also unsuccessful. We therefore selected human candidate genes by bióinformatic approaches and by proteomic analysis of partially purified enzyme and tested alkyiglycerol monooxygenase activity in CHO cells transfected with expression plasmids. Transmembrane protein 195. a predicted membrane protein with unassigned function which occurs in bilateral animals, was found to encode for tetrahydrobiopterin-dependent alkylglycerol monooxygenase. This sequence assignment was confirmed by injection of transmembrane protein 195 cRNA into Xenopus Iaevis oocytes. Transmembrane protein 195 shows no sequence homology to aromatic amino acid hydroxylases or nitric oxide synthases, but contains the fatty acid hydroxylase motif. This motif is found in enzymes which contain a diiron center and which carry out hydroxylations of lipids at aliphatic carbon atoms like alkylglycerol monooxygenase. This sequence assignment suggests that alkylglycerol monooxygenase forms a distinct third group among tetrahydrobiopterin-dependent enzymes. [ABSTRACT FROM AUTHOR]

Glyceryl ether monooxygenase is a tetrahydrobiopterin-dependent membrane-bound enzyme which catalyses the cleavage of lipid ethers into glycerol and the corresponding aldehyde. Despite many different characterisation and purification attempts, so far no gene and primary sequence have been assigned to this enzyme. The seven other tetrahydrobiopterin-dependent enzymes can be divided in the family of aromatic amino acid hydroxylases – comprising phenylalanine hydroxylase, tyrosine hydroxylase and the two tryptophan hydroxylases – and into the three nitric oxide synthases. We tested the influences of different metal ions and metal ion chelators on glyceryl ether monooxygenase, phenylalanine hydroxylase and nitric oxide synthase activity to elucidate the relationship of glyceryl ether monooxygenase to these two families. 1,10-Phenanthroline, an inhibitor of non-heme iron-dependent enzymes, was able to potently block glyceryl ether monooxygenase as well as phenylalanine hydroxylase, but had no effect on inducible nitric oxide synthase. Two tetrahydrobiopterin analogues, N5-methyltetrahydrobiopterin and 4-aminotetrahydrobiopterin, had a similar impact on glyceryl ether monooxygenase activity, as has already been shown for phenylalanine hydroxylase. These observations point to a close analogy of the role of tetrahydrobiopterin in glyceryl ether monooxygenase and in aromatic amino acid hydroxylases and suggest that glyceryl ether monooxygenase may require a non-heme iron for catalysis. [ABSTRACT FROM AUTHOR]

Channelopathies are often linked to defective protein folding and trafficking. Among them, the calcium channelopathy episodic ataxia type-2 (EA2) is an autosomal dominant disorder related to mutations in the pore-forming Cav2.1 subunit of P/Q-type calcium channels. Although EA2 is linked to loss of Cav2.1 channel activity, the molecular mechanism underlying dominant inheritance remains unclear. Here, we show that EA2 mutants as well as a truncated form (DI-II) of the Cav3.2 subunit of T-type calcium channel are misfolded, retained in the endoplasmic reticulum, and subject to proteasomal degradation. Pulse-chase experiments revealed that misfolded mutants bind to nascent wild-type Cav subunits and induce their subsequent degradation, thereby abolishing channel activity. We conclude that this destructive interaction mechanism promoted by Cav mutants is likely to occur in EA2 and in other inherited dominant channelopathies. [ABSTRACT FROM AUTHOR]

The mechanism of action of the antiepileptic and antinociceptive drugs of the gabapentinoid family has remained poorly under- stood. Gabapentin (GBP) binds to an exofacial epitope of the α2δ-1 and α2δ-2 auxiliary subunits of voltage-gated calcium channels, but acute inhibition of calcium currents by GBP is either very minor or absent. We formulated the hypothesis that GBP impairs the ability of α2δ subunits to enhance voltage-gated Ca2+channel plasma membrane density by means of an effect on trafficking. Our results conclusively demonstrate that GBP inhibits calcium currents, mimicking a lack ofα2δ only when applied chronically, but not acutely, both in heterologous expression systems and in dorsal root-ganglion neurons. GBP acts primarily at an intracellular location, requiring uptake, because the effect of chronically applied GBP is blocked by an inhibitor of the system-L neutral amino acid transporters and enhanced by coexpression of a transporter. However, it is mediated by α2δ subunits, being prevented by mutations in either α2δ-1 or α2δ-2 that abolish GBP binding, and is not observed for α2δ-3, which does not bind GBP. Furthermore, the trafficking of α2δ-2 and Cav2 channels is disrupted both by GBP and by the mutation in α2δ-2, which prevents GBP binding, and we find that GBP reduces cell-surface expression of α2δ-2 and Ca\v2.1 subunits. Our evidence indicates that GBP may act chronically by displacing an endogenous ligand that is normally a positive modulator of α2δ subunit function, thereby impairing the trafficking function of the α2δ subunits to which it binds. [ABSTRACT FROM AUTHOR]

Alkylglycerol monooxygenase (AGMO) is a tetrahydrobiopterin (BH4)-dependent enzyme with major expression in the liver and white adipose tissue that cleaves alkyl ether glycerolipids. The present study describes the disclosure and biological characterization of a candidate compound (Cp6), which inhibits AGMO with an IC50 of 30–100 µM and 5–20-fold preference of AGMO relative to other BH4-dependent enzymes, i.e., phenylalanine-hydroxylase and nitric oxide synthase. The viability and metabolic activity of mouse 3T3-L1 fibroblasts, HepG2 human hepatocytes and mouse RAW264.7 macrophages were not affected up to 10-fold of the IC50. However, Cp6 reversibly inhibited the differentiation of 3T3-L1 cells towards adipocytes, in which AGMO expression was upregulated upon differentiation. Cp6 reduced the accumulation of lipid droplets in adipocytes upon differentiation and in HepG2 cells exposed to free fatty acids. Cp6 also inhibited IL-4-driven differentiation of RAW264.7 macrophages towards M2-like macrophages, which serve as adipocyte progenitors in adipose tissue. Collectively, the data suggest that pharmacologic AGMO inhibition may affect lipid storage. [ABSTRACT FROM AUTHOR]

