A live imaging system to analyze spatiotemporal dynamics of RNA polymerase II modification in Arabidopsis thaliana

Mio K. Shibuta, Takuya Sakamoto, Tamako Yamaoka, Mayu Yoshikawa, Shusuke Kasamatsu, Noriyoshi Yagi, Satoru Fujimoto, Takamasa Suzuki, Satoshi Uchino, Yuko Sato, Hiroshi Kimura, Sachihiro Matsunaga

Spatiotemporal changes in general transcription levels play a vital role in the dynamic regulation of various critical activities. Phosphorylation levels at Ser2 in heptad repeats within the C-terminal domain of RNA polymerase II, representing the elongation form, is an indicator of transcription. However, rapid transcriptional changes during tissue development and cellular phenomena are difficult to capture in living organisms. We introduced a genetically encoded system termed modification-specific intracellular antibody (mintbody) into Arabidopsis thaliana. We developed a protein processing- and 2A peptide-mediated two-component system for real-time quantitative measurement of endogenous modification level. This system enables quantitative tracking of the spatiotemporal dynamics of transcription. Using this method, we observed that the transcription level varies among tissues in the root and changes dynamically during the mitotic phase. The approach is effective for achieving live visualization of the transcription level in a single cell and facilitates an improved understanding of spatiotemporal transcription dynamics.