CRISPR/Cas9 is well known for its use in genome editing, but it was discovered in the adaptive immune system of bacteria, where it protects them from viruses or other pathogens by cutting foreign DNA at specific sites. However, it was unknown how the system is regulated in its natural environment. New research from Johns Hopkins School of Medicine in Maryland suggests the existence of a 'genetic brake'.
'From an immunity perspective, bacteria need to ramp up CRISPR/Cas9 activity to identify and rid the cell of threats, but they also need to dial it down to avoid autoimmunity – when the immune system mistakenly attacks components of the bacteria themselves,' explained first author Rachael Workman.
The research, published in Cell, found that when a specific gene was deactivated in the bacterium Streptococcus pyogenes, the CRISPR system became more active. The gene codes for a long-form transactivating CRISPR RNA (tracrRNA) the function of which was previously unknown. The tracrRNA – whose short-form usually guides the Cas9 enzyme to cut DNA – acts as a molecular switch to dampen gene activity.
In a further experiment, the long tracrRNA was re-programmed to recognise and bind to a gene that produces green fluorescence. Bacteria with the altered version of the tracrRNA glowed less green than bacteria with the normal long tracrRNA, suggesting that the molecule can be manipulated to dial down other genes.
'The dimmer capability that the experiments uncovered offers opportunities to design new or better CRISPR/Cas9 tools aimed at regulating gene activity for research purposes,' said lead author Dr Joshua Modell.
The team hope to discover whether the same mechanism applies in other bacterial strains or if different control mechanisms exist.