Ion channels like voltage gated ether á gogo (Eag1) K+ channels or Ca2+ activated Cl- channels have been shown to support cell proliferation. Bestrophin 1 (Best1) has been proposed to form Ca2+ activated Cl- channels in epithelial cells. Here we show that original T84 colonic carcinoma cells grow slow (T84-slow) and express low amounts of Eag1 and Best1, while spontaneously transformed T84 cells grow fast (T84-fast) and express high levels of both proteins. Both Eag1 and Best1 currents are upregulated in T84-fast cells. Eag1 currents were cell cycle dependent, with upregulation during G1/S transition. T84-slow but not T84-fast formed tight monolayers when grown on permeable supports. RNAi inhibition of Eag1 and Best1 reduced proliferation of T84-fast cells, while overexpression of Best1 turned T84-slow into fast growing cells. Eag1 and Best1 improve intracellular Ca2+ signaling and cell volume regulation. These results establish a novel role of bestrophins for cell proliferation.