Normal cell cycle progression and proliferation of palatal mesenchymal cells are important for palatal development. As targets of miR-17-92, E2F transcription factors family has been suggested to induce the transcription of miR-17-92 in several cell types. In the present study, we sought to investigate whether this negative feedback loop exists in mouse PMCs and what the function of this negative feedback loop would be in palatal mesenchymal cells. Using GeneMANIA, we revealed that the most important function of experimentally verified targets of miR-17-92 is cell cycle regulation. E2F1 and E2F3, but not E2F2, were extensively expressed in mouse palate. Over-expression of E2F1 significantly increased the expression of all the members of miR-17-92. After increased by E2F1, miR-17 and miR-20a may negatively target E2F1, and thereby prevent the cells from excessive proliferation. We suggest that the negative feedback loop between E2F1 and miR-17-92 may contribute to palatal development by regulating the proliferation and cell cycle of palatal mesenchymal cells.