Updated project metadata. CVB 3C protease (3Cpro) plays a specific cleavage role on AU-rich binding factor (AUF1, also called as hnRNP D), consequently disputes the regulations of AUF1 on downstream molecules stabilities and transcriptions. In our work, iTRAQ approach was firstly used to quantitatively identify differentially expressed cellular proteins in AUF1-overexpressed HeLa cells, which would provide us a new insight into the role of AUF1 on cellular progress. We identified 1290 differentially expressed proteins (DEPs), including 882 upregulated and 408 downregulated DEPs at 24 h post infection (p.i.), The DEPs are involved in a variety of cellular functions such as metabolic processes, transcription, and RNA binding and exhibit different subcellular localization via GO term, protein interaction network, and series analysis. Among DEPs, DDX5, POLR2, and PPP2CA showed important roles in cellular metabolism. Especially, DDX5 was negatively regulated by AUF1 and increased during CVB infection, which in turns played an inhibitory effect on CVB replication. These findings are particularly important in the development new anti-viral therapy strategies targeting on series of new molecules.