Biology subjects

Martin Depken

Publications and source records attributed to Martin Depken.

2 recordsLinked to original sources

bioRxiv2016

Real-time observation of replicative helicase assembly onto single-stranded DNA

Replicative helicases load onto DNA at the start of replication, and play a vital role by driving the replication fork forward. These helicases assemble into closed multimeric rings that need to encircle single-stranded (ss)DNA to be activated. Though helicase loading on substrates with accessible free ends has been well characterized for the T7 gp4 helicase, a model system for superfamily IV replicative helicases, the physiologically more relevant loading onto exposed ssDNA without free ends remains less well understood. Here, using a label-free assay that exploits changes in the DNA hairpin hopping dynamics to detect gp4 binding and activity, we characterize loading and activation of gp4 on exposed ssDNA without free ends, and find clear evidence of stepwise assembly of the helicase at the fork at physiologically relevant concentrations. The gradual loading onto ssDNA, rather than pre-forming in solution followed by spontaneous ring opening which appears favored at higher concentrations, suggests a new paradigm of stepwise assembly for the helicases in superfamily IV that do not require a separate loading enzyme.

Biophysics↗

bioRxiv2016

Crowding-induced transcriptional bursts dictate nucleosome and polymerase density profiles along genes

During transcription, RNA polymerase competes for space on the DNA with other DNA binding proteins and higher order DNA structures acting as roadblocks. Though it is known that individual polymerases often slow down when forcing roadblocks, the effect of crowding on transcription as a whole is not clear. Based on quantitative theoretical modeling, we show that interactions with roadblocks induce a strong kinetic attraction between polymerases, causing them to self-organize into pelotons. Peloton formation explains observed nucleosome and polymerase density profiles close to the initiation site on highly transcribed genes, and how these densities depend on induction levels. At termination, pelotons translate into transcriptional bursts that dispaly the same characteristics as those observed in vivo. Our model thus unifies common spatial and temporal transcription patterns as arising from a non-specific interaction between roadblocks and polymerases. The generality of our model suggests that peloton formation might be ubiquitous in systems where molecular motors interact with dynamic roadblocks.

Biophysics↗