Tag Archives: NIGMS-funded

Alternative transcription cycle for bacterial RNA polymerase

DNA transcription is the most important nexus of gene regulation in all living organisms.  In recent years, single-molecule experiments have given us a new window into the mechanisms of transcription and have revealed novel, previously unsuspected molecular behaviors and mechanisms.  Ph.D. student Tim Harden used multi-color single-molecule fluorescence imaging to reveal a completely new transcription cycle for bacterial RNA polymerase.  Harden and co-authors showed that an RNA polymerase molecule in vitro frequently (in >90% of transcription events) remains bound to DNA and may again initiate transcription after it has terminated the first round of transcription.  Even more unexpectedly, this “secondary initiation” is not restricted to the same RNA.  After the first round, the polymerase can scan thousands of basepairs along the DNA and can initiate at a different start site, frequently one that is oriented in the opposite direction and produces an antisense transcript.

cannonical and alternative bacterial transcription cycles

To complement the single-molecule studies in vitro, the manuscript reports new analyses of whole-transcriptome cellular RNAs revealed by the Rend-seq method that measures transcript initiation and termination frequencies across the genome with single basepair resolution.  These provide evidence that the new transcription cycle may be responsible for initiating antisense transcription at hundreds of genomic locations in the two widely divergent bacterial species examined.  The work defines a new mechanism for the regulated production of antisense RNAs, many of which are now recognized as important agents of gene-specific regulation through control of transcription, mRNA decay, and translation.  In addition, the new transcription cycle provides a mechanism through which transcription initiation can be controlled not just through feedback networks involving multiple genes, but also through production of multiple different primary transcripts consequent to a single RNA polymerase-to-DNA recruitment event.

10.1038/s41467-019-14208-9
Harden, T.T., et al. Alternative transcription cycle for bacterial RNA polymerase.
Nature Communications 11, 450 (2020).

Synergy between cyclase-associated protein and cofilin accelerates actin filament depolymerization by two orders of magnitude

From Science at Brandeis:

“All animal and plant cells contain a highly elaborate system of filamentous protein polymers called the actin cytoskeleton, a scaffold that can be rapidly transformed to alter a cell’s shape and function. A critical step in reconfiguring this scaffold is the rapid disassembly (or turnover) of the actin filaments. But how is this achieved? It has long been known that the protein Cofilin plays a central role in this process, but it has been unclear how Cofilin achieves this feat. Cofilin can sever actin filaments into smaller fragments to promote their disassembly, but whether it also catalyzes subunit dissociation from filament ends has remained uncertain and controversial. Until now, this problem has been difficult to address because of limitations in directly observing Cofilin’s biochemical effects at filament ends….”  Dr. Shashank Shekhar, working together with Dr. Johnson Chung and “jointly mentored by Bruce Goode, Jeff Gelles and Jane Kondev, use[d] microfluidics-assisted single molecule TIRF imaging to tackle the problem.

The new study shows that Cofilin and one other protein (Srv2/CAP) intimately collaborate at one end of the actin filament to accelerate subunit dissociation by over 300-fold! These are the fastest rates of actin depolymerization ever observed. Further, these results establish a new paradigm in which a protein that decorates filament sides (Cofilin) works in concert with a protein that binds to filament ends (Srv2/CAP) to produce an activity that is orders of magnitude stronger than the that of either protein alone.”

10.1038/s41467-019-13268-1
Shekhar S. et al. Synergy between cyclase-associated protein and cofilin accelerates actin filament depolymerization by two orders of magnitude.
Nature Communications 10, 5319 (2019).