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Anil K Agarwal - One of the best experts on this subject based on the ideXlab platform.
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messenger rna levels of enzymes involved in glycerolipid synthesis in the brain of the mouse and its alterations in AGPAT2 and db db mice
bioRxiv, 2020Co-Authors: Lila Gonzalezhodar, Anil K Agarwal, Victor CortesAbstract:Aims: Expression of genes encoding enzymes involved in glycerolipid and monoacylglycerol pathways in specific brain regions is poorly known and its impact in insulin resistance (IR) and type 2 diabetes (T2D) in the brain remains unreported. We determined mRNA levels of enzymes involved in glycerolipid synthesis in different regions of the mouse brain and evaluated their changes in two models of IR and T2D, the AGPAT2-/- and Leprdb/db mice. Methods: Cerebral cortex, hypothalamus, hippocampus and cerebellum were dissected from adult AGPAT2-/- mice, Leprdb/db mice and their respective wild type littermates. Total RNA was isolated and mRNA abundance was measured by RT-qPCR. Key findings: GPAT1, AGPAT1-4, LIPIN1/2, DGAT1/2 and MOGAT1 mRNAs were detected in all studied brain regions, whereas GPAT2, LIPIN3 and MOGAT2 were undetectable. Abundance of AGPATs, LIPIN1 and DGAT1, was higher in cerebellum and hypothalamus. LIPIN2 and MOGAT1 levels were higher in hypothalamus, and DGAT2 was higher in cortex and hypothalamus. In AGPAT2-/- mice, LIPIN1 levels were increased in all the brain regions. By contrast, GPAT1 in cortex and hypothalamus, AGPAT3 in hippocampus and hypothalamus, AGPAT4 in hypothalamus, and MOGAT1 in cortex, hypothalamus and cerebellum were lower in AGPAT2-/- mice. Leprdb/db mice showed fewer and milder changes, with increased levels of GPAT1 and LIPIN1 in cerebellum, and AGPAT3 in hypothalamus. Conclusions and Significance: Enzymes of glycerolipids synthesis are differentially expressed across regions of the mouse brain. Two mouse models of IR and T2D have altered gene expression of glycerolipid enzymes in the brain.
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AGPAT2 is essential for postnatal development and maintenance of white and brown adipose tissue
Elsevier, 2016Co-Authors: Kelly M. Cautivo, Anil K Agarwal, Carlos O. Lizama, Pablo J. Tapia, Abhimanyu Garg, Jay D. Horton, Víctor A. CortésAbstract:Objective: Characterize the cellular and molecular events responsible for lipodystrophy in AGPAT2 deficient mice. Methods: Adipose tissue and differentiated MEF were assessed using light and electron microscopy, followed by protein (immunoblots) and mRNA analysis (qPCR). Phospholipid profiling was determined by electrospray ionization tandem mass spectrometry (ESI-MS/MS). Results: In contrast to adult AGPAT2−/− mice, fetuses and newborn AGPAT2−/− mice have normal mass of white and brown adipose tissue. Loss of both the adipose tissue depots occurs during the first week of postnatal life as a consequence of adipocyte death and inflammatory infiltration of the adipose tissue. At the ultrastructural level, adipose tissue of newborn AGPAT2−/− mice is virtually devoid of caveolae and has abnormal mitochondria and lipid droplets. Autophagic structures are also abundant. Consistent with these findings, differentiated AGPAT2−/− mouse embryonic fibroblasts (MEFs) also have impaired adipogenesis, characterized by a lower number of lipid-laden cells and ultrastructural abnormalities in lipid droplets, mitochondria and plasma membrane. Overexpression of PPARγ, the master regulator of adipogenesis, increased the number of AGPAT2−/− MEFs that differentiated into adipocyte-like cells but did not prevent morphological abnormalities and cell death. Furthermore, differentiated AGPAT2−/− MEFs have abnormal phospholipid compositions with 3-fold increased levels of phosphatidic acid. Conclusion: We conclude that lipodystrophy in AGPAT2−/− mice results from postnatal cell death of adipose tissue in association with acute local inflammation. It is possible that AGPAT2 deficient adipocytes have an altered lipid filling or a reduced capacity to adapt the massive lipid availability associated with postnatal feeding. Keywords: AGPAT2, Adipose tissue, Adipogenesis, Phospholipid, Lipodystroph
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0021-972X/03/$15.00/0 The Journal of Clinical Endocrinology & Metabolism 88(10):4840–4847 Printed in U.S.A. Copyright © 2003 by The Endocrine Society doi: 10.1210/jc.2003-030855 Phenotypic and Genetic Heterogeneity in Congenital Generalized Lipod
2013Co-Authors: Anil K Agarwal, Vinaya Simha, Phillip Gorden, Elif A Oral, Stephanie Ann Moran, Zohra Zaidi, Figen Gurakan, Silva A ArslanianAbstract:Congenital generalized lipodystrophy (CGL) is a rare autosomal recessive disorder characterized by near complete absence of adipose tissue from birth. Recently, mutations in 1-acylglycerol-3-phosphate O-acyltransferase 2 (AGPAT2) and Berardinelli-Seip congenital lipodystrophy 2 (BSCL2) gene
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novel subtype of congenital generalized lipodystrophy associated with muscular weakness and cervical spine instability
American Journal of Medical Genetics Part A, 2008Co-Authors: Vinaya Simha, Patricia A Aronin, Anil K Agarwal, Susan T. IannacconeAbstract:Congenital generalized lipodystrophy (CGL) is a rare autosomal recessive disorder characterized by extreme paucity of adipose tissue from birth, and early onset of metabolic complications related to insulin resistance. Mutations in three genes, 1-acylglycerol 3-phosphate-O-acyltransferase 2 (AGPAT2), Berardinelli Seip Congenital Lipodystrophy 2 (BSCL2), and Caveolin-1 (CAV1) are associated with the three subtypes of this disorder, CGL1, CGL2 and CGL3, respectively. We report two siblings of Hispanic origin who displayed characteristic features of CGL such as generalized loss of subcutaneous fat from birth, acanthosis nigricans, acromegaloid habitus, umbilical prominence, hepatosplenomegaly, hypoleptinemia, dyslipidemia, and insulin resistance. However, no disease causing variants were detected in the DNA sequence of AGPAT2, BSCL2 or CAV1 genes. Further, whole body magnetic resonance imaging (MRI) in the two siblings revealed marked loss of subcutaneous, intraabdominal and intrathoracic fat like in other patients with CGL, but preservation of bone marrow fat which is invariably lost in all patients with CGL1 and CGL2, but not in the patient reported with CGL3. They also had generalized muscle weakness during infancy and early childhood associated with a nearly fivefold increase in serum creatine kinase (CK) levels, but with normal muscle biopsy and electrophysiologic studies. Both patients were also found to have atlantoaxial dislocation requiring surgical intervention. Thus, this pedigree represents a novel subtype of CGL characterized by generalized loss of body fat but with preservation of bone marrow fat, congenital muscular weakness and cervical spine instability. The genetic basis of this novel subtype remains to be determined.
