To investigate the ability of the retinoblastoma gene product (RB protein, pRB) to regulate its own expression, cotransfection assays using human RB promoter-luciferase fusion plasmids and a human pRB expression plasmid were employed. In B104, a rat neuroblastoma cell line, pRB stimulated luciferase activity about 2-fold from the wild-type promoter, and about 4-fold from a mutant promoter with a mutation in the retinoblastoma binding factor 1 (RBF-1) site. The RB-responsive region was mapped to a novel 44 bp sequence in the 5' untranslated region in both wild-type and mutant promoters. When apparent stimulation of luciferase activity by pRB was observed, the luciferase mRNA level did not increase, suggesting that through this 44 bp region, pRB could post-transcriptionally regulate its own expression.