Publications

• Molecular identification of microsomal acyl-CoA:glycerol-3-phosphate acyltransferase, a key enzyme in de novo triacylglycerol synthesis.

  Cao J., Li J.-L., Li D., Tobin J.F., Gimeno R.E.

  Acyl-CoA:glycerol-3-phosphate acyltransferase (GPAT) catalyzes the first step during de novo synthesis of triacylglycerol. It has been well recognized that mammals possess multiple enzymatically distinct proteins with GPAT activity. Although the mitochondrial-associated GPAT has been cloned and ex ... >> More

  Acyl-CoA:glycerol-3-phosphate acyltransferase (GPAT) catalyzes the first step during de novo synthesis of triacylglycerol. It has been well recognized that mammals possess multiple enzymatically distinct proteins with GPAT activity. Although the mitochondrial-associated GPAT has been cloned and extensively characterized, the molecular identity of the endoplasmic reticulum (ER)-associated GPAT, which accounts for the majority of total GPAT activity in most tissues, has remained elusive. Here we report the identification of genes encoding human and mouse ER-associated GPAT (termed GPAT3). GPAT3 is a member of the acyltransferase family predominantly expressed in tissues characterized by active lipid metabolism, such as adipose tissue, small intestine, kidney, and heart. Ectopic expression of GPAT3 leads to a significant increase in N-ethylmaleimide-sensitive GPAT activity, whereas acyltransferase activity toward a variety of other lysophospholipids, as well as neutral lipid substrates, is not altered. Overexpression of GPAT3 in mammalian cells results in increased triacylglycerol, but not phospholipid, formation. GPAT3 is localized to the ER when overexpressed in COS-7 cells. GPAT3 mRNA is dramatically up-regulated during adipocyte differentiation, is reciprocally regulated in adipose tissue and liver of ob/ob mice, and is up-regulated in mice treated with a peroxisome proliferator-activated receptor gamma (PPARgamma) agonist. A substantial loss of GPAT activity in 3T3-L1 adipocytes was achieved by reducing GPAT3 mRNA levels through GPAT3-specific siRNA knockdown. These findings identify GPAT3 as a previously uncharacterized triacylglycerol biosynthetic enzyme. Similar to other lipogenic enzymes, GPAT3 may be useful as a target for the treatment of obesity. << Less

  Proc. Natl. Acad. Sci. U.S.A. 103:19695-19700(2006) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• Functional characterization of the human 1-acylglycerol-3-phosphate-O-acyltransferase isoform 10/glycerol-3-phosphate acyltransferase isoform 3.

  Sukumaran S., Barnes R.I., Garg A., Agarwal A.K.

  Synthesis of phospholipids can occur de novo or via remodeling of the existing phospholipids. Synthesis of triglycerides, a form of energy storage in cells, is an end product of these pathways. Several 1-acylglycerol-3-phosphate-O-acyltransferases (AGPATs) acylate lysophosphatidic acid (LPA) at th ... >> More

  Synthesis of phospholipids can occur de novo or via remodeling of the existing phospholipids. Synthesis of triglycerides, a form of energy storage in cells, is an end product of these pathways. Several 1-acylglycerol-3-phosphate-O-acyltransferases (AGPATs) acylate lysophosphatidic acid (LPA) at the sn-2 (carbon 2) position to produce phosphatidic acid (PA). These enzymes are involved in phospholipids and triglyceride synthesis through an evolutionary conserved process involving serial acylations of glycerol-3-phosphate. We cloned a cDNA predicted to be an AGPAT isoform (AGPAT10). This cDNA has been recently identified as glycerol-3-phosphate-O-acyltransferase isoform 3 (GPAT3). When this AGPAT10/GPAT3 cDNA was expressed in Chinese Hamster ovary cells, the protein product localizes to the endoplasmic reticulum. In vitro enzymatic activity using lysates of human embryonic kidney-293 cells infected with recombinant AGPAT10/GPAT3 adenovirus show that the protein has a robust AGPAT activity with an apparent V(max) of 2 nmol/min per mg protein, but lacks GPAT enzymatic activity. This AGPAT has similar substrate specificities for LPA and acyl-CoA as shown for another known isoform, AGPAT2. We further show that when overexpressed in human Huh-7 cells depleted of endogenous AGPAT activity by sh-RNA-AGPAT2-lentivirus, the protein again demonstrates AGPAT activity. These observations strongly suggest that the cDNA previously identified as GPAT3 has AGPAT activity and thus we prefer to identify this clone as AGPAT10 as well. << Less

  J. Mol. Endocrinol. 42:469-478(2009) [PubMed] [EuropePMC]

  This publication is cited by 11 other entries.

• AGPAT6 is a novel microsomal glycerol-3-phosphate acyltransferase.

  Chen Y.Q., Kuo M.-S., Li S., Bui H.H., Peake D.A., Sanders P.E., Thibodeaux S.J., Chu S., Qian Y.-W., Zhao Y., Bredt D.S., Moller D.E., Konrad R.J., Beigneux A.P., Young S.G., Cao G.