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enzymatic activity of naturally occurring 1 acylglycerol 3 phosphate o acyltransferase 2 mutants associated with congenital generalized lipodystrophy
Biochemical and Biophysical Research Communications, 2005Co-Authors: Wasim A Haque, Anil K AgarwalAbstract:Mutations in the gene encoding 1-acylglycerol-3-phosphate-O-acyltransferase 2 (AGPAT2) have been reported in patients with congenital generalized lipodystrophy (CGL). AGPAT2, a 278 amino acid protein, belongs to the acyltransferase enzyme family, and has two conserved motifs, NHX(4)D and EGTR, involved in the enzymatic activity. The AGPATs catalyze acylation of lysophosphatidic acid (LPA) to phosphatidic acid (PA) during the biosynthesis of glycerophospholipids and triglycerides from glycerol-3-phosphate. The present studies were designed to determine the enzymatic activity of AGPAT2 mutants found in CGL patients to provide a molecular explanation for the phenotype and to obtain additional information about the structure-function relationship of AGPAT2 protein. The enzymatic activities of the wild type AGPAT2 and mutants were determined in cell lysates of overexpressing Chinese hamster ovary cells by measuring the conversion of [(3)H]LPA to [(3)H]PA in the presence of oleoyl-coenzyme A. Whereas, the R68X, 221delGT, 252delMRT, D180fsX251, and V167fsX183 mutants had markedly reduced enzymatic activity (median <15% of the wild type), the mutants, 140delF, G136R, and L228P, retained median activity ranging from 15% to 40% of the wild type enzyme. However, the missense mutant, A239V, had 90% of the wild type activity. We suggest that reduction in AGPAT2 enzymatic activity underlies the loss of adipose tissue in CGL. Our observations reveal an important role of various carboxy-terminal residues in determining the enzymatic activity of AGPAT2.
Rosalind A. Coleman - One of the best experts on this subject based on the ideXlab platform.
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glycerol 3 phosphate acyltranferase 2 behaves as a cancer testis gene and promotes growth and tumorigenicity of the breast cancer mda mb 231 cell line
PLOS ONE, 2014Co-Authors: Magali Pellonmaison, Rosalind A. Coleman, Elizabeth Renee Cattaneo, Ezequiel Lacunza, Mauro Aldo Montanaro, Maria Belen Garciafabiani, Mercedes C Solergerino, Ivana Yoseli Quiroga, Martin Carlos Abba, Maria R GonzalezbaroAbstract:The de novo synthesis of glycerolipids in mammalian cells begins with the acylation of glycerol-3-phosphate, catalyzed by glycerol-3-phosphate acyltransferase (GPAT). GPAT2 is a mitochondrial isoform primarily expressed in testis under physiological conditions. Because it is aberrantly expressed in multiple myeloma, it has been proposed as a novel cancer testis gene. Using a bioinformatics approach, we found that GPAT2 is highly expressed in melanoma, lung, prostate and breast cancer, and we validated GPAT2 expression at the protein level in breast cancer by immunohistochemistry. In this case GPAT2 expression correlated with a higher histological grade. 5-Aza-2′ deoxycytidine treatment of human cells lines induced GPAT2 expression suggesting epigenetic regulation of gene expression. In order to evaluate the contribution of GPAT2 to the tumor phenotype, we silenced its expression in MDA-MB-231 cells. GPAT2 knockdown diminished cell proliferation, anchorage independent growth, migration and tumorigenicity, and increased staurosporine-induced apoptosis. In contrast, GPAT2 over-expression increased cell proliferation rate and resistance to staurosporine-induced apoptosis. To understand the functional role of GPAT2, we performed a co-expression analysis in mouse and human testis and found a significant association with semantic terms involved in cell cycle, DNA integrity maintenance, piRNA biogenesis and epigenetic regulation. Overall, these results indicate the GPAT2 would be directly associated with the control of cell proliferation. In conclusion, we confirm GPAT2 as a cancer testis gene and that its expression contributes to the tumor phenotype of MDA-MB-231 cells.
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glycerol 3 phosphate acyltransferase 2 is expressed in spermatic germ cells and incorporates arachidonic acid into triacylglycerols
PLOS ONE, 2012Co-Authors: Elizabeth Renee Cattaneo, Rosalind A. Coleman, Magali Pellonmaison, Martin E Rabassa, Ezequiel Lacunza, Maria R GonzalezbaroAbstract:Background De novo glycerolipid synthesis begins with the acylation of glycerol-3 phosphate catalyzed by glycerol-3-phosphate acyltransferase (GPAT). In mammals, at least four GPAT isoforms have been described, differing in their cell and tissue locations and sensitivity to sulfhydryl reagents. In this work we show that mitochondrial GPAT2 overexpression in CHO-K1 cells increased TAG content and both GPAT and AGPAT activities 2-fold with arachidonoyl-CoA as a substrate, indicating specificity for this fatty acid. Methods and Results Incubation of GPAT2-transfected CHO-K1 cells with [1-14C]arachidonate for 3 h increased incorporation of [14C]arachidonate into TAG by 40%. Consistently, arachidonic acid was present in the TAG fraction of cells that overexpressed GPAT2, but not in control cells, corroborating GPAT2's role in synthesizing TAG that is rich in arachidonic acid. In rat and mouse testis, Gpat2 mRNA was expressed only in primary spermatocytes; the protein was also detected in late stages of spermatogenesis. During rat sexual maturation, both the testicular TAG content and the arachidonic acid content in the TAG fraction peaked at 30 d, matching the highest expression of Gpat2 mRNA and protein. Conclusions These results strongly suggest that GPAT2 expression is linked to arachidonoyl-CoA incorporation into TAG in spermatogenic germ cells.
