Autotaxin (ATX) is a cancer-associated motogen that has multiple biological activities in vitro through the production of bioactive small lipids, lysophosphatidic acid (LPA). ATX and LPA are abundantly present in circulating blood. However, their roles in circulation remain to be solved. To uncover the physiological role of ATX we analyzed ATX knock-out mice. In ATXnull embryos, early blood vessels appeared to form properly, but they failed to develop into mature vessels. As a result ATX-null mice are lethal around embryonic day 10.5. The phenotype is much more severe than those of LPA receptor knock-out mice reported so far. In cultured allantois explants, neither ATX nor LPA was angiogenic. However, both of them helped to maintain preformed vessels by preventing disassembly of the vessels that was not antagonized by Ki16425, an LPA receptor antagonist. In serum from heterozygous mice both lysophospholipase D activity and LPA level were about half of those from wild-type mice, showing that ATX is responsible for the bulk of LPA production in serum. The present study revealed a previously unassigned role of ATX in stabilizing vessels through novel LPA signaling pathways. Autotaxin (ATX)2 is a motogen-like phosphodiesterase originally isolated from conditioned medium of human melanoma cells (1). Enforced expression of ATX in Ras-transformed NIH3T3 cells greatly enhances their invasive, tumorigenic, and metastatic potentials (2). In addition, enhanced expression of ATX has been demonstrated in various malignant tumor tissues (3). Thus, ATX is implicated in tumorigenic and metastatic potentials of cancer cells. ATX is also expressed in various tissues and is present at high concentration in various biological fluids including plasma, serum, and seminal plasma (4), implying specific roles of ATX in circulation.Recently, ATX was shown to have lysophospholipase D (lysoPLD) activity, which converts lysophosphatidylcholine to a bioactive lysophospholipid, lysophosphatidic acid (LPA) (5, 6). ATX also converts sphingosylphosphorylcholine into another bioactive lysophospholipid, sphingosine 1-phosphate (S1P) in vitro (7). Because LPA and S1P are regulators of cell motility and proliferation in various cell systems, they might be the effectors of the motogenic actions of ATX. LPA and S1P have been shown to have diverse roles in many biological processes that are mediated by G protein-coupled receptors (GPCRs) specific to LPA or S1P; there are five GPCRs for LPA (LPA 1-5 ) and five for S1P (S1P 1-5 ) with a number of putative GPCRs (8). Thus, ATX may exert its functions through these receptors. Indeed, ATX stimulates cell motility of tumor cells through one of the LPA receptors, LPA 1 (9), and ATX positively or negatively modulates cell motility depending on S1P receptor subtypes (7, 10). To uncover the physiological role of ATX and to identify the endogenous product of ATX, we investigated ATX knock-out mice. In this study we show that ATX produces LPA, but not S1P, in circulating blood and that it contributes to blood vess...

Autotaxin (ATX, also known as Enpp2) is a secreted lysophospholipase D that hydrolyzes lysophosphatidylcholine to generate lysophosphatidic acid (LPA), a lipid mediator that activates G protein-coupled receptors to evoke various cellular responses. Here, we report the crystal structures of mouse ATX alone and in complex with LPAs with different acyl-chain lengths and saturations. These structures reveal that the multidomain architecture helps to maintain the structural rigidity of the lipid-binding pocket, which accommodates the respective LPA molecules in distinct conformations. They indicate that a loop region in the catalytic domain is a major determinant for the substrate specificity of the Enpp family enzymes. Furthermore, along with biochemical and biological data, these structures suggest that the produced LPAs are delivered from the active site to cognate G protein-coupled receptors through a hydrophobic channel.

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Autotaxin (ATX) is a multifunctional phosphodiesterase originally isolated from melanoma cells as a potent cell motility-stimulating factor. ATX is identical to lysophospholipase D, which produces a bioactive phospholipid, lysophosphatidic acid (LPA), from lysophosphatidylcholine (LPC). Although enhanced expression of ATX in various tumor tissues has been repeatedly demonstrated, and thus, ATX is implicated in progression of tumor, the precise role of ATX expressed by tumor cells was unclear. In this study, we found that ATX is highly expressed in glioblastoma multiforme (GBM), the most malignant glioma due to its high infiltration into the normal brain parenchyma, but not in tissues from other brain tumors. In addition, LPA 1 , an LPA receptor responsible for LPAdriven cell motility, is predominantly expressed in GBM. One of the glioblastomas that showed the highest ATX expression (SNB-78), as well as ATX-stable transfectants, showed LPA 1 -dependent cell migration in response to LPA in both Boyden chamber and wound healing assays. Interestingly these ATX-expressing cells also showed chemotactic response to LPC. In addition, knockdown of the ATX level using small interfering RNA technique in SNB-78 cells suppressed their migratory response to LPC. These results suggest that the autocrine production of LPA by cancer cell-derived ATX and exogenously supplied LPC contribute to the invasiveness of cancer cells and that LPA 1 , ATX, and LPC-producing enzymes are potential targets for cancer therapy, including GBM.

