Many elongation factors in eukaryotes promote gene expression by increasing the processivity of RNA polymerase II (Pol II). However, the stability of RNA Pol II elongation complexes suggests that such complexes are not inherently prone to prematurely terminating transcription, particularly at physiological nucleotide concentrations. We show that the termination factor, Pcf11, causes premature termination on an HIV provirus. The transcription that occurs when Pcf11 is depleted from cells or an extract is no longer sensitive to 6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB), a compound that causes premature termination. Hence, Pcf11 can act as a negative elongation factor to repress RNA Pol II gene expression in eukaryotic cells.