Protein synthesis initiation factor 4AI (eIF-4AI) is ubiquitously expressed in eucaryotic cells. We characterized the eIF-4AI gene promoter cloned from human fibroblasts. The minimal promoter, localized to a region in the 5'-noncoding region adjacent to the first exon, consisted of approximately 300 base pairs (bp), 80% of which were identical with those in the corresponding mouse promoter. The minimal promoter contained TATA and CAAT motifs and consensus sequences binding to SP1 (three sites) and AP2 (one site). Deletion analyses of the promoter revealed that a 24-bp region near 5'-end of the minimal promoter was essential for the efficient transcription, although the AP2 site in the region was dispensable. A fluorescence polarization assay suggested that the plus strand of the 24-bp region, despite the lack of known consensus sequences binding to transcription factors except for AP2, bound to unknown nuclear protein(s) in a sequence specific manner.