Bulky DNA lesions not only block the progress of RNAPII locally, but also affect transcription genome-wide so that even genes that are not damaged temporarily cease to be expressed.
The mechanisms and factors that underlie the more general, DNA damaged-induced repression of gene expression, and its eventual re-start, are still very poorly understood.
In order to uncover factors with a role in the transcription-related DNA damage response, we carried out several general and more specific proteomic screens, as well as a functional genomics screen.
The proteomic screens all made use of quantitative SILAC proteomics, enabling us to distinguish between ‘constitutive’ and UV-induced interactors of RNAPII and CSB, respectively. As well as the well-known TC-NER factors, this multi-omic approach has uncovered numerous new factors that had not previously been connected to the transcription-related DNA damage response, many of which we are presently investigating in detail.
Discovery of new factors via the multi-omic approach is complemented by genome-wide deep sequencing techniques such as ChIP-Seq, RNA-Seq, CLIP-Seq, 4SU-Seq and DRB/GRO-Seq to investigate the role of these new factors in the transcriptional response to UV-induced DNA damage at the genome-wide level.
Because UV-damage has turned out to trigger a dramatic change in mRNA splicing, termination, and RNA biology in general, much of our present focus has shifted more and towards RNA biology, including the function of stable RNAs.