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identification of a novel sn glycerol 3 phosphate acyltransferase isoform gpat4 as the enzyme deficient in agpat6 mice
Journal of Lipid Research, 2008Co-Authors: Cynthia A Nagle, Tal M. Lewin, Shuli Wang, Laurent Vergnes, Karen Reue, Hendrik Dejong, Rosalind A. ColemanAbstract:The regulation of triacylglycerol (TAG) synthesis and metabolism plays an important role in whole body energy homeostasis in mammals, and dysregulation of TAG synthesis and oxidation pathways has been implicated in the pathogenesis of obesity, lipodystrophy, cardiovascular disease, insulin resistance, and type 2 diabetes (1). De novo TAG synthesis begins with the formation of lysophosphatidic acid (LPA) through the acylation of the sn-1 position of glycerol-3-phosphate by sn-glycerol-3-phosphate acyltransferase (GPAT) (2). The second enzyme in de novo TAG synthesis, sn-1-acylglycerol-3-phosphate O-acyltransferase (AGPAT), acylates LPA to form phosphatidic acid (PA). Phosphatidic acid phosphohydrolase (also called lipin) converts PA to diacylglycerol (DAG) (3, 4), and diacylglycerol acyltransferase acylates DAG to form TAG. PA and DAG are also precursors for the glycerophospholipids. Three mammalian GPAT isoforms have been cloned (5–7). GPAT1 and GPAT2 are located in the outer mitochondrial membrane, whereas GPAT3 is located in the endoplasmic reticulum. GPAT1 is resistant to inactivation by sulfhydryl agents like N-ethylmaleimide (NEM), prefers palmitoyl-CoA as a substrate compared with oleoyl-CoA, and plays a regulatory role in liver TAG synthesis (2). GPAT2 is NEM-sensitive, has no preference for palmitoyl-CoA, and is highly expressed in rodent testis (8, 9). Recently, the gene product previously called AGPAT8 in the National Center for Biotechnology Information database was shown to have NEM-sensitive GPAT activity and to lack AGPAT activity; this gene product was renamed GPAT3 (7). Because small interfering RNA knockdown of Gpat3 in 3T3-L1 adipocytes reduces total GPAT activity by ∼60% (7, 10), it is likely that GPAT3 is important for TAG synthesis in white adipose tissue (WAT). However, GPAT3 is unlikely to play a major role in TAG synthesis in liver, where its mRNA expression is very low (7, 10). In liver, GPAT1 accounts for 30–50% of the total GPAT activity (2). The remaining hepatic GPAT activity is NEM-sensitive and microsomal, suggesting that one or more additional microsomal NEM-sensitive GPAT isoforms must exist. Eight genes with predicted amino acid sequence similarities to GPAT1 have been named AGPAT isoforms 1–8. AGPAT1 and -2 are well characterized and have high AGPAT activity (11, 12). Mutations in AGPAT2 cause generalized human congenital lipodystrophy, with absent abdominal and subcutaneous adipose tissue, systemic insulin resistance, and hypertriglyceridemia (13). Compared with AGPAT1 and -2, the AGPAT activities reported for AGPAT3 to -5 are extremely low (14); no enzyme activity has been reported for AGPAT6 or -7. We previously characterized mice with a targeted deletion in Agpat6 (Agpat6−/− mice; {"type":"entrez-protein","attrs":{"text":"NP_848934","term_id":"30520329","term_text":"NP_848934"}}NP_848934) (10, 15). These mice have a subdermal lipodystrophy, resistance to high-fat diet-induced obesity, and a 50% reduction of TAG content in liver, adipose tissue, and mammary epithelium, suggesting an important role for AGPAT6 in TAG synthesis in these tissues. AGPAT6 shares amino acid sequence homology with other glycerolipid acyltransferases at conserved motifs I–V (Table 1) and is closely related to GPAT1 and -2 in motifs I–IV, which were identified as important for acyltransferase catalysis and glycerol-3-phosphate binding domains (16). AGPAT6 shares 66% amino acid homology with “AGPAT8,” recently identified as GPAT3 (7), and confocal microscopy shows that AGPAT6 localizes to the endoplasmic reticulum (15). These observations raised the possibility that AGPAT6, like AGPAT8, is in fact a GPAT enzyme. Here, we demonstrate that expression of this enzyme in Cos-7 cells increases GPAT, but not AGPAT, activity. Furthermore, tissues from Agpat6−/− mice show reduced GPAT activity but normal AGPAT activity. Our studies thus establish that Agpat6 encodes the primary microsomal NEM-sensitive GPAT activity in liver, GPAT4. TABLE 1 Acyltransferase motifs in Escherichia coli and mouse AGPAT and GPAT isoforms
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cloning and functional characterization of a novel mitochondrial n ethylmaleimide sensitive glycerol 3 phosphate acyltransferase gpat2
Archives of Biochemistry and Biophysics, 2007Co-Authors: Shuli Wang, Tal M. Lewin, Nicole M J Schwerbrock, Douglas P Lee, Nan Gong, Douglas G Mashek, Maria R Gonzalezbaro, Cliona Stapleton, Rosalind A. ColemanAbstract:Abstract Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial and rate-limiting step in glycerolipid synthesis. Several mammalian GPAT activities have been recognized, including N-ethylmaleimide (NEM)-sensitive isoforms in microsomes and mitochondria and an NEM-resistant form in mitochondrial outer membrane (GPAT1). We have now cloned a second mitochondrial isoform, GPAT2 from mouse testis. The open-reading frame encodes a protein of 798 amino acids with a calculated mass of 88.8 kDa and 27% amino acid identity to GPAT1. Testis mRNA expression was 50-fold higher than in liver or brown adipose tissue, but the specific activity of NEM-sensitive GPAT in testis mitochondria was similar to that in liver. When Cos-7 cells were transiently transfected with GPAT2, NEM-sensitive GPAT activity increased 30%. Confocal microscopy confirmed a mitochondrial location. Incubation of GPAT2-transfected Cos-7 cells with trace (3 μM; 0.25 μCi) [1-14C]oleate for 6 h increased incorporation of [14C]oleate into TAG 84%. In contrast, incorporation into phospholipid species was lower than in control cells. Although a polyclonal antibody raised against full-length GPAT1 detected an ∼89-kDa band in liver and testis from GPAT1 null mice and both 89- and 80-kDa bands in BAT from the knockout animals, the GPAT2 protein expressed in Cos-7 cells was only 80 kDa. In vitro translation showed a single product of 89 kDa. Unlike GPAT1, GPAT2 mRNA abundance in liver was not altered by fasting or refeeding. GPAT2 is likely to have a specialized function in testis.
