CRISPR, transposons and the role of RNAs as genomic targeting agents
Israel Fernández
Biofisika Institute (CSIC, EHU), Leioa, Bizkaia
CRISPR systems evolved in Bacteria and Archaea as defense mechanisms against Mobile Genetic Elements (MGE) such as viruses. The diversity of CRISPR systems as well as their prevalence among prokaryotic species suggest a scenario of rampant conflict between cells and MGE at the microbiology domain. With conflicts of such wingspan, arms-races at the molecular level are common, with shuffling of genes and/or gene exaptation events providing defense/attack systems with novel strategies to overcome adversarial situations. Using cryoEM, we studied in detail a bacterial transposon that exapted the DNA binding machinery of a CRISPR system for precise DNA location, binding and mobilization. Driven by accurate base pairing specified by the RNA component of the CRISPR system, these transposons are able to mobilize large DNA cargo molecules ranging in size from 0.5 to 5 kilo-bases to distant locations in bacterial genomes. We will focus our discussion on technical aspects of cryoEM, describing the problems we faced in the characterization of this system in order to attain high-resolution regimens. Additionally, we will discuss two new problematic samples for cryoEM we are working with that exemplify our approach to hard cryoEM targets.