Alternative transcription cycle for bacterial RNA polymerase

Abstract

RNA polymerases (RNAPs) transcribe genes through a cycle of recruitment to promoter DNA, initiation, elongation, and termination. After termination, RNAP is thought to initiate the next round of transcription by detaching from DNA and rebinding a new promoter. We used single-molecule fluorescence microscopy to observe individual RNAP molecules after 25 transcript release at a terminator. Following termination, RNAP almost always remained bound to DNA and sometimes exhibited one-dimensional sliding over thousands of basepairs. Unexpectedly, the DNA-bound RNAP often restarted transcription, usually in reverse direction, thus producing an antisense transcript. Furthermore, we report evidence of this "secondary initiation" in live cells, using genome-wide RNA sequencing. These findings reveal an 30 alternative transcription cycle that allows RNAP to reinitiate without dissociating from DNA, which is likely to have important implications for gene regulation.

Defining kinetic roles of transcriptional activators in the early Drosophila embryo

“…The data from two imaging replicates for each transgenic reporter were combined and treated as a single dataset, as is typical for analysis of fluorescence spectroscopy experiments (e.g. (Harden et al, 2016) ).…”

“…There, biochemical and structural approaches have elucidated detailed chemical and physical mechanisms for many individual TFs (e.g. the sigma factors (Bae et al, 2015;Chen et al, 2020;Friedman and Gelles, 2012;Harden et al, 2016) ). Within animal transcription, research has largely focused on a tissue-specific paradigm of TF function that identifies TFs responsible for developmental patterning and cell type specification and characterizes them as activators or repressors (Guo et al, 2018;Lambert et al, 2018;Villar et al, 2014) .…”