hPuf-A is a member of RNA-binding PUF family that regulates mRNA translation. Redistribution of hPuf-A from the nucleolus to the nucleoplasm upon genotoxic stress modulates the poly(ADP-ribosyl)ation activity of PARP-1. Here, we report a novel function of hPuf-A involved in promoting breast cancer progression. Immunohistochemical studies showed higher expression levels of hPuf-A in stage I, II, III, and IV breast cancer specimens in contrast with those of hPuf-A in ductal carcinoma in situ. The presence of hPuf-A is highly associated with colony formation capacities in breast cancer T47D and MDA-MB-231 cells. Xenograft growth of hPuf-A-silenced and hPuf-A overexpressing MDA-MB-231 cells in nude mice was substantially in concert with colony formation capacities. This promoting effect of hPuf-A in tumorigenesis might be correlated with the regulation of its associated mRNAs, such as RbAp48 and DDX3. Collectively, hPuf-A may have diagnostic values in breast cancer progression.