Abstract—Transforming growth factor-&bgr; (TGF-&bgr;) increases expression of endothelial nitric oxide synthase (eNOS), although the precise mechanism by which it does so is unclear. We report that Smad2, a transcription factor activated by TGF-&bgr;, mediates TGF-&bgr; induction of eNOS in endothelial cells. TGF-&bgr; induces Smad2 translocation from cytoplasm to nucleus, where it directly interacts with a specific region of the eNOS promoter. Overexpression of Smad2 increases basal levels of eNOS, and further increases TGF-&bgr; stimulation of eNOS expression. Ectopic expression of Smurf, an antagonizer of Smad2, decreases Smad2 expression and blocks TGF-&bgr; induction of eNOS. Because Smad2 can interact with a variety of transcription factors, coactivators, and corepressors, Smad2 may thus act as an integrator of multiple signals in the regulation of eNOS expression.