E1AF transcription factor, a member of Ets family, is deregulated in many tumors and widely known to play critical roles in tumor metastasis via directly binding to the promoter of genes involved in tumor migration and invasion. Here, we found that E1AF overexpression promoted breast cancer cell cycle progression and growth in vivo as well as the transcription of cell cycle-related protein Cyclin D3. And, the interference of Cyclin D3 expression by transfecting with Cyclin D3 RNAi inhibited the positive role of E1AF in cell cycle progression. We further showed that decreasing the expression of E1AF by E1AF RNAi reduced Cyclin D3 transcription and expression, and inhibited cell cycle progression that was abrogated by Cyclin D3 overexpression. Taken together, E1AF increases cell cycle progression via upregulation of Cyclin D3 transcription, which elicits a new mechanism of breast cancer growth and a new mechanism of Cyclin D3 transcription.