The promoter specificity of transcriptional activators is generally thought to be conferred by the specificity of the DNA-binding domain, which brings the activation domain to the appropriate promoter sequence. We show here, however, that Oct-1 and Oct-2 can differentially activate transcription not through DNA binding specificity but instead through the use of promoter-selective activation domains. These distinct activation domains lead to stimulation of the U2 small nuclear RNA promoter by Oct-1 and an mRNA promoter by Oct-2. An Oct-2 variant, called Oct-2B, differs from Oct-2 by an Oct-1-related C-terminal extension that results from alternative splicing. This variant gains the ability to activate the U2 small nuclear RNA promoter. Thus, the promoter selectivity of a transcriptional activator can be changed, in this case by alternative splicing, without affecting its DNA binding specificity.