The transient receptor potential melastatin (TRPM) family belongs to the superfamily of TRP ion channels. It consists of eight family members that are involved in a plethora of cellular functions. TRPM2 is a homotetrameric Ca2+-permeable cation channel activated upon oxidative stress and is important, among others, for body heat control, immune cell activation and insulin secretion. Invertebrate TRPM2 proteins are channel enzymes; they hydrolyze the activating ligand, ADP-ribose, which is likely important for functional regulation. Since its cloning in 1998, the understanding of the biophysical properties of the channel has greatly advanced due to a vast number of structure–function studies. The physiological regulators of the channel have been identified and characterized in cell-free systems. In the wake of the recent structural biochemistry revolution, several TRPM2 cryo-EM structures have been published. These structures have helped to understand the general features of the channel, but at the same time have revealed unexplained mechanistic differences among channel orthologues. The present review aims at depicting the major research lines in TRPM2 structure-function. It discusses biophysical properties of the pore and the mode of action of direct channel effectors, and interprets these functional properties on the basis of recent three-dimensional structural models. [ABSTRACT FROM AUTHOR]

The gene encoding alkylglycerol monooxygenase (AGMO) was assigned 10 years ago. So far, AGMO is the only known enzyme capable of catalysing the breakdown of alkylglycerols and lyso-alkylglycerophospholipids. With the knowledge of the genetic information, it was possible to relate a potential contribution for mutations in the AGMO locus to human diseases by genome-wide association studies. A possible role for AGMO was implicated by genetic analyses in a variety of human pathologies such as type 2 diabetes, neurodevelopmental disorders, cancer, and immune defence. Deficient catabolism of stored lipids carrying an alkyl bond by an absence of AGMO was shown to impact on the overall lipid composition also outside the ether lipid pool. This review focuses on the current evidence of AGMO in human diseases and summarises experimental evidence for its role in immunity, energy homeostasis, and development in humans and several model organisms. With the progress in lipidomics platform and genetic identification of enzymes involved in ether lipid metabolism such as AGMO, it is now possible to study the consequence of gene ablation on the global lipid pool and further on certain signalling cascades in a variety of model organisms in more detail. [ABSTRACT FROM AUTHOR]

Cardiolipin (CL) is a phospholipid specific for mitochondrial membranes and crucial for many core tasks of this organelle. Its acyl chain configurations are tissue specific, functionally important, and generated via post-biosynthetic remodeling. However, this process lacks the necessary specificity to explain CL diversity, which is especially evident for highly specific CL compositions in mammalian tissues. To investigate the so far elusive regulatory origin of CL homeostasis in mice, we combine lipidomics, integrative transcriptomics, and data-driven machine learning. We demonstrate that not transcriptional regulation, but cellular phospholipid compositions are closely linked to the tissue specificity of CL patterns allowing artificial neural networks to precisely predict cross-tissue CL compositions in a consistent mechanistic specificity rationale. This is especially relevant for the interpretation of disease-related perturbations of CL homeostasis, by allowing differentiation between specific aberrations in CL metabolism and changes caused by global alterations in cellular (phospho-)lipid metabolism. • Mouse tissue mitochondria are defined by characteristic cardiolipin architectures • Transcriptional regulation does not explain cardiolipin diversity • Machine learning models explain cardiolipin states in tissues and cultured cells • Phospholipid pools, particularly linoleoyl content, control cardiolipin specificity The lipid architecture of biomembranes is crucial for their cellular functions. The regulatory origins of the strong tissue specificity of cardiolipins, a vital mitochondrial phospholipid class, were so far largely unresolved. Oemer et al. find that a single mechanism explains cardiolipin diversity across tissues on basis of the phospholipid environment. [ABSTRACT FROM AUTHOR]

Transplant vasculopathy (TV) represents a major obstacle to long-term graft survival and correlates with severity of ischemia reperfusion injury (IRI). Donor administration of the nitric oxide synthases (NOS) co-factor tetrahydrobiopterin has been shown to prevent IRI. Herein, we analysed whether tetrahydrobiopterin is also involved in TV development. Using a fully allogeneic mismatched (BALB/c to C57BL/6) murine aortic transplantation model grafts subjected to long cold ischemia time developed severe TV with intimal hyperplasia (α-smooth muscle actin positive cells in the neointima) and endothelial activation (increased P-selectin expression). Donor pretreatment with tetrahydrobiopterin significantly minimised these changes resulting in only marginal TV development. Severe TV observed in the non-treated group was associated with increased protein oxidation and increased occurrence of endothelial NOS monomers in the aortic grafts already during graft procurement. Tetrahydrobiopterin supplementation of the donor prevented all these early oxidative changes in the graft. Non-treated allogeneic grafts without cold ischemia time and syngeneic grafts did not develop any TV. We identified early protein oxidation and impaired endothelial NOS homodimer formation as plausible mechanistic explanation for the crucial role of IRI in triggering TV in transplanted aortic grafts. Therefore, targeting endothelial NOS in the donor represents a promising strategy to minimise TV. [ABSTRACT FROM AUTHOR]