David Bikard

– Abstract –

CRISPR tools to study and control bacteria

Over the past few years CRISPR systems have been derived into powerful tools to edit genomes, control gene expression, visualize nucleic acids in vivo, modify epigenetic marks, kill cells and more. Most of these tools rely on the RNA-guided nuclease known as Cas9 which can be easily reprogrammed to bind and cut almost any position in a genome. We focus on the application of these technologies to Bacteria. We have recently investigated the properties of Cas9 as well as the catalytic dead variant known as dCas9 in E. coli. The introduction of double strand breaks by Cas9 at a specific genomic position leads to cell death as such breaks cannot be repaired through homologous recombination, the main DNA repair pathway in bacteria. We used this property as a selection tool in both E. coli and S. pneumoniae, and to develop sequence-specific antimicrobials against S. aureus. Work on the dCas9 protein has revealed how it can be used to precisely tune the expression level of several target genes independently and with low noise levels. Finally, the ability to knockdown gene expression with dCas9 can also be used in high-throughput screens to unravel gene function and interaction. All in all, CRISPR technology provides a fantastic toolkit to study and fight pathogenic bacteria.