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agpat6 a novel lipid biosynthetic gene required for triacylglycerol production in mammary epithelium
Journal of Lipid Research, 2006Co-Authors: Anne P Beigneux, Rosalind A. Coleman, Laurent Vergnes, Xin Qiao, Steven Quatela, Ryan Davis, Steven M Watkins, Rosemary L Walzem, Mark R Philips, Karen ReueAbstract:In analyzing the sequence tags for mutant mouse embryonic stem (ES) cell lines in BayGenomics (a mouse gene-trapping resource), we identified a novel gene, 1-acylglycerol-3-phosphate O-acyltransferase (Agpat6), with sequence similarities to previously characterized glycerolipid acyltransferases. Agpat6's closest family member is another novel gene that we have provisionally designated Agpat8. Both Agpat6 and Agpat8 are conserved from plants, nematodes, and flies to mammals. AGPAT6, which is predicted to contain multiple membrane-spanning helices, is found exclusively within the endoplasmic reticulum (ER) in mammalian cells. To gain insights into the in vivo importance of Agpat6, we used the Agpat6 ES cell line from BayGenomics to create Agpat6-deficient (Agpat6 -/- ) mice. Aipat6 -/- mice lacked full-length Agpat6 transcripts, as judged by northern blots. One of the most striking phenotypes of Agpat6 -/- mice was a defect in lactation. Pups nursed by Agpat6 -/- mothers die perinatally. Normally, Agpat6 is expressed at high levels in the mammary epithelium of breast tissue, but not in the surrounding adipose tissue. Histological studies revealed that the aveoli and ducts of Agpat6 -/- lactating mammary glands were underdeveloped, and there was a dramatic decrease in the size and number of lipid droplets within mammary epithelial cells and ducts. Also, the milk from Agpat6 -/- mice was markedly depleted in diacylglycerols and triacylglycerols. Thus, we identified a novel glycerolipid acyltransferase of the ER, AGPAT6, which is crucial for the production of milk fat by the mammary gland.
Vial Thomas - One of the best experts on this subject based on the ideXlab platform.
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Le virus de la dengue détourne le métabolisme des phospholipides du moustique pour sa réplication
2020Co-Authors: Vial ThomasAbstract:Plus de la moitié de la population mondiale est exposée au risque d'infection par le virus de la dengue (DENV) en raison de la distribution mondiale de ses moustiques vecteurs. Il n'existe ni vaccin ni traitement efficace. La seule stratégie disponible repose sur les insecticides, contre lesquels les moustiques développent une résistance. Les virus utilisent le métabolome de l'hôte pour la réplication et la dissémination. C'est particulièrement vrai pour les virus enveloppés comme le DENV qui dépend des membranes lipidiques de l'hôte pour compléter son cycle de vie. Pour atteindre un environnement métabolique optimal, les virus perturbent le métabolome de l'hôte. La compréhension de ces altérations chez les moustiques vecteurs pourrait révéler de nouvelles stratégies pour bloquer la transmission du DENV. Ici, nous avons caractérisé comment le DENV détourne le lipidome du moustique Aedes aegypti. Pour décrire les changements métaboliques tout au long du cycle du DENV chez le moustique, nous avons débeloppé une méthode de chromatographie liquide et de spectrométrie de masse à haute résolution (LC-HRMS) à différents stades de l'infection chez le vecteur. Nous avons révélé une reconfiguration majeure des phospholipides tout au long du cycle du DENV chez le moustique, dans les cellules, l'intestin moyen et le moustique entier. Pour déchiffrer la façon dont le virus reconfigure les phospholipides, nous avons caractérisé phylogénétiquement les isoformes de l'enzyme acylglycerol-phosphate acyltransférase (AGPAT) et identifié celles qui catalysent une étape limitante dans la biogenèse des phospholipides. Nous avons constaté que l'infection par le DENV diminuait l'expression de AGPAT1, dont la déplétion renforce l'infection en maintenant des concentrations élevées d'aminophospholipides (aminoPL), en particulier la phosphatidylcholine (PC) et la phosphatidyléthanolamine (PE), pendant le cycle du DENV chez le moustique. En démontrant que la sous-régulation de AGPAT1, causé par le virus, fournit un environnement proviral, nous révèlons le premier facteur métabolique hôte chez les moustiques et soulignent le rôle des aminophospholipides dans le cycle cellulaire viral. Nous avons ensuite cherché à confirmer que le virus influence la biosynthèse des aminoPL et déterminer à quel stade du cycle viral la reconfiguration des aminoPL est nécessaire. La biosynthèse de novo de PC et de PE est connue sous le nom de voie de Kennedy, où un diacylglycérol (DAG) incorpore soit un groupe choline, soit un groupe éthanolamine. Le remodelage des AminoPL par déacylation/réacylation assure ensuite un dynamisme des membranes qui participe aux réarrangements membranaires. En utilisant un marquage isotopique avec une supplémentation en éthanolamine ou en choline, nous avons montré que le virus module la biosynthèse des PC et des PE en interagissant avec le remodelage membranaire. Soulignant l'importance de la voie de Kennedy dans l'infection par le DENV, la supplémentation en éthanolamine a réduit le titre du virus dans les cellules de moustiques en modifiant la composition de PC et PE. Bien que la supplémentation en éthanolamine n'ait pas modifié l'attachement, l'internalisation ou la traduction, elle réduit la réplication et entraîne un ratio plus faible de particules infectieuses, probablement en raison d'une réplication déficiente. Ces résultats confirment l'importance des aminoPL dans l'infection des moustiques par le DENV et révèlent l'importance de la composition des aminoPL dans la réplication. Les PC et PE sont les espèces de phospholipides les plus abondantes dans les cellules eucaryotes et contribuent à l'architecture de la membrane cellulaire, en particulier dans le réticulum endoplasmique, où la réplication a lieu. L'inhibition de la reconfiguration des aminoPL par la supplémentation en éthanolamine pourrait représenter une nouvelle stratégie pour interférer avec la perturbation du métabolome des moustiques par le virus de la dengue.More than half of the world population is at risk of dengue virus (DENV) infection because of the global distribution of its mosquito vectors. There is neither effective vaccine nor therapeutics. The only available strategy relies on insecticides, against which mosquitoes are developing resistance. Viruses utilize the host metabolome for replication and dissemination. This is particularly true for envelope viruses like DENV that relies on host lipid membranes to complete their life-cycle. To reach an optimal metabolic environment, viruses subvert the host metabolome. Understanding DENV-mosquito metabolic interactions will reveal novel strategies to stop DENV transmission. Here, we characterized how DENV hijacks the Aedes aegypti mosquito lipidome to identify targets for novel transmission-blocking interventions. To describe metabolic changes throughout the mosquito DENV cycle, we deployed a Liquid chromatography-high resolution mass spectrometry (LC-HRMS) workflow at different stages of vector infection. We revealed a major phospholipid reconfiguration throughout the DENV mosquito cycle, in cells, midguts, and whole mosquitoes. To decipher how DENV