To investigate mechanisms that control fruit development, we have begun experiments to identify proteins that control gene expression during tomato fruit ripening. We focused on the regulation of two different genes, E4 and E8, whose transcription is coordinately activated at the onset of fruit ripening. We report here that a DNA-binding protein specifically reacts with similar sequences flanking the E4 and E8 genes. The E4 binding site is at position -34 to -18 and, therefore, overlaps the region (TATA box) that in many eukaryotic genes serves to determine the efficiency and initiation site of transcription. In contrast, the E8 binding site is distal, located at -936 to -920 relative to the start of E8 gene transcription. Gel electrophoresis mobility retardation experiments indicate that the DNA binding activity that interacts with these two sites increases at the onset of fruit ripening. Taken together, these results suggest that this DNA-binding protein may function to coordinate E4 and E8 gene expression during fruit ripening.