Mitochondrial Glycerol Phosphate Acyltransferase Directs the Incorporation of Exogenous Fatty Acids into Triacylglycerol

Igal, R. Ariel; Wang, Shuli; González-Baró, María R.; Coleman, Rosalind A.

doi:10.1074/jbc.m103386200

Cited by 82 publications

(75 citation statements)

References 36 publications

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“…We (25) have shown that a 3.8-fold increase in mtGPAT1 activity in CHO cells increases TAG content 2.7-fold and [ 14 C]oleate incorporation into TAG 3.4-fold. Although that study strongly suggested that mtGPAT1 directs fatty acid toward TAG synthesis, CHO cells provide limited information because their oxidation rate is low and exogenous fatty acids are primarily metabolized to glycerolipids.…”

Section: Discussionmentioning

confidence: 99%

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Mitochondrial glycerol-3-phosphate acyltransferase-1 directs the metabolic fate of exogenous fatty acids in hepatocytes

Lewin¹,

Wang²,

Nagle³

et al. 2005

American Journal of Physiology-Endocrinology and Metabolism

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(TAG) in nonadipose tissues is closely associated with the development of insulin resistance, interest has increased in the metabolism of longchain acyl-CoAs toward ␤-oxidation or the synthesis and storage of TAG. To learn whether a mitochondrial isoform of glycerol-3-phosphate acyltransferase (mtGPAT1) competes with carnitine palmitoyltransferase I (CPT I) for acyl-CoAs and whether it contributes to the formation of TAG, we overexpressed rat mtGPAT1 13-fold in primary hepatocytes obtained from fasted rats. When 100, 250, or 750 M oleate was present, both TAG mass and the incorporation of [ 14 C]oleate into TAG increased more than twofold in hepatocytes overexpressing mtGPAT1 compared with vector controls. Although the incorporation of [ 14 C]oleate into CO2 and acid-soluble metabolites increased with increasing amounts of oleate in the media, these metabolites were ϳ40% lower in the Ad-mtGPAT1 infected cells, consistent with competition for acyl-CoAs between CPT I and mtG-PAT1. A 50 -60% decrease was also observed in [14 C]oleate incorporation into cholesteryl ester. With increasing amounts of exogenous oleate, [14 C]TAG secretion increased appropriately in vector controlinfected hepatocytes, suggesting that the machinery for VLDL-TAG biogenesis and secretion was unaffected. Despite the marked increases in TAG synthesis and storage in the Ad-mtGPAT1 cells, however, the Ad-mtGPAT1 cells secreted the same amount of [ 14 C

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Section: Discussionmentioning

confidence: 99%

“…14 C]oleate incorporation into TAG 3.4-fold (25). In the Ad-GPAT1-infected hepatocytes, a 13-fold increase in GPAT activity resulted in 2.4-, 3-, and 2.4-fold increases in TAG mass at 100, 250, and 750 M oleate, respectively (Fig.…”

Section: Adenoviral Mediated Overexpression Of Mtgpat1 In Hepatocytesmentioning

confidence: 99%

Mitochondrial glycerol-3-phosphate acyltransferase-1 directs the metabolic fate of exogenous fatty acids in hepatocytes

Lewin¹,

Wang²,

Nagle³

et al. 2005

American Journal of Physiology-Endocrinology and Metabolism

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“…This result was surprising for several reasons. First, expression of mitochondrial GPAT1 or the newly identified mitochondrial GPAT2 increases TG synthesis in cultured cells (14,28). Second, GPAT3 has been reported by others to increase triglyceride synthesis (14).…”

Section: Discussionmentioning

confidence: 99%

AGPAT6 Is a Novel Microsomal Glycerol-3-phosphate Acyltransferase

Chen

Kuo

et al. 2008

Journal of Biological Chemistry

128

141

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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 wildtype 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.

