Molecular recycling is essential to gene expression.  In all organisms, after each RNA molecule is made, the RNA polymerase (RNAP) enzyme that made it must be reset and repositioned to make subsequent RNAs.  The steps involved in this RNAP recycling process are fully understood in any organism, not even in simple bacteria.

Ph.D. student Koe Inlow and collaborators studied the RapA enzyme, one of the only bacterial homologs of the large Swi2/Snf2 family of eukaryotic chromatin remodelers.  RapA has long been known as an abundant RNAP binding protein.  It has been proposed to function in RNAP recycling (and also in multiple other roles), but how it does so is unclear. Using multi-color single-molecule fluorescence microscopy, she made unprecedented direct observations in vitro of the dynamics of individual molecules of fluorescently labeled RNAP and RapA as they interacted with each other and with template DNA during and following transcript synthesis.  These studies show for the first time that RapA acts on a key intermediate in the transcription cycle: the recently discovered post-termination complex (PTC) in which RNAP slides along DNA and from which it can reinitiate transcription on nearby genes either sense or antisense relative to the prior round of transcript synthesis.  RapA thus competes against local reinitiation by RNAP sliding and is likely to promote replenishment of the global pool of free RNAP holoenzyme.  Further, the studies reveal that even tiny (nanomolar) concentrations of RapA efficiently use ATP hydrolysis to disassemble the PTC and uncover the essential features of the mechanism by which this removal occurs. These studies fill in the essential missing pieces in the current understanding of the events that occur after RNA is released and that enable RNAP recycling. Furthermore, we rationalize RapA genetics by explaining how RapA facilitates global transcriptional reprogramming as cells enter and leave stress conditions.