Retinoic acid (RA) plays a key role in pluripotent cell differentiation. In F9 embryonic carcinoma cells, RA can induce differentiation towards somatic lineages via the Ras-extracellular signal-regulated kinase (Ras/Erk) pathway, but the mechanism through which it induces the Erk1/2 phosphorylation is unclear. Here, we show that miR-485 is a positive regulator that targets α/β-hydrolase domain-containing protein 2 (Abhd2), which can result in Erk1/2 phosphorylation and triggers differentiation. RA up-regulates miR-485 and concurrently down-regulates Abhd2. We verified that Abhd2 is targeted by miR-485 and they both can influence the phosphorylation of Erk1/2. In summary, RA can mediate cell differentiation by phosphorylating Erk1/2 via miR-485 and Abhd2.