Molecular Microbiology seminars

Virginijus Šikšnys traces genome-editing tools to bacterial antiviral defence. Programmable CRISPR-Cas nucleases offer powerful ways to alter DNA, but delivery, unintended effects, immune interactions and insertion efficiency remain challenges. The lecture discusses compact TnpB nucleases and how further study of defence systems may reveal useful enzymatic functions.

Macromolecular protein complexes perform essential biological functions across life forms. A fundamental, though yet unsolved question in biology is how the function of such complexes is regulated by intracellular or extracellular signals. For instance, we have little understanding of how forces affect multi-protein machines whose function is often mechanical in nature. We address this question by studying the bacterial flagellar motor, a large complex that powers swimming motility in many bacteria. This rotary motor autonomously adapts to changes in mechanical load by adding or removing force-generating ‘stator’ units that power rotation. In the bacterium Escherichia coli, up to 11 units drive the motor at high load while all the units are released at low load. We manipulate motor load using electrorotation, a technique in which a rapidly rotating electric field applies an external torque on the motor. This allows us to change motor load at will and measure the resulting stator dynamics at single-unit resolution. We found that the force generated by the stator units controls their unbinding, forming a feedback loop that leads to autoregulation of the assembly. We complemented our experiments with theoretical models that provide insight into the underlying molecular interactions. Torque-dependent remodeling takes place within seconds, making it a highly responsive control mechanism, one that is mediated by the mechano-chemical tuning of protein interactions.

Emmanuelle Charpentier and Jennifer Doudna explain the discovery and biochemical development of CRISPR-Cas9 as a programmable tool for modifying DNA. The scientific story begins with bacterial defence against invading genetic material and the role of tracrRNA in the CRISPR system. It proceeds through the reconstruction and simplification of the RNA-guided machinery and the demonstration that a chosen guide can direct DNA cleavage to a specified sequence. The lectures connect basic research on microbes and RNA with a method that transformed experimental genetics. They explain the molecular logic of the system and the possibilities it opened for studying gene function, engineering organisms and developing therapeutic approaches.