In the study, the authors show that DSIF consists of two loops, which wrap around the DNA and RNA strand respectively, and a stabilizing domain (Nature Structural and Molecular Biology doi: 10.1038/nsmb.3465). The protein keeps all molecules involved in transcribing genes into RNA in place: it positions incoming DNA and outgoing RNA, and recruits factors for RNA processing. By physically contacting all key players, including the polymerase itself, it exerts its influence. The positioning of nucleic acids is done by forming clamps, like rings, around the DNA and RNA.

Beyond the resolution of light

To capture images of this process, Bernecky and her colleagues resorted to cryo-electron microscopy and X-ray crystallography. Instead of light, these techniques use electrons and X-rays, respectively, allowing for incredibly high resolution. The much smaller wavelengths of electrons and X-rays overcome the resolution limit that can be achieved with light.

The experiment highlighted DSIF's interaction with the outgoing RNA that keeps this RNA in the exit channel of Pol II. If this was not the case, the produced RNA would compete with the non-template DNA strand for the template DNA strand and destabilize the interaction between RNA, DNA and polymerase. As a result, transcription would stop.

DSIF pauses and un-pauses Pol II

Another noteworthy feature is the involvement of DSIF in pausing Pol II and releasing it from pause positions. The pausing regulates the rate of transcription and ensures that genes are expressed synchronously, for example to form new tissues during the development of embryos. This can be achieved by un-pausing Pol II simultaneously in many neighbouring cells. Whether DSIF initiates pausing or un-pausing depends on its phosphorylation state.

The DNA clamp, as observed by Bernecky and her co-workers, is important for maintaining stability of the elongation complex. Indeed, without it, Pol II dissociates from the template and transcription pauses. The RNA clamp of DSIF contributes more actively to the regulation of pausing, mediated by phosphorylation. This makes this RNA clamp a major developmental regulator.