Microbial genome defence pathways: from molecular mechanisms to next-generation molecular tools
The constant arms race between prokaryotic microbes and their molecular parasites such as viruses, plasmids and transposons has driven the evolution of complex genome defence mechanisms. The CRISPR-Cas defence systems provide adaptive RNA-guided immunity against invasive nucleic acid elements. CRISPR-associated effector nucleases such as Cas9, Cas12a and Cas13 have emerged as powerful tools for precision genome editing, gene expression control and nucleic acid detection. However, these technologies suffer from drawbacks that limit their efficacy and versatility, necessitating the search for additional exploitable molecular activities. Building on our recent structural and biochemical studies, the goal of this project is to investigate the molecular architectures and mechanisms of CRISPR-associated systems and other genome defence mechanisms, aiming not only to shed light on their biological roles but also inform their technological development. Specifically, the proposed studies will examine (i) the molecular basis of cyclic oligoadenylate signalling in type III CRISPR-Cas systems, (ii) the mechanism of transposon-associated type I CRISPR-Cas systems and their putative function in RNA-guided DNA transposition, and (iii) molecular activities associated with recently described non-CRISPR defence systems. Collectively, the proposed studies will advance our understanding of the molecular functions of genome defence mechanisms in shaping the evolution of prokaryotic genomes and make critical contributions to their development as novel genetic engineering tools.