Strong evidence indicates that transcription elongation by RNA polymerase II (pol II) is a highly regulated process. Here we present genetic results that indicate a role for the Saccharomyces cerevisiae Rtf1 protein in transcription elongation. A screen for synthetic lethal mutations was carried out with an rtf1 deletion mutation to identify factors that interact with Rtf1 or regulate the same process as Rtf1. The screen uncovered mutations in SRB5, CTK1, FCP1, and POB3. These genes encode an Srb/mediator component, a CTD kinase, a CTD phosphatase, and a protein involved in the regulation of transcription by chromatin structure, respectively. All of these gene products have been directly or indirectly implicated in transcription elongation, indicating that Rtf1 may also regulate this process. In support of this view, we show that RTF1 functionally interacts with genes that encode known elongation factors, including SPT4, SPT5, SPT16, and PPR2. We also show that a deletion of RTF1 causes sensitivity to 6-azauracil and mycophenolic acid, phenotypes correlated with a transcription elongation defect. Collectively, our results suggest that Rtf1 may function as a novel transcription elongation factor in yeast.