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“…The role for mtGPAT in up-regulating TAG synthesis is also supported by studies of overexpressed rat mtGPAT in Chinese hamster ovary cells. Expression of recombinant rat mtGPAT in Chinese hamster ovary cells results in a 3.8-fold increase in NEM-resistant GPAT activity, a 4-fold increase in [ 14 C]oleate incorporation into TAG, and a 30% decrease in [ 14 C]oleate incorporation into phospholipids (10). To confirm the critical role of mtGPAT in the regulation of TAG synthesis, mtGPAT null mice were generated (8).…”

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Identification of a New Glycerol-3-phosphate Acyltransferase Isoenzyme, mtGPAT2, in Mitochondria

Lewin

Schwerbrock

Lee

et al. 2004

Journal of Biological Chemistry

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Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial and rate-limiting step of glycerolipid synthesis. Two distinct GPAT isoenzymes had been identified in mammalian tissues, an N-ethylmaleimide (NEM)-sensitive isoform in the endoplasmic reticulum membrane (microsomal GPAT) and an NEM-resistant form in the outer mitochondrial membrane (mtGPAT). Although only mtGPAT has been cloned, the microsomal and mitochondrial GPAT isoforms can be distinguished, because they differ in acyl-CoA substrate preference, sensitivity to inhibition by dihydroxyacetone phosphate and polymixin B, temperature sensitivity, and ability to be activated by acetone. The preponderance of evidence supports a role for mtGPAT in synthesizing the precursors for triacylglycerol synthesis. In mtGPAT ؊/؊ mice, PCR genotyping and Northern analysis showed successful knockout of mtGPAT; however, we detected a novel NEM-sensitive GPAT activity in mitochondrial fractions and an anti-mtGPAT immunoreactive protein in liver mitochondria, but not in microsomes. Rigorous analysis using two-dimensional gel electrophoresis revealed that the anti-mtGPAT immunoreactive proteins in wild type and mtGPAT ؊/؊ liver mitochondria have different isoelectric points. These results suggested the presence of a second GPAT in liver mitochondria from mtGPAT ؊/؊ mice. Characterization of this GPAT activity in liver from mtGPAT null mice showed that, unlike the mtG-PAT activity in wild type samples, activity in mtGPAT knockout mitochondria did not prefer palmitoyl-CoA, was sensitive to inactivation by NEM, was inhibited by dihydroxyacetone phosphate and polymixin B, was temperature-sensitive, and was not activated by acetone. We conclude that a novel GPAT (mtGPAT2) with antigenic epitopes similar to those of mtGPAT is detectable in mitochondria from the livers of mtGPAT ؊/؊ mice.The initial and rate-limiting step of glycerolipid synthesis is the acylation of glycerol 3-phosphate with long-chain fatty acylCoA to form 1-acyl-glycerol 3-phosphate (LPA). 1 This reaction is catalyzed by two glycerol-3-phosphate acyltransferase (GPAT; EC 2.3.1.15) isoenzymes that are encoded by different genes (1). One isoform is present in the endoplasmic reticulum membrane (microsomal GPAT), and the other is located in the outer mitochondrial membrane (mtGPAT). Although the microsomal GPAT has not been cloned or purified, its activity is easily distinguished, because, unlike mtGPAT, the microsomal isoform is inhibited by sulfhydryl reagents (2). In most tissues, the microsomal GPAT activity is 10 times higher than that found in the mitochondrial fraction, but in liver, mtGPAT contributes 30 -50% of the total activity (1).Microsomal GPAT and mtGPAT also differ in their acyl-CoA substrate preference. The microsomal isoform esterifies both saturated and unsaturated long-chain acyl-CoAs equally well, but mtGPAT prefers C16:0-CoA (2). In rat liver, kidney, and heart, mtGPAT activity is 3-10-fold higher with C16:0-CoA than with other long-chain saturated or unsaturated acyl-CoA substrates (3...

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Mitochondrial Glycerol Phosphate Acyltransferase Directs the Incorporation of Exogenous Fatty Acids into Triacylglycerol

Cited by 82 publications

References 36 publications

Mitochondrial glycerol-3-phosphate acyltransferase-1 directs the metabolic fate of exogenous fatty acids in hepatocytes

Mitochondrial glycerol-3-phosphate acyltransferase-1 directs the metabolic fate of exogenous fatty acids in hepatocytes

AGPAT6 Is a Novel Microsomal Glycerol-3-phosphate Acyltransferase

Identification of a New Glycerol-3-phosphate Acyltransferase Isoenzyme, mtGPAT2, in Mitochondria

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