A transcription dynamics imaging system in Arabidopsis thaliana

Abstract

Spatiotemporal regulation of transcription plays important roles in the dynamic regulation of various biological activities. However, rapid transcriptional changes are difficult to observe in living cells. Our group established a new system for imaging transcription activity, focusing on the RNA polymerase II (RNAPII). RNAPII is a large multiprotein complex that transcribes DNA into precursors of messenger RNA. The C-terminal domain of RNAPII contains highly conserved heptad repeats that undergo several post-translational modifications during the transcription cycle. Phosphorylation of the second Ser residue (Ser2P) in the repeat signals transcription elongation and termination, making Ser2P an important indicator of the transcription. To monitor Ser2P levels in living cells, a genetically encoded system termed modification-specific intracellular antibody (mintbody) is introduced into Arabidopsis thaliana. Immunostaining and ChIP-seq analysis confirm the function of the mintbody as an intracellular antibody for Ser2P (Ser2P-mintbody). Ser2P-mintbody can change its localization in response to rapid changes in transcriptional activities under the conditions of transcription inhibitor treatment. For quantitative measurement of endogenous Ser2P levels, a two-component system is being developed. This system allows quantitative tracking of transcriptional dynamics, such as the dynamic change of transcription level during the mitotic phase. In addition, Ser2P-mintbody is useful for exploring tissue- or cell-type-specific variation of transcription activity. The observation of Ser2P-mintbody in pollen leads to the discovery of the interesting distribution pattern of the transcription active region in sperm nuclei. The approach is effective for achieving live visualization of the transcription level and facilitates a better understanding of cellular phenomena and tissue development.