A Saccharomyces cerevisiae protein fraction which binds specifically to the internal promoter regions of genes that are transcribed by RNA polymerase III is shown to function as a transcription factor. We postulate that the stable DNA binding of the factor confers stability on polymerase III transcription complexes. Analysis of the binding by DNase 'foot-printing' distinguishes three segments of the S. cerevisiae tRNALeu3 gene: a region surrounding the so-called A block of the internal promoter, a region surrounding the B block and an intermediate segment. Binding to the A and B block regions is connected, but the B block region exerts a dominant effect.