Autotaxin (ATX), or nucleotide pyrophosphatase-phosphodiesterase 2, is a secreted lysophospholipase D that promotes cell migration, metastasis, and angiogenesis. ATX generates lysophosphatidic acid (LPA), a lipid mitogen and motility factor that acts on several G protein-coupled receptors. Here we report that ATX-deficient mice die at embryonic day 9.5 (E9.5) with profound vascular defects in yolk sac and embryo resembling the G␣ 13 knockout phenotype. Furthermore, at E8.5, ATX-deficient embryos showed allantois malformation, neural tube defects, and asymmetric headfolds. The onset of these abnormalities coincided with increased expression of ATX and LPA receptors in normal embryos. ATX heterozygous mice appear healthy but show half-normal ATX activity and plasma LPA levels. Our results reveal a critical role for ATX in vascular development, indicate that ATX is the major LPA-producing enzyme in vivo, and suggest that the vascular defects in ATX-deficient embryos may be explained by loss of LPA signaling through G␣ 13 .Autotaxin (ATX), also known as ectonucleotide pyrophosphatase-phosphodiesterase 2, belongs to the nucleotide pyrophosphatase (NPP) family of ectoenzymes and exoenzymes, originally defined by their ability to hydrolyze nucleotides in vitro (8,15,44). Full-length ATX is cleaved along the classical export pathway and secreted as a catalytically active glycoprotein (21, 52). ATX was initially isolated as an autocrine motility factor for melanoma cells (45) and later found to promote metastasis and tumor vascularization in nude mice as well as eliciting an angiogenic response in Matrigel assays (31, 32). Hence, ATX may contribute to tumor progression by providing an invasive and/or angiogenic microenvironment for both malignant and stromal cells, a notion supported by growing evidence that ATX expression is upregulated in various invasive and metastatic cancers (4,18,22,28,43,55).The physiological substrate of ATX had remained elusive until it was discovered that ATX is identical to lysophospholipase D (lysoPLD), a secreted enzyme present in plasma and conditioned media that converts lysophosphatidylcholine (LPC) into bioactive lysophosphatidic acid (LPA) (11,47,48). LPA stimulates cell proliferation, migration, and survival by acting on specific G protein-coupled receptors (GPCRs) that are linked to multiple G proteins, including G q/11 , G i/o , and G 12/13 (20,30). LPA promotes wound healing in vivo and has been implicated in tumor progression, inflammation, vascular disease, and neural development (5,23,28,42,51). It has now become clear that LPA production, rather than nucleotide metabolism, accounts for the growth factor-like effects of ATX observed in cell culture. Strikingly, the other NPP family members lack intrinsic lysoPLD activity despite the similarity between their catalytic domain and that of ATX (14), implying that ATX/NPP2 is a unique lysoPLD with no functional redundancy within the NPP family.In addition to converting LPC into LPA, ATX can also hydrolyze sphingosyl-phosphorycholine (...