Polymerase Recycling and Transcriptional Noise

Transcription is a highly fluctuating process subject to stochastic variation, which produces great variations in the numbers of mRNAs that are found in individual, otherwise identical cells. This transcriptional cell-to-cell variability, often referred to as transcriptional noise, is incompletely understood, in particular in mammalian systems. This variability is implicated in many clinically relevant settings such as stem cell differentiation, immune regulation, or cancer formation. Transcription by RNA polymerase II (PolII) is a complex process that integrates mechanisms involved in initiation, RNA processing, and termination . In particular, the C-terminal domain (CTD) of the main polymerase subunit Rpb1 is of importance for local clustering of polymerases and undergoes different types of phosphorylation in a defined sequence throughout the transcriptional cycle. Dephosphorylation of the CTD upon termination is a prerequisite for reinitiation and thus recycling of the polymerase.
Recycling of PolII should permit it to repeatedly start transcribing again after termination, potentially producing a rapid succession of new mRNAs. Our hypothesis is that this polymerase recycling is a significant contributor to transcriptional bursting, which is strongly supported by pilot experiments we conducted. We will conduct genome-wide experimental analysis of PolII recycling and stochastic transcription. To achieve this, we will be doing ChIP-seq, and single-cell RNA-seq. Finally, data gained will be processed and analysed using standard bioinformatics approaches. Mathematical model fitting will also be used to determine transcriptional parameters including the frequency in dependency of polymerase recycling, based on master equations.