Natural Killer (NK) cell function is compromised by prostaglandin E₂ (PGE₂), but very little is known about the mechanism by which PGE₂ affects NK effector activity. Specifically, nothing is known regarding which PGE₂ receptor (EP1-4) mediates these effects. We have examined the role of individual EPs in regulating NK cells. Murine splenic or human NK cells express all four EP1-4 receptors. In endogenous NK cells from normal mice (N-NK), we show that activating all four NK-EP receptors with PGE₂ leads to less migration, reduced ability to lyse tumor targets, inhibited IFNγ secretion and decreased TNFα production. The ability of PGE₂ to inhibit N-NK cells is most likely through the EP4 receptor and, to a lesser degree, the EP2 receptor. Like PGE₂, the EP4 agonist PGE1-OH blocked NK cell migration, inhibited cytotoxicity, and prevented cytokine secretion. The EP2 agonist, Butaprost, was able to inhibit cytotoxicity but did not blunt migration or effectively inhibit IFNγ secretion. In contrast to the inhibitory actions of PGE₂, the EP1/EP3 agonist, Sulprostone, increased migration of N-NK cells. Thus, EP4 and EP1/3 may have opposing roles in regulating NK cells. NK cells from tumor bearing mice (T-NK) were compromised in many functions and showed reduced EP receptor expression. PGE₂ inhibits the potential of T-NK cells to migrate, exert cytotoxic effects, and secrete IFNγ. This ability of PGE₂ to inhibit NK cells from tumor bearing mice is mimicked by EP2 and EP4 agonists and, therefore most likely through EP2 and EP4 receptors. T-NK cells stimulated with PGE₂, EP2 agonist (Butaprost), and EP4 agonist (PGE1-OH) were more sensitive to inhibition compared to N-NK cells. In contrast to the inhibitory effects of PGE₂ on cytotoxicity, and IFNγ production, TNFα secretion was actually induced in T-NK. Thus, PGE₂ inhibits TNFα secretion from N-NK, and further increases the constitutively high TNFα secretion from T-NK. Taken together our results show that tumor-derived PGE₂ is able to suppress many NK functions that are critical to tumor control and much of this inhibition is mediated through the EP4 receptor on NK cells. These data, combined with our published studies showing that tumor-expressed EP4 promotes metastasis, support the rationale to develop more effective EP antagonists as novel therapeutic agents.