RESUMO
Transcription is a dynamic process. To detect the dynamic relationship among protein clusters of RNA polymerase II and coactivators, gene loci, and transcriptional activity, we insert an MS2 repeat, a TetO repeat, and inteins with a selection marker just downstream of the transcription start site. By optimizing the individual elements, we develop the Spliced TetO REpeAt, MS2 repeat, and INtein sandwiched reporter Gene tag (STREAMING-tag) system. Clusters of RNA polymerase II and BRD4 are observed proximal to the transcription start site of Nanog when the gene is transcribed in mouse embryonic stem cells. In contrast, clusters of MED19 and MED22 tend to be located near the transcription start site, even without transcription activity. Thus, the STREAMING-tag system reveals the spatiotemporal relationships between transcriptional activity and protein clusters near the gene. This powerful tool is useful for quantitatively understanding transcriptional regulation in living cells.