reconfigures phospholipids, we phylogenetically characterized acylglycerolphosphate acyltransferase (AGPAT) enzyme isoforms and identified those (i.e., AGPAT1) that catalyze a central rate-limiting step in phospholipid biogenesis. We found that DENV infection decreased AGPAT1 expression, which depletion enhances infection by maintaining high aminophospholipid (aminoPL) concentrations, especially phosphatidylcholine (PC) and phosphatidylethanolamine (PE), during DENV mosquito cycle. By demonstrating that DENV-mediated AGPAT1 downregulation provides a proviral environment, these results reveal the first metabolic host factor in mosquitoes and emphasize the role of aminophospholipids in DENV cellular cycle. We then undertook to precise how DENV influences aminoPL biosynthesis and what stage of DENV cellular cycle requires aminoPL reconfiguration. De novo biosynthesis of PC and PE is known as the Kennedy pathway, where a diacylglycerol (DAG) incorporates either a choline or an ethanolamine group. AminoPL remodeling by deacylation/reacylation then ensures membrane dynamism that participates in membrane rearrangements. Using isotopic labelling through ethanolamine or choline supplementation, we showed that DENV modulates PC and PE biosynthesis by interacting with membrane remodeling. Further supporting the importance of the Kennedy pathway in DENV infection, ethanolamine supplementation reduced virus titer in mosquito cells by altering composition of specific PC and PE. While ethanolamine-mediated aminoPL disruption did not alter attachment, internalization or translation, it reduced replication and resulted in a lower ratio of infectious particles, likely because of deficient replication. These results strongly support the importance of aminoPLs in DENV infection of mosquitoes and reveal the importance of aminoPL composition in replication. PC and PE are the most abundant phospholipid species in eukaryotic cells and contribute to cell membrane architecture, especially in the endoplasmic reticulum, where replication takes place. Disruption of aminoPL reconfiguration may represent a novel strategy to interfere with DENV subversion of mosquito metabolome
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Le virus de la dengue détourne le métabolisme des phospholipides du moustique pour sa réplication
HAL CCSD, 2020Co-Authors: Vial ThomasAbstract:More than half of the world population is at risk of dengue virus (DENV) infection because of the global distribution of its mosquito vectors. There is neither effective vaccine nor therapeutics. The only available strategy relies on insecticides, against which mosquitoes are developing resistance. Viruses utilize the host metabolome for replication and dissemination. This is particularly true for envelope viruses like DENV that relies on host lipid membranes to complete their life-cycle. To reach an optimal metabolic environment, viruses subvert the host metabolome. Understanding DENV-mosquito metabolic interactions will reveal novel strategies to stop DENV transmission. Here, we characterized how DENV hijacks the Aedes aegypti mosquito lipidome to identify targets for novel transmission-blocking interventions. To describe metabolic changes throughout the mosquito DENV cycle, we deployed a Liquid chromatography-high resolution mass spectrometry (LC-HRMS) workflow at different stages of vector infection. We revealed a major phospholipid reconfiguration throughout the DENV mosquito cycle, in cells, midguts, and whole mosquitoes. To decipher how DENV reconfigures phospholipids, we phylogenetically characterized acylglycerolphosphate acyltransferase (AGPAT) enzyme isoforms and identified those (i.e., AGPAT1) that catalyze a central rate-limiting step in phospholipid biogenesis. We found that DENV infection decreased AGPAT1 expression, which depletion enhances infection by maintaining high aminophospholipid (aminoPL) concentrations, especially phosphatidylcholine (PC) and phosphatidylethanolamine (PE), during DENV mosquito cycle. By demonstrating that DENV-mediated AGPAT1 downregulation provides a proviral environment, these results reveal the first metabolic host factor in mosquitoes and emphasize the role of aminophospholipids in DENV cellular cycle. We then undertook to precise how DENV influences aminoPL biosynthesis and what stage of DENV cellular cycle requires aminoPL reconfiguration. De novo biosynthesis of PC and PE is known as the Kennedy pathway, where a diacylglycerol (DAG) incorporates either a choline or an ethanolamine group. AminoPL remodeling by deacylation/reacylation then ensures membrane dynamism that participates in membrane rearrangements. Using isotopic labelling through ethanolamine or choline supplementation, we showed that DENV modulates PC and PE biosynthesis by interacting with membrane remodeling.[...]Plus de la moitié de la population mondiale est exposée au risque d'infection par le virus de la dengue (DENV) en raison de la distribution mondiale de ses moustiques vecteurs. Il n'existe ni vaccin ni traitement efficace. La seule stratégie disponible repose sur les insecticides, contre lesquels les moustiques développent une résistance. Les virus utilisent le métabolome de l'hôte pour la réplication et la dissémination. C'est particulièrement vrai pour les virus enveloppés comme le DENV qui dépend des membranes lipidiques de l'hôte pour compléter son cycle de vie. Pour atteindre un environnement métabolique optimal, les virus perturbent le métabolome de l'hôte. La compréhension de ces altérations chez les moustiques vecteurs pourrait révéler de nouvelles stratégies pour bloquer la transmission du DENV. Ici, nous avons caractérisé comment le DENV détourne le lipidome du moustique Aedes aegypti. Pour décrire les changements métaboliques tout au long du cycle du DENV chez le moustique, nous avons débeloppé une méthode de chromatographie liquide et de spectrométrie de masse à haute résolution (LC-HRMS) à différents stades de l'infection chez le vecteur. Nous avons révélé une reconfiguration majeure des phospholipides tout au long du cycle du DENV chez le moustique, dans les cellules, l'intestin moyen et le moustique entier. Pour déchiffrer la façon dont le virus reconfigure les phospholipides, nous avons caractérisé phylogénétiquement les isoformes de l'enzyme acylglycerol-phosphate acyltransférase (AGPAT) et identifié celles qui catalysent une étape limitante dans la biogenèse des phospholipides. Nous avons constaté que l'infection par le DENV diminuait l'expression de AGPAT1, dont la déplétion renforce l'infection en maintenant des concentrations élevées d'aminophospholipides (aminoPL), en particulier la phosphatidylcholine (PC) et la phosphatidyléthanolamine (PE), pendant le cycle du DENV chez le moustique. En démontrant que la sous-régulation de AGPAT1, causé par le virus, fournit un environnement proviral, nous révèlons le premier facteur métabolique hôte chez les moustiques et soulignent le rôle des aminophospholipides dans le cycle cellulaire viral. Nous avons ensuite cherché à confirmer que le virus influence la biosynthèse des aminoPL et déterminer à quel stade du cycle viral la reconfiguration des aminoPL est nécessaire. La biosynthèse de novo de PC et de PE est connue sous le nom de voie de Kennedy, où un diacylglycérol (DAG) incorpore soit un groupe choline, soit un groupe éthanolamine. [...
