After observing that expression of two NR4A orphan nuclear receptors, NR4A3 and NR4A1, was altered by insulin in cDNA microarray analyses of human skeletal muscle, we studied whether these receptors could modulate insulin sensitivity. We found that both NR4A3 and NR4A1 were induced by insulin and by thiazolidinedione drugs (pioglitazone and troglitazone) in 3T3-L1 adipocytes. Furthermore, gene expression of NR4A3 and NR4A1 was reduced in skeletal muscles and adipose tissues from multiple rodent models of insulin resistance. To determine whether NR4A3 could modulate insulin sensitivity, 3T3-L1 adipocytes were stably transduced with NR4A3 or LacZ (control) lentiviral vectors. Compared with LacZ expressing cells, hyperexpression of NR4A3 increased the ability of insulin to augment glucose transport activity, and the mechanism involved increased recruitment of GLUT4 glucose transporters to the plasma membrane. NR4A3 hyperexpression also led to an increase in insulin-mediated tyrosine phosphorylation of insulin receptor substrate-1 as well as Akt phosphorylation. Suppression of NR4A3 using lentiviral short hairpin RNA constructs reduced the ability of insulin to stimulate glucose transport and phosphorylate Insulin receptor substrate-1 and Akt. Thus, NR4A3 and NR4A1 are attractive novel therapeutic targets for potential amelioration of insulin resistance, and treatment and prevention of type 2 diabetes and the metabolic syndrome.