DSIF contributes to transcriptional activation by DNA-binding activators by preventing pausing during transcription elongation.
The transcription elongation factor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor (DSIF) regulates RNA polymerase II (RNAPII) processivity by promoting, in concert with negative elongation factor (NELF), promoter-proximal pausing of RNAPII. DSIF is also reportedly involved in transcriptional activation. However, the role of DSIF in transcriptional activation by DNA-binding activators is unclear. Here we show that DSIF acts cooperatively with a DNA-binding activator, Gal4-VP16, to promote transcriptional activation. In the absence of DSIF, Gal4-VP16-activated transcription resulted in frequent pausing of RNAPII during elongation in vitro. The presence of DSIF reduced pausing, thereby supporting Gal4-VP16-mediated activation. We found that DSIF exerts its positive effects within a short time-frame from initiation to elongation, and that NELF does not affect the positive regulatory function of DSIF. Knockdown of the gene encoding the large subunit of DSIF, human Spt5 (hSpt5), in HeLa cells reduced Gal4-VP16-mediated activation of a reporter gene, but had no effect on expression in the absence of activator. Together, these results provide evidence that higher-level transcription has a stronger requirement for DSIF, and that DSIF contributes to efficient transcriptional activation by preventing RNAPII pausing during transcription elongation.
Zhu
Wenyan
W
Graduate School of Bioscience and Biotechnology and Integrated Research Institute, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
Wada
Tadashi
T
Okabe
Sachiko
S
Taneda
Takuya
T
Yamaguchi
Yuki
Y
Handa
Hiroshi
H
D004268:DNA-Binding Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D009687:Nuclear Proteins / Q000502:physiology*; D012319:RNA Polymerase II / Q000378:metabolism; D015534:Trans-Activators / Q000502:physiology*; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D015533:Transcriptional Activation; D035602:Transcriptional Elongation Factors
D004268:DNA-Binding Proteins; C070633:Gal-VP16; D009687:Nuclear Proteins; C112224:SUPT5H protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; C118437:negative elongation factor; D012319:RNA Polymerase II