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Dengue virus diverts the mosquito phospholipid metabolism for replication
2020Co-Authors: Vial ThomasAbstract:Plus de la moitié de la population mondiale est exposée au risque d'infection par le virus de la dengue (DENV) en raison de la distribution mondiale de ses moustiques vecteurs. Il n'existe ni vaccin ni traitement efficace. La seule stratégie disponible repose sur les insecticides, contre lesquels les moustiques développent une résistance. Les virus utilisent le métabolome de l'hôte pour la réplication et la dissémination. C'est particulièrement vrai pour les virus enveloppés comme le DENV qui dépend des membranes lipidiques de l'hôte pour compléter son cycle de vie. Pour atteindre un environnement métabolique optimal, les virus perturbent le métabolome de l'hôte. La compréhension de ces altérations chez les moustiques vecteurs pourrait révéler de nouvelles stratégies pour bloquer la transmission du DENV. Ici, nous avons caractérisé comment le DENV détourne le lipidome du moustique Aedes aegypti. Pour décrire les changements métaboliques tout au long du cycle du DENV chez le moustique, nous avons débeloppé une méthode de chromatographie liquide et de spectrométrie de masse à haute résolution (LC-HRMS) à différents stades de l'infection chez le vecteur. Nous avons révélé une reconfiguration majeure des phospholipides tout au long du cycle du DENV chez le moustique, dans les cellules, l'intestin moyen et le moustique entier. Pour déchiffrer la façon dont le virus reconfigure les phospholipides, nous avons caractérisé phylogénétiquement les isoformes de l'enzyme acylglycerol-phosphate acyltransférase (AGPAT) et identifié celles qui catalysent une étape limitante dans la biogenèse des phospholipides. Nous avons constaté que l'infection par le DENV diminuait l'expression de AGPAT1, dont la déplétion renforce l'infection en maintenant des concentrations élevées d'aminophospholipides (aminoPL), en particulier la phosphatidylcholine (PC) et la phosphatidyléthanolamine (PE), pendant le cycle du DENV chez le moustique. En démontrant que la sous-régulation de AGPAT1, causé par le virus, fournit un environnement proviral, nous révèlons le premier facteur métabolique hôte chez les moustiques et soulignent le rôle des aminophospholipides dans le cycle cellulaire viral. Nous avons ensuite cherché à confirmer que le virus influence la biosynthèse des aminoPL et déterminer à quel stade du cycle viral la reconfiguration des aminoPL est nécessaire. La biosynthèse de novo de PC et de PE est connue sous le nom de voie de Kennedy, où un diacylglycérol (DAG) incorpore soit un groupe choline, soit un groupe éthanolamine. [...]More than half of the world population is at risk of dengue virus (DENV) infection because of the global distribution of its mosquito vectors. There is neither effective vaccine nor therapeutics. The only available strategy relies on insecticides, against which mosquitoes are developing resistance. Viruses utilize the host metabolome for replication and dissemination. This is particularly true for envelope viruses like DENV that relies on host lipid membranes to complete their life-cycle. To reach an optimal metabolic environment, viruses subvert the host metabolome. Understanding DENV-mosquito metabolic interactions will reveal novel strategies to stop DENV transmission. Here, we characterized how DENV hijacks the Aedes aegypti mosquito lipidome to identify targets for novel transmission-blocking interventions. To describe metabolic changes throughout the mosquito DENV cycle, we deployed a Liquid chromatography-high resolution mass spectrometry (LC-HRMS) workflow at different stages of vector infection. We revealed a major phospholipid reconfiguration throughout the DENV mosquito cycle, in cells, midguts, and whole mosquitoes. To decipher how DENV reconfigures phospholipids, we phylogenetically characterized acylglycerolphosphate acyltransferase (AGPAT) enzyme isoforms and identified those (i.e., AGPAT1) that catalyze a central rate-limiting step in phospholipid biogenesis. We found that DENV infection decreased AGPAT1 expression, which depletion enhances infection by maintaining high aminophospholipid (aminoPL) concentrations, especially phosphatidylcholine (PC) and phosphatidylethanolamine (PE), during DENV mosquito cycle. By demonstrating that DENV-mediated AGPAT1 downregulation provides a proviral environment, these results reveal the first metabolic host factor in mosquitoes and emphasize the role of aminophospholipids in DENV cellular cycle. We then undertook to precise how DENV influences aminoPL biosynthesis and what stage of DENV cellular cycle requires aminoPL reconfiguration. De novo biosynthesis of PC and PE is known as the Kennedy pathway, where a diacylglycerol (DAG) incorporates either a choline or an ethanolamine group. AminoPL remodeling by deacylation/reacylation then ensures membrane dynamism that participates in membrane rearrangements. Using isotopic labelling through ethanolamine or choline supplementation, we showed that DENV modulates PC and PE biosynthesis by interacting with membrane remodeling.[...