Enpp1 is a membrane-bound glycoprotein that regulates bone mineralization by hydrolyzing extracellular nucleotide triphosphates to produce pyrophosphate. Enpp1 dysfunction causes human diseases characterized by ectopic calcification. Enpp1 also inhibits insulin signaling, and an Enpp1 polymorphism is associated with insulin resistance. However, the precise mechanism by which Enpp1 functions in these cellular processes remains elusive. Here, we report the crystal structures of the extracellular region of mouse Enpp1 in complex with four different nucleotide monophosphates, at resolutions of 2.7-3.2 Å. The nucleotides are accommodated in a pocket formed by an insertion loop in the catalytic domain, explaining the preference of Enpp1 for an ATP substrate. Structural mapping of disease-associated mutations indicated the functional importance of the interdomain interactions. A structural comparison of Enpp1 with Enpp2, a lysophospholipase D, revealed marked differences in the domain arrangements and active-site architectures. Notably, the Enpp1 mutant lacking the insertion loop lost the nucleotide-hydrolyzing activity but instead gained the lysophospholipid-hydrolyzing activity of Enpp2. Our findings provide structural insights into how the Enpp family proteins evolved to exert their diverse cellular functions. molecular evolution | X-ray crystallography

Body weight is tightly regulated by food intake and energy dissipation, and obesity is related to decreased energy expenditure (EE). Herein, we show that nucleotide pyrophosphatase/phosphodiesterase 2 (ENPP2, autotaxin) is an adipose-derived, secreted enzyme that controls adipose expansion, brown adipose tissue (BAT) function, and EE. In mice, Enpp2 was highly expressed in visceral white adipose tissue and BAT and is downregulated in hypertrophied adipocytes/adipose tissue. Enpp2 +/2 mice and adipocyte-specific Enpp2 knockout mice fed a highfat diet showed smaller body weight gains and less insulin resistance than control mice fed the same diet. BAT was functionally more active and EE was increased in Enpp2-deficient mice. In humans, ENPP2 expression in subcutaneous fat and ENPP2 levels in serum were reduced in obese subjects. Taken together, our results establish ENPP2 as an adipose-derived, secreted enzyme that regulates adipose obesity and systemic metabolism. They also suggest ENPP2 could be a useful therapeutic target for the treatment of metabolic disease.Until recently, adipose tissue was viewed as a passive energy storage organ, but with the discovery of leptin and the adipose-derived humoral factors now known as "adipokines," it has become apparent that adipose tissue is an active endocrine organ that is essential for energy homeostasis (1). Moreover, obese adipose tissue secretes various inflammatory cytokines, including interleukin-6 (IL-6) and tumor necrosis factor-a (TNF-a), whose activities are known to contribute to the development of metabolic and cardiovascular diseases (2).Enpp2, also designated autotaxin, phosphodiesterase I a/autotaxin, and nucleotide pyrophosphatase/phosphodiesterase 2, was originally discovered as an autocrine motility-stimulating factor released from cancer cells (3). ENPP2 catalyzes the conversion of lysophosphatidylcholine to lysophosphatidic acid (LPA), which exerts a variety of biological effects, in part via G-protein-coupled receptors (3,4). In addition, the COOH-terminal noncatalytic domain of ENPP2 also has biological effects independent of LPA (3). Homozygous Enpp2-deficient mice die in utero due to profound vascular defects, but heterozygous Enpp2-deficient (Enpp2 +/2 ) mice are apparently healthy, with plasma LPA levels about half those in wild-type (WT) mice (5). Enpp2 is reportedly expressed in mouse adipose tissue and 3T3-F442A preadipocytes, and medium conditioned by Enpp2-expressing COS7 cells increased proliferation of 3T3-F442A cells. Recently, Dusaulcy et al. (6) reported that adipocytespecific Enpp2 knockout (KO) mice fed a high-fat diet showed greater adiposity and less systemic insulin resistance

The serum ATX activity and plasma LPA level are increased in chronic hepatitis C in association with liver fibrosis. Our study may provide the first evidence showing a significant increase of both ATX and LPA in the blood under a specific disease.

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