The paradoxical role of ATP in viral infections and cellular defense

Abstract

Conflict between prokaryotes and viruses has driven the evolution of diverse defense systems. The study of these systems has not only expanded our knowledge of the mechanisms underlying genetic conflict, but has also revolutionized biotechnologies. I joined the Wiedenheft lab in early 2021 during the height of the SARS-CoV-2 pandemic. As such, the lab was focused on developing technologies to aid in the identification and study of SARS-CoV-2. I joined a team that was actively repurposing a type III CRISPR system as a molecular diagnostic. Type III CRISPRs are RNA targeting systems that unleash antiviral signaling cascades upon recognition of a target. Previous work in the lab had shown that this activity could be repurposed as a molecular diagnostic, but the first generation technology was not sensitive enough for clinical use. To address this need, we developed a technology termed Capture and Concentrate, which utilized 'CRISPR magnets' to concentrate target RNAs from complex mixtures. Using this technology, we increased diagnostic sensitivity by 1,000 fold. In addition to my work at the bench, I developed an in silico model of the diagnostic to inform future development efforts. While type III CRISPRs are unique due to their ability to generate antiviral signaling cascades, they also contain intrinsic endonuclease activity. In 2023 we demonstrated that this activity could be repurposed as an RNA editing technology. Using the type III system from Streptococcus thermophilus, we introduced precise genomic modifications in a recombinant sindbis virus. This technology removed the need for reverse genetics systems, which hampered the study of the rapidly evolving SARS-CoV-2 virus. In the final year of my graduate studies, I worked with a team focused on determining how, PARIS, a bacterial innate immune system, induced host death upon viral infection. PARIS is a two-component system comprised of the ATPase, AriA, and a nuclease, AriB. Previous studies had shown that PARIS induced abortive infection in response to viral proteins, but the mechanism was not known. We determined that upon antigen recognition, AriB is activated and cleaves host tRNAs to induce cell death and prevent community spread.