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Dengue virus reduces AGPAT1 expression to alter phospholipids and enhance infection in Aedes aegypti
'Public Library of Science (PLoS)', 2019Co-Authors: Vial Thomas, Tan Wei-lian, Xiang, Benjamin Wong Wei, Missé Dorothée, Deharo Eric, Marti Guillaume, Pompon JulienAbstract:International audienceMore than half of the world population is at risk of dengue virus (DENV) infection because of the global distribution of its mosquito vectors. DENV is an envelope virus that relies on host lipid membranes for its life-cycle. Here, we characterized how DENV hijacks the mosquito lipidome to identify targets for novel transmission-blocking interventions. To describe metabolic changes throughout the mosquito DENV cycle, we deployed a Liquid chromatography–high resolution mass spectrometry (LC-HRMS) workflow including spectral similarity annotation in cells, midguts and whole mosquitoes at different times post infection. We revealed a major aminophospholipid reconfiguration with an overall early increase, followed by a reduction later in the cycle. We phylogenetically characterized acylglycerolphosphate acyltransferase (AGPAT) enzyme isoforms to identify those that catalyze a rate-limiting step in phospholipid biogenesis, the acylation of lysophosphatidate to phosphatidate. We showed that DENV infection decreased AGPAT1, but did not alter AGPAT2 expression in cells, midguts and mosquitoes. Depletion of either AGPAT1 or AGPAT2 increased aminophospholipids and partially recapitulated DENV-induced reconfiguration before infection in vitro. However, only AGPAT1 depletion promoted infection by maintaining high aminophospholipid concentrations. In mosquitoes, AGPAT1 depletion also partially recapitulated DENV-induced aminophospholipid increase before infection and enhanced infection by maintaining high aminophospholipid concentrations. These results indicate that DENV inhibition of AGPAT1 expression promotes infection by increasing aminophospholipids, as observed in the mosquito’s early DENV cycle. Furthermore, in AGPAT1-depleted mosquitoes, we showed that enhanced infection was associated with increased consumption/redirection of aminophospholipids. Our study suggests that DENV regulates aminophospholipids, especially phosphatidylcholine and phosphatidylethanolamine, by inhibiting AGPAT1 expression to increase aminophospholipid availability for virus multiplication
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Dengue virus reduces AGPAT1 expression to alter phospholipids and enhance infection in Aedes aegypti
2019Co-Authors: Vial Thomas, Missé Dorothée, Deharo Eric, Tan W. L., Xiang B. W. W., Marti G., Pompon JulienAbstract:Author summary Dengue is endemic in tropical and subtropical regions, and has now encroached onto temperate regions because of the geographic expansion of its vector, Aedes aegypti. In the absence of effective vaccine and curative drug, the sole intervention relies on containment strategies using insecticide. However, occurrence of insecticide resistance diminishes vector control efficacy. Here, we explore the nascent field of mosquito metabolomics as part of our discovery effort for new transmission-blocking targets. Dengue virus (DENV) relies on host metabolome, specifically the lipid membrane to complete its life-cycle. However, little is known about how DENV subverts the mosquito physiology. Using high-resolution mass spectrometry, we described metabolic changes incurred by DENV throughout the mosquito cycle, from cellular replication onset to systemic infection. Membrane phospholipids were highly reconfigured and were associated with reduced expression of AGPAT1, an enzyme involved in their biogenesis. AGPAT1 depletion partially recapitulated DENV-induced metabolic reconfiguration and enhanced infection by maintaining high phospholipid concentrations. These phospholipids were then consumed/redirected later in the mosquito DENV cycle. Our work comprehensively describes metabolic changes associated with DENV infection. In addition, we reveal how DENV subdues the lipidome for its benefit by demonstrating the role of phospholipids in mosquito infection. More than half of the world population is at risk of dengue virus (DENV) infection because of the global distribution of its mosquito vectors. DENV is an envelope virus that relies on host lipid membranes for its life-cycle. Here, we characterized how DENV hijacks the mosquito lipidome to identify targets for novel transmission-blocking interventions. To describe metabolic changes throughout the mosquito DENV cycle, we deployed a Liquid chromatography-high resolution mass spectrometry (LC-HRMS) workflow including spectral similarity annotation in cells, midguts and whole mosquitoes at different times post infection. We revealed a major aminophospholipid reconfiguration with an overall early increase, followed by a reduction later in the cycle. We phylogenetically characterized acylglycerolphosphate acyltransferase (AGPAT) enzyme isoforms to identify those that catalyze a rate-limiting step in phospholipid biogenesis, the acylation of lysophosphatidate to phosphatidate. We showed that DENV infection decreased AGPAT1, but did not alter AGPAT2 expression in cells, midguts and mosquitoes. Depletion of either AGPAT1 or AGPAT2 increased aminophospholipids and partially recapitulated DENV-induced reconfiguration before infection in vitro. However, only AGPAT1 depletion promoted infection by maintaining high aminophospholipid concentrations. In mosquitoes, AGPAT1 depletion also partially recapitulated DENV-induced aminophospholipid increase before infection and enhanced infection by maintaining high aminophospholipid concentrations. These results indicate that DENV inhibition of AGPAT1 expression promotes infection by increasing aminophospholipids, as observed in the mosquito's early DENV cycle. Furthermore, in AGPAT1-depleted mosquitoes, we showed that enhanced infection was associated with increased consumption/redirection of aminophospholipids. Our study suggests that DENV regulates aminophospholipids, especially phosphatidylcholine and phosphatidylethanolamine, by inhibiting AGPAT1 expression to increase aminophospholipid availability for virus multiplication
Maria R Gonzalezbaro - One of the best experts on this subject based on the ideXlab platform.
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glycerol 3 phosphate acyltranferase 2 behaves as a cancer testis gene and promotes growth and tumorigenicity of the breast cancer mda mb 231 cell line
PLOS ONE, 2014Co-Authors: Magali Pellonmaison, Rosalind A. Coleman, Elizabeth Renee Cattaneo, Ezequiel Lacunza, Mauro Aldo Montanaro, Maria Belen Garciafabiani, Mercedes C Solergerino, Ivana Yoseli Quiroga, Martin Carlos Abba, Maria R GonzalezbaroAbstract:The de novo synthesis of glycerolipids in mammalian cells begins with the acylation of glycerol-3-phosphate, catalyzed by glycerol-3-phosphate acyltransferase (GPAT). GPAT2 is a mitochondrial isoform primarily expressed in testis under physiological conditions. Because it is aberrantly expressed in multiple myeloma, it has been proposed as a novel cancer testis gene. Using a bioinformatics approach, we found that GPAT2 is highly expressed in melanoma, lung, prostate and breast cancer, and we validated GPAT2 expression at the protein level in breast cancer by immunohistochemistry. In this case GPAT2 expression correlated with a higher histological grade. 5-Aza-2′ deoxycytidine treatment of human cells lines induced GPAT2 expression suggesting epigenetic regulation of gene expression. In order to evaluate the contribution of GPAT2 to the tumor phenotype, we silenced its expression in MDA-MB-231 cells. GPAT2 knockdown diminished cell proliferation, anchorage independent growth, migration and tumorigenicity, and increased staurosporine-induced apoptosis. In contrast, GPAT2 over-expression increased cell proliferation rate and resistance to staurosporine-induced apoptosis. To understand the functional role of GPAT2, we performed a co-expression analysis in mouse and human testis and found a significant association with semantic terms involved in cell cycle, DNA integrity maintenance, piRNA biogenesis and epigenetic regulation. Overall, these results indicate the GPAT2 would be directly associated with the control of cell proliferation. In conclusion, we confirm GPAT2 as a cancer testis gene and that its expression contributes to the tumor phenotype of MDA-MB-231 cells.
