During pre-mRNA splicing, a central step in the expression and regulation of eukaryotic genes, the spliceosome selects splice sites for intron excision and exon ligation. In doing so, the spliceosome must distinguish optimal from suboptimal splice sites. At the catalytic stage of splicing, suboptimal splice sites are repressed by the DEAH-box ATPases Prp16 and Prp22. Here, using budding yeast, we show that these ATPases function further by enabling the spliceosome to search for and utilize alternative branch sites and 3′ splice sites. The ATPases facilitate this search by remodeling the splicing substrate to disengage candidate splice sites. Our data support a mechanism involving 3′ to 5′ translocation of the ATPases along substrate RNA and toward a candidate site, but, surprisingly, not across the site. Thus, our data implicate DEAH-box ATPases in acting at a distance by pulling substrate RNA from the catalytic core of the spliceosome. Display omitted •The DEAH-box ATPase Prp16 enables alternative branch site selection•The DEAH-box ATPase Prp22 promotes alternative 3′ splice site selection•Prp16 and Prp22 both repress suboptimal sites by disengaging the splice sites•Prp16 and Prp22 remodel the substrate without translocating through their targets Two DEAH-box ATPases enable the spliceosome to search for and utilize alternative branch sites and 3′ splice sites by disengaging suboptimal sites from the splicing machinery through a mechanism that suggests action at a distance by RNA pulling.