The present study showed that Transforming growth factor beta 1 (TGF-${eta}_1$) can induce apoptosis of bovine mammary epithelial cells. This apoptosis was also observed with phosphorylation of Smad2/3 within 0.5-2 h. Afterwards the signal transferred into the nucleus. Moreover, intracellular free $Ca^{2+}$ concentration was significantly elevated as well as Caspase-3 activated and DNA lysised, thereby inducing the programmed cell death. This signaling pathway of TGF-${eta}_1$ was blocked by SB-431542 ($10^{-2}{mu}M$) via inhibiting ALK-5 kinase activity, which thus reversed the anti-proliferation and apoptosis effect of TGF-${eta}_1$ in mammary epithelial cells. These results indicated that TGF-${eta}_1$ induced apoptosis of bovine mammary epithelial cells through the ALK-5-Smad2/3 pathway, which plays an important role in inhibiting survival of mammary epithelial cells. Moreover, intracellular $Ca^{2+}$ also played a critical role in TGF-${eta}_1$-induced cell apoptosis.