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glycerol 3 phosphate acyltransferase 2 is expressed in spermatic germ cells and incorporates arachidonic acid into triacylglycerols
PLOS ONE, 2012Co-Authors: Elizabeth Renee Cattaneo, Rosalind A. Coleman, Magali Pellonmaison, Martin E Rabassa, Ezequiel Lacunza, Maria R GonzalezbaroAbstract:Background De novo glycerolipid synthesis begins with the acylation of glycerol-3 phosphate catalyzed by glycerol-3-phosphate acyltransferase (GPAT). In mammals, at least four GPAT isoforms have been described, differing in their cell and tissue locations and sensitivity to sulfhydryl reagents. In this work we show that mitochondrial GPAT2 overexpression in CHO-K1 cells increased TAG content and both GPAT and AGPAT activities 2-fold with arachidonoyl-CoA as a substrate, indicating specificity for this fatty acid. Methods and Results Incubation of GPAT2-transfected CHO-K1 cells with [1-14C]arachidonate for 3 h increased incorporation of [14C]arachidonate into TAG by 40%. Consistently, arachidonic acid was present in the TAG fraction of cells that overexpressed GPAT2, but not in control cells, corroborating GPAT2's role in synthesizing TAG that is rich in arachidonic acid. In rat and mouse testis, Gpat2 mRNA was expressed only in primary spermatocytes; the protein was also detected in late stages of spermatogenesis. During rat sexual maturation, both the testicular TAG content and the arachidonic acid content in the TAG fraction peaked at 30 d, matching the highest expression of Gpat2 mRNA and protein. Conclusions These results strongly suggest that GPAT2 expression is linked to arachidonoyl-CoA incorporation into TAG in spermatogenic germ cells.
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cloning and functional characterization of a novel mitochondrial n ethylmaleimide sensitive glycerol 3 phosphate acyltransferase gpat2
Archives of Biochemistry and Biophysics, 2007Co-Authors: Shuli Wang, Tal M. Lewin, Nicole M J Schwerbrock, Douglas P Lee, Nan Gong, Douglas G Mashek, Maria R Gonzalezbaro, Cliona Stapleton, Rosalind A. ColemanAbstract:Abstract Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial and rate-limiting step in glycerolipid synthesis. Several mammalian GPAT activities have been recognized, including N-ethylmaleimide (NEM)-sensitive isoforms in microsomes and mitochondria and an NEM-resistant form in mitochondrial outer membrane (GPAT1). We have now cloned a second mitochondrial isoform, GPAT2 from mouse testis. The open-reading frame encodes a protein of 798 amino acids with a calculated mass of 88.8 kDa and 27% amino acid identity to GPAT1. Testis mRNA expression was 50-fold higher than in liver or brown adipose tissue, but the specific activity of NEM-sensitive GPAT in testis mitochondria was similar to that in liver. When Cos-7 cells were transiently transfected with GPAT2, NEM-sensitive GPAT activity increased 30%. Confocal microscopy confirmed a mitochondrial location. Incubation of GPAT2-transfected Cos-7 cells with trace (3 μM; 0.25 μCi) [1-14C]oleate for 6 h increased incorporation of [14C]oleate into TAG 84%. In contrast, incorporation into phospholipid species was lower than in control cells. Although a polyclonal antibody raised against full-length GPAT1 detected an ∼89-kDa band in liver and testis from GPAT1 null mice and both 89- and 80-kDa bands in BAT from the knockout animals, the GPAT2 protein expressed in Cos-7 cells was only 80 kDa. In vitro translation showed a single product of 89 kDa. Unlike GPAT1, GPAT2 mRNA abundance in liver was not altered by fasting or refeeding. GPAT2 is likely to have a specialized function in testis.
Maria R. Gonzalez-baro - One of the best experts on this subject based on the ideXlab platform.
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Glycerol-3-phosphate acyltransferase 2 is essential for normal spermatogenesis.
The Biochemical journal, 2017Co-Authors: Maria Belen Garcia-fabiani, Elizabeth Renee Cattaneo, Ezequiel Lacunza, Mauro A. Montanaro, Pablo Stringa, Marianela Santana, Magali Pellon-maison, Maria R. Gonzalez-baroAbstract:Glycerol-3-phosphate acyltransferases (GPATs) catalyze the first and rate-limiting step in the de novo glycerolipid synthesis. The GPAT2 isoform differs from the other isoforms because its expression is restricted to male germ cells and cancer cells. It has been recently reported that GPAT2 expression in mouse testis fluctuates during sexual maturation and that it is regulated by epigenetic mechanisms in combination with vitamin A derivatives. Despite progress made in this field, information about GPAT2 role in the developing male germ cells remains unclear. The aim of the present study was to confirm the hypothesis that GPAT2 is required for the normal physiology of testes and male germ cell maturation. The gene was silenced in vivo by inoculating lentiviral particles carrying the sequence of a short-hairpin RNA targeting Gpat2 mRNA into mouse testis. Histological and gene expression analysis showed impaired spermatogenesis and arrest at the pachytene stage. Defects in reproductive fitness were also observed, and the analysis of apoptosis-related gene expression demonstrated the activation of apoptosis in Gpat2-silenced germ cells. These findings indicate that GPAT2 protein is necessary for the normal development of male gonocytes, and that its absence triggers apoptotic mechanisms, thereby decreasing the number of dividing germ cells.
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Glycerol-3-phosphate acyltransferase 2 expression modulates cell roughness and membrane permeability: An atomic force microscopy study
PLoS ONE, 2017Co-Authors: Elizabeth R. Cattaneo, Mauro Aldo Montanaro, Eduardo Prieto, Maria B. Garcia-fabiani, Hervé Guillou, Maria R. Gonzalez-baroAbstract:In mammalian cells, de novo glycerolipid synthesis begins with the acylation of glycerol-3-phosphate, catalyzed by glycerol-3-phosphate acyltransferases (GPAT). GPAT2 is a mitochondrial isoform primarily expressed in testis under physiological conditions, and overexpressed in several types of cancers and cancer-derived human cell lines where its expression contributes to the tumor phenotype. Using gene silencing and atomic force microscopy, we studied the correlation between GPAT2 expression and cell surface topography, roughness and membrane permeability in MDA-MB-231 cells. In addition, we analyzed the glycerolipid composition by gas-liquid chromatography. GPAT2 expression altered the arachidonic acid content in glycerolipids, and the lack of GPAT2 seems to be partially compensated by the overexpression of another arachidonic-acid-metabolizing enzyme, AGPAT11. GPAT2 expressing cells exhibited a rougher topography and less membrane damage than GPAT2 silenced cells. Pore-like structures were present only in GPAT2 subexpressing cells, correlating with higher membrane damage evidenced by lactate dehydrogenase release. These GPAT2-induced changes are consistent with its proposed function as a tumor-promoting gene, and might be used as a phenotypic differentiation marker. AFM provides the basis for the identification and quantification of those changes, and demonstrates the utility of this technique in the study of cancer cell biology.