A Regulatory Role for 1-Acylglycerol-3-phosphate-O-acyltransferase 2 in Adipocyte Differentiation

Abstract

Mutations in the 1-acylglycerol-3-phosphate-O-acyltransferase 2 (AGPAT2) gene have been identified in individuals affected with congenital generalized lipodystrophy (CGL). AGPAT2 catalyzes acylation of lysophosphatidic acid to phosphatidic acid, a precursor for both triacylglycerol (TAG) and phospholipid synthesis. Recent studies suggest that reduced AGPAT2 enzymatic activity may underlie the CGL clinical phenotype. To gain insight into how altered AGPAT2 activity causes lipodystrophy, we examined the effect of knockdown of AGPAT2 expression in preadipocytes on TAG synthesis and storage, and on adipocyte differentiation. We show that AGPAT2 mRNA expression is induced 30-fold during adipocyte differentiation and that AGPAT2 enzymatic activity is required for TAG mass accumulation in mature adipocytes. We demonstrate that small interference RNA-mediated knockdown of AGPAT2 expression prevents appropriate early induction of C/EBP␤ and PPAR␥, key transcriptional activators of the adipogenic program, and delays expression of multiple adipocyte-related genes. The unexpected finding, that levels of several phospholipid species, including phosphatidic acid (PA), are elevated in TAG-depleted adipocytes with AGPAT2 knockdown, suggests that impaired AGPAT2 activity affects availability of PA for TAG synthesis but not overall PA synthesis nor utilization of PA for phospholipid synthesis. These findings underscore the importance of an AGPAT2-mediated metabolic pathway in adipocyte differentiation. Congenital generalized lipodystrophy (CGL),3 also referred to as Berardinelli-Seip syndrome, is an autosomal recessive disorder characterized by paucity of adipose tissue, severe insulin resistance, hypertriglyceridemia, and hepatic steatosis in childhood (1). Positional cloning strategies have identified mutations in the Berardinelli-Seip congential lipodistrophy 2 gene, which encodes the seipin protein of unknown function, and in the 1-acylglycerol-3-phosphate-O-acyltransferase 2 (AGPAT2) gene in individuals affected with CGL (2, 3). AGPAT2 (also referred to as lysophosphatidic acid acyltransferase or LPAAT-␤) encodes an enzyme that catalyzes the acylation of lysophosphatidic acid (LPA) to phosphatidic acid (PA), which serves as a precursor for triacylglycerol (TAG) and phospholipid synthesis (4, 5).AGPAT2 is one of five murine lysophosphatidic acid acyltransferase isoforms that have been identified (AGPAT1-5), each of which contains a highly conserved catalytic (NHX 4 D) and substrate binding motif (EGTR) (6). Although there are differences in the tissue distribution of the AGPAT isoforms, the physiological significance of this diversity is not understood. AGPAT1 and AGPAT3 are ubiquitously expressed, whereas AGPAT2, AGPAT4, and AGPAT5 are expressed in a tissue-specific manner (6). In human tissues AGPAT2 is highly expressed in liver, pancreas, skeletal muscle, and small intestine (7) and is the most highly expressed AGPAT isoform in adipose tissue (3).Structure-function studies of AGPAT2 mutations identified in CGL patients dem...

Hepatic Gluconeogenesis Is Enhanced by Phosphatidic Acid Which Remains Uninhibited by Insulin in Lipodystrophic Agpat2−/− Mice

“…Similar observations were reported in cultured OP9 cells, a cultured cell model for adipogenesis (8). Inhibiting Agpat2 expression in these cells using siRNA indicated that additional pathway(s) might exist to generate PA (8).…”

“…Surprisingly, total glycerophospholipid levels in the livers of Agpat2 Ϫ/Ϫ mice were not very different from those of WT mice (7). Similar observations were reported in cultured OP9 cells, a cultured cell model for adipogenesis (8). Inhibiting Agpat2 expression in these cells using siRNA indicated that additional pathway(s) might exist to generate PA (8).…”

Pathology of Congenital Generalized Lipodystrophy in Agpat2–/– Mice

“…In vitro studies have shown that AGPAT2 is involved in the metabolism and differentiation of adipocytes; as such, knockdown of AGPAT2 expression reduces the synthesis and storage of triglycerides in preadipocytes and adipocytes, thus preventing the development of normal adipocyte morphology. 15 The abnormal appearance and reduced numbers of WAT adipocytes that we Figure 27. Pancreas, islet of Langerhans; aged wild-type (WT) Agpat2 þ/þ mouse.…”

Lipodystrophy: metabolic insights from a rare disorder

“…The failure of TAG synthesis therefore provides a very plausible explanation for lipodystrophy in patients with genetic defects in the AGPAT2 gene. However, cellular studies have also demonstrated that disruption of AGPAT2 or BSCL2 inhibits not only lipid synthesis but also the normal induction of adipogenic gene expression during fat cell development (Gale et al 2006, Payne et al 2008. This may reflect the tight co-regulation of gene expression and lipid droplet formation in developing adipocytes or indicate that these proteins possess functions in addition to their proposed roles in lipogenesis.…”