  AGPAT6 is a member of the 1-acylglycerol-3-phosphate O-acyltransferase (AGPAT) family that appears to be important in triglyceride biosynthesis in several tissues, but the precise biochemical function of the enzyme is unknown. In the current study, we show that AGPAT6 is a microsomal glycerol-3-ph ... >> More

  AGPAT6 is a member of the 1-acylglycerol-3-phosphate O-acyltransferase (AGPAT) family that appears to be important in triglyceride biosynthesis in several tissues, but the precise biochemical function of the enzyme is unknown. In the current study, we show that AGPAT6 is a microsomal glycerol-3-phosphate acyltransferase (GPAT). Membranes from HEK293 cells overexpressing human AGPAT6 had higher levels of GPAT activity. Substrate specificity studies suggested that AGPAT6 was active against both saturated and unsaturated long-chain fatty acyl-CoAs. Both glycerol 3-phosphate and fatty acyl-CoA increased the GPAT activity, and the activity was sensitive to N-ethylmaleimide, a sulfhydryl-modifying reagent. Purified AGPAT6 protein possessed GPAT activity but not AGPAT activity. Using [(13)C(7)]oleic acid labeling and mass spectrometry, we found that overexpression of AGPAT6 increased both lysophosphatidic acid and phosphatidic acid levels in cells. In these studies, total triglyceride and phosphatidylcholine levels were not significantly altered, although there were significant changes in the abundance of specific phosphatidylcholine species. Human AGPAT6 is localized to endoplasmic reticulum and is broadly distributed in tissues. Membranes of mammary epithelial cells from Agpat6-deficient mice exhibited markedly reduced GPAT activity compared with membranes from wild-type mice. Reducing AGPAT6 expression in HEK293 cells through small interfering RNA knockdown suggested that AGPAT6 significantly contributed to HEK293 cellular GPAT activity. Our data indicate that AGPAT6 is a microsomal GPAT, and we propose renaming this enzyme GPAT4. << Less

  J. Biol. Chem. 283:10048-10057(2008) [PubMed] [EuropePMC]

  This publication is cited by 4 other entries.

• Glycerol-3-phosphate acyltransferase-2 is expressed in spermatic germ cells and incorporates arachidonic acid into triacylglycerols.

  Cattaneo E.R., Pellon-Maison M., Rabassa M.E., Lacunza E., Coleman R.A., Gonzalez-Baro M.R.

  <h4>Background</h4>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 reag ... >> More

  <h4>Background</h4>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.<h4>Methods and results</h4>Incubation of GPAT2-transfected CHO-K1 cells with [1-(14)C]arachidonate for 3 h increased incorporation of [(14)C]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.<h4>Conclusions</h4>These results strongly suggest that GPAT2 expression is linked to arachidonoyl-CoA incorporation into TAG in spermatogenic germ cells. << Less

  PLoS ONE 7:e42986-e42986(2012) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.

• The initial step of the glycerolipid pathway: identification of glycerol-3-phosphate / dihydroxyacetone phosphate dual substrate acyltransferases in Saccharomyces cerevisiae.

  Zheng Z., Zou J.

  The initial step of phospholipid biosynthesis in yeast is carried out through the acylation of glycerol 3-phosphate (G-3-P) and dihydroxyacetone phosphate by stereospecific sn-1 acyltransferases. Here we report the identification of two key fatty acyltransferases of the glycerolipid biosynthesis p ... >> More

  The initial step of phospholipid biosynthesis in yeast is carried out through the acylation of glycerol 3-phosphate (G-3-P) and dihydroxyacetone phosphate by stereospecific sn-1 acyltransferases. Here we report the identification of two key fatty acyltransferases of the glycerolipid biosynthesis pathway in Saccharomyces cerevisiae. Disruption of the open reading frame YBL011w, corresponding to a gene previously identified as a choline transporter suppressor (SCT1), resulted in a substantial decrease of total cellular G-3-P acyltransferase activity. A yeast strain disrupted at the open reading frame YKR067w, which encodes a protein closely related to Sct1p, also exhibited a dramatic reduction in G-3-P acyltransferase activity. Molecular characterizations of the genes revealed that a missense mutation in YKR067w accounted for a defect in the activities of the G-3-P acyltransferase in the yeast mutant strain TTA1. Heterologous expression of YKR067w in Escherichia coli further confirmed its enzyme activity. These results indicate that YKR067w and YBL011w, designated herein as GAT1 and GAT2(SCT1), respectively, are yeast G-3-P acyltransferase genes. Furthermore, biochemical results are presented to show that both Gat1p and Gat2p(Sct1p) are G-3-P/dihydroxyacetone phosphate dual substrate-specific sn-1 acyltransferases. The fatty acyl specificity of Gat1p is similar to that of the mammalian microsomal G-3-P acyltransferase, as it can effectively utilize a broad range of fatty acids as acyl donors. In contrast, Gat2p(Sct1p) displayed preference toward 16-carbon fatty acids. The most notable of the altered phospholipid compositions of the gat1Delta and gat2(sct1)Delta strains are a decreased phosphatidic acid pool and an increased phosphatidylserine/phosphatidylinositol ratio. This did not appear to affect the mutants as no growth defect was found. However, null mutations of both GAT1 and GAT2(SCT1) are synthetically lethal to yeast. << Less

  J. Biol. Chem. 276:41710-41716(2001) [PubMed] [EuropePMC]

  This publication is cited by 5 other entries.