The long terminal repeats of murine intracisternal A particles (IAPs) contain an IAP proximal enhancer (IPE) element that is inactive in murine F9 embryonal carcinoma cells and active in the parietal endoderm cell line PYS-2. The element binds efficiently to a 60-kDa IPE-binding protein (IPEB) present in PYS-2 cells but poorly to F9 proteins, suggesting a role for IPEB in regulating IAP expression. We have purified calf thymus IPEB, which binds to the IPE and transactivates a reporter gene in HeLa cell extracts. Based on the peptide sequence of the purified calf IPEB, we have cloned a 420-bp cDNA and showed that the encoded protein is the homolog of human p54^nrb and mouse NonO, which are characterized by the presence of two RNA recognition motifs. We show that p54^nrb is an IPE-binding transcription activator with its DNA-binding and activation domains in the N- and C-terminal halves, respectively. The activation domain of p54^nrb is active in HeLa, PYS-2, and F9 cells, whereas p54^nrb as a whole molecule is active in HeLa and PYS-2 cells but not in F9 cells. Thus, the lack of activity of p54^nrb in F9 cells is due to an ineffective DNA-binding domain. We demonstrate that p54^nrb also binds to a pre-mRNA. Based on the close sequence relatedness of this protein to PSF, which is required for pre-mRNA splicing in vitro, we discuss the possibility that p54^nrb has dual roles in transcription and splicing.