The complexity of eukaryotic life forms is driven by RNA polymerase II (Pol II), which transcribes all protein-coding genes and thousands of non-coding RNA loci. In vivo, Pol II cannot transcribe genes alone – it needs the help of various regulatory factors to initiate transcription, pause, be released from pausing and then elongate though a complex chromatin environment that may be crowded with obstacles, until finally reaching gene end and terminating transcription.

Around 30 “core” Pol II regulatory factors are known – they bind Pol II directly at different stages of transcription and are thought to be necessary for the transcription process. Depletion of many core Pol II-regulatory factors is not lethal in cell lines or even in animals, despite the critical tasks they perform on genes when analysed in vitro. This shows that multiple mechanisms must exist, and compensate for each other in cells. To understand the relationships between different Pol II regulators, and to discover new regulatory factors, the PhD candidate will perform synthetic lethal CRISPR knock-out/knock-down screens. The function and mechanism of identified hits will be investigated further using a combination of genomic, biochemical and structural approaches.

References

Tufegdzic Vidakovic, A., Mitter, R., Kelly, G.P., Neumann, M., Harreman, M., Rodriguez-Martinez, M., Herlihy, A., Weems, J.C., Boeing, S., Encheva, V., et al. (2020)
Regulation of the RNAPII Pool Is Integral to the DNA Damage Response.
Cell 180: 1245-1261 e1221.

Tufegdzic Vidakovic, A., Harreman, M., Dirac-Svejstrup, A.B., Boeing, S., Roy, A., Encheva, V., Neumann, M., Wilson, M., Snijders, A.P., and Svejstrup, J.Q. (2019)
Analysis of RNA polymerase II ubiquitylation and proteasomal degradation.
Methods 159